PHYSIOLOGICAL REVIEWS

Vol. 81, No. 1, January 2001

Printed in U.S.A.

Small GTP-Binding Proteins

YOSHIMI TAKAI, TAKUYA SASAKI, AND TAKASHI MATOZAKI
Department of Molecular Biology and Biochemistry, Osaka University Graduate School
of Medicine/Faculty of Medicine, Suita, Japan

I. Introduction
II. General Properties
A. Structure
B. A role as molecular switches
C. Localization
III. Ras Proteins as Regulators of Gene Expression
A. Outline
B. Ras protein cycle: activation/inactivation
C. Raf protein kinase activation by Ras proteins
D. Modifiers of the Ras protein-induced Raf protein kinase activation
E. Other effectors of Ras proteins
F. Transport of newly synthesized Ras proteins from the endoplasmic reticulum
to the plasma membrane
G. Ras proteins and cancer
H. Rap proteins
I. Ral proteins
J. Other Ras family members
IV. Rho/Rac/Cdc42 Proteins as Regulators of Both Cytoskeletal Reorganization and Gene Expression
A. Outline
B. Reorganization of the actin cytoskeleton
C. Rho/Rac/Cdc42 protein cycle: cyclical activation/inactivation
D. Mode of action of Rho proteins in cytoskeletal reorganization
E. Mode of action of Rac/Cdc42 proteins in cytoskeletal reorganization
F. Mode of action of Rho/Rac/Cdc42 proteins in gene expression
G. Other functions of Rho/Rac/Cdc42 proteins
V. Rab Proteins as Regulators of Vesicle Trafficking
A. Outline
B. Vesicle trafficking
C. Localization of Rab proteins
D. Rab protein cycle: cyclical activation/inactivation and translocation
E. SNAREs and tethering proteins in vesicle targeting/docking/fusion
F. Mode of action of Rab proteins in vesicle targeting/docking/fusion
G. Rab3A in Ca2-dependent exocytosis
H. Rab proteins and cytoskeleton
I. Rab proteins in vesicle budding
VI. Sar1/Arf Proteins as Regulators of Vesicle Budding
A. Outline
B. Coat proteins and vesicle budding
C. Arf protein cycle: cyclical activation/inactivation and translocation
D. Arf proteins in vesicle budding
E. Arf6 in endocytic recycling and cytoskeletal reorganization
F. Sar1 as a regulator of vesicle budding
VII. Ran Function in Nucleocytoplasmic Transport and Microtubule Organization
A. Outline
B. Nucleocytoplasmic transport
C. Ran cycle: cyclical activation/inactivation and translocation
D. Mode of action of Ran in nucleocytoplasmic transport
E. A role for Ran in microtubule organization
http://physrev.physiology.org

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VIII. Small G Protein Cascades and Cross-talks

A. Small G protein cascades
B. Cross-talk between small G proteins
IX. Conclusions and Perspectives
A. Roles in two types of cell regulation
B. A role as biotimers rather than as molecular switches
C. A role as spatial determinants

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Takai, Yoshimi, Takuya Sasaki, and Takashi Matozaki. Small GTP-Binding Proteins. Physiol Rev 81: 153208,
2001.Small GTP-binding proteins (G proteins) exist in eukaryotes from yeast to human and constitute a superfamily consisting of more than 100 members. This superfamily is structurally classified into at least five families: the
Ras, Rho, Rab, Sar1/Arf, and Ran families. They regulate a wide variety of cell functions as biological timers
(biotimers) that initiate and terminate specific cell functions and determine the periods of time for the continuation
of the specific cell functions. They furthermore play key roles in not only temporal but also spatial determination of
specific cell functions. The Ras family regulates gene expression, the Rho family regulates cytoskeletal reorganization and gene expression, the Rab and Sar1/Arf families regulate vesicle trafficking, and the Ran family regulates
nucleocytoplasmic transport and microtubule organization. Many upstream regulators and downstream effectors of
small G proteins have been isolated, and their modes of activation and action have gradually been elucidated.
Cascades and cross-talks of small G proteins have also been clarified. In this review, functions of small G proteins
and their modes of activation and action are described.

I. INTRODUCTION
Small GTP-binding proteins (G proteins) are monomeric G proteins with molecular masses of 20 40 kDa.
The Ha-Ras and Ki-Ras genes were first discovered as the
v-Ha-Ras and v-Ki-Ras oncogenes of sarcoma viruses
around 1980 (111, 660). Their cellular oncogenes were
then identified in humans, and their mutations were furthermore found in some human carcinomas (146, 252,
499, 561, 626, 661). The mutated forms were subsequently
shown to stimulate proliferation and transformation of
cultured cells (71, 84, 182, 681). Moreover, the mutated
forms were shown to induce cell differentiation in neuronal cells (41, 249, 523). These findings drew the attention
of many scientists not only in the cancer research field but
also in many other fields. Finally, these Ras proteins were
shown to be related to the heterotrimeric G proteins, such
as Gs and Gi, and G proteins involved in protein synthesis,
such as elongation factor Tu (EF-Tu) (222, 644, 659).
The Rho gene was discovered as a homolog of the
Ras gene in Aplysia in 1985 (421); the YPT1 gene, which
had been discovered as an open reading frame between
the actin and tubulin genes in the yeast Saccharomyces
cerevisiae (S. cerevisiae) in 1983 (208), was identified to
encode a small G protein in 1986 (641); Arf protein, which
was purified as a cofactor for the cholera toxin-catalyzed
ADP-ribosylation of Gs in 1984 (326), was identified to
encode a small G protein in 1986 (327). The SEC4 gene,
which had been isolated as a gene involved in secretion in
the yeast in 1980 (527), was identified to encode a small G
protein in 1987 (623). These results suggested the presence of a big family of Ras-like small G proteins. Actually,
many small G proteins were systematically isolated by

molecular biological (100, 101, 105, 551, 573) and biochemical (291, 337, 344, 534, 535, 793, 797) methods.
Now, more than 100 small G proteins have been
identified in eukaryotes from yeast to human, and they
comprise a superfamily (60, 250, 701). The members of
this superfamily are structurally classified into at least five
families: the Ras, Rho, Rab, Sar1/Arf, and Ran families
(Table 1 and Fig. 1). In the yeast S. cerevisiae, sequence
analysis against complete genomic sequence has revealed
that there are 4 Ras family members, 6 Rho family members, 11 Rab family members, 7 Sar1/Arf family members,
and 2 Ran family members (210, 383). The functions of
many small G proteins have recently been elucidated: the
Ras subfamily members (Ras proteins) of the Ras family
mainly regulate gene expression, the Rho/Rac/Cdc42 subfamily members (Rho/Rac/Cdc42 proteins) of the Rho
family regulate both cytoskeletal reorganization and gene
expression, the Rab and Sar1/Arf family members (Rab
and Sar1/Arf proteins) regulate intracellular vesicle trafficking, and the Ran family members (Ran) regulate nucleocytoplasmic transport during the G1, S, and G2 phases
of the cell cycle and microtubule organization during the
M phase. Many upstream regulators and downstream effectors of small G proteins have been identified, and
modes of activation and actions have gradually been elucidated. In this review, functions of small G proteins and
their modes of activation and action are described. However, this review may not cover all detailed information
regarding each small G protein; readers may refer to other
recent excellent reviews (3, 49, 58, 80, 82, 139, 325, 417,
420, 428, 483, 491, 519, 536, 630, 713, 744, 758).
As to nomenclature, the term small GTPases is often
used, but small G proteins is used here because small G

TABLE

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1. The small G protein superfamily

Ras Family

Mammal

Ha-Ras
Ki-Ras
N-Ras
R-Ras
M-Ras
RalA
RalB
Rap1A
Rap1B
Rap2A
Rap2B
Tc21
Rit
Rin
Rad
Kir/Gem

Yeast

Ras1
Ras2
Rsr1
Yct7

Rheb
B-Ras1
B-Ras2

Rho Family

RhoA
RhoB
RhoC
RhoD
RhoE/
Rnd3/
Rho8
RhoG
RhoH/
TTF
Rac1
Rac2
Rac3
Cdc42
Rnd1/
Rho6

Rnd2/
Rho7
Tc10

Rho1
Rho2
Rho3
Rho4
Cdc42
Yns0

Rab Family

Rab1A
Rab1B
Rab2
Rab3A
Rab3B
Rab3C
Rab3D
Rab4
Rab5A
Rab5B
Rab5C
Rab6
Rab7
Rab8
Rab9
Rab10

Rab11A
Rab11B
Rab12
Rab13
Rab14
Rab15
Rab16
Rab17
Rab18
Rab19
Rab20
Rab21
Rab22
Rab23
Rab24
Rab25

Ypt1
Sec4
Ypt31/
Ypt8
Ypt32/
Ypt9
Ypt51/
Vps21

Ypt52
Ypt53
Ypt6
Ypt7
Ypt10
Ypt11

Rab26
Rab27A
Rab27B
Rab28
Rab29
Rab30
Rab31
Rab32
Rab33A
Rab33B

Sar1/Arf
Family

Ran
Family

Arf1
Arf2
Arf3
Arf4
Arf5
Arf6
Sar1a
Sar1b
Arl1
Arl2
Arl3
Arl4
Arl5
Arl6
Arl7
Ard1

Ran

Arf1
Arf2
Arf3
Sar1
Arl1
Arl2
Cin4

Gsp1
Gsp2

See References 251a and 810a.

proteins have both GDP/GTP-binding and GTPase activities. In many cases, the GTPase activity is necessary for
the termination of the functions of small G proteins, but
not essential for them to perform their functions. From
this point of view, the term GTPase is misleading. G
proteins represent heterotrimeric G proteins and small
GTP-binding proteins should be used, but just for simplicity small G proteins is used here. G proteins used
here include heterotrimetric G proteins (223), G proteins
involved in protein synthesis (338), and small G proteins.
Guanine nucleotide exchange factor (GEF) is often used,
but guanine nucleotide exchange protein (GEP) is used
here, because all GEFs thus far found are proteins and
GEPs is a more correct term.
II. GENERAL PROPERTIES
A. Structure
A comparison of the amino acid sequences of Ras
proteins from various species has revealed that they are
conserved in primary structures and are 30 55% homologous to each other. Among Ras proteins, each protein
shares relatively high (50 55%) amino acid identity,
whereas Rab and Rho/Rac/Cdc42 proteins share 30%
amino acid identity with Ras proteins (250, 742). Nevertheless, like other G proteins, all small G proteins have

consensus amino acid sequences responsible for specific

interaction with GDP and GTP and for GTPase activity,
which hydrolyzes bound GTP to GDP and Pi (61, 701, 742)
(Fig. 2A). Moreover, they have a region interacting with
downstream effectors. In addition, small G proteins belonging to Ras, Rho/Rac/Cdc42, and Rab proteins have
sequences at their COOH termini that undergo posttranslational modifications with lipid, such as farnesyl, geranylgeranyl, palmitoyl, and methyl moieties, and proteolysis (89, 228, 427, 701, 811) (Fig. 3). Arf proteins have an
NH2-terminal Gly residue that is modified with myristic
acid (493). Sar1 and Ran do not have such sequences to
direct posttranslational modifications.
Crystallographic and NMR analyses of some small G
proteins, including Ha-Ras, N-Ras, Rap2A, RhoA, Rac1,
Rab3A, Rab7, Arf1, and Ran, have revealed that all GDP/
GTP-binding domains have a common topology (219)
(Fig. 2B). By comparison of the structure of Ha-Ras in the
GTP-bound conformation and the GDP-bound conformation, two highly flexible regions surrounding the -phosphate of GTP have been established (471, 560): the switch
I region within loop L2 and 2 (the effector region) and
the switch II region within loop L4 and helix 2. A twostate model for the movement of the effector loop in the
GTP-bound form of Ha-Ras has been established; the
flexibility of the loop can conveniently be monitored by a
large shift of Tyr-32 relative to the phosphate groups,
because the hydroxyl group of Tyr-32 forms hydrogen

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FIG.

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1. Dendrogram of the small G protein superfamily. [Modified from Garcia-Ranea and Valencia (210).]

bond with the -phosphate of GTP. Binding of c-Raf-1, an

effector of Ras proteins (see below), stabilizes the effector loop in the active conformation (459).
The COOH-terminal regions are classified into at
least four groups: 1) Cys-A-A-X (A, aliphatic acid; X, any
amino acid); 2) Cys-A-A-Leu/Phe; 3) Cys-X-Cys; and 4)
Cys-Cys (89, 228, 427, 701) (Fig. 3). The Cys-A-A-X struc-

ture is furthermore subclassified into two groups: one has

an additional Cys residue upstream of the Cys residue of
the Cys-A-A-X structure (1a), and the other has a polybasic region (1b). In the case of the Cys-A-A-X structure,
Ha-Ras and Ki-Ras are first farnesylated at the Cys residue
followed by the proteolytic removal of the A-A-X portion
and the carboxylmethylation of the exposed Cys residue

FIG. 2. Structure of small G proteins. A: consensus

amino acid sequences responsible for specific interaction
with GDP and GTP and for GTPase activity. B: crystallographic structure of small G proteins. The crystallographic
structure of Ha-Ras is representatively shown. A, Ala; D,
Asp; E, Glu; G, Gly; K, Lys; N, Asn; S, Ser; X, any amino
acid. [Modified from Pai et al. (560).]

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SMALL G PROTEINS

FIG. 3. The COOH-terminal structures and posttranslational modifications of small G proteins. The COOH-terminal regions of small G
proteins are classified into at least four groups (1 4). The Cys-A-A-X
structure is furthermore subclassified into two groups (1a and 1b). A,
aliphatic acid; X, any amino acid; P, palmitoyl; F, farnesyl; GG, geranylgeranyl.

(90, 201, 247, 256, 313). Ha-Ras has an additional Cys

residue that is further palmitoylated (256). The Cys-A-ALeu structure of Rap1 is first geranylgeranylated followed
by the same modifications (336). Both Cys residues of the
Cys-X-Cys structure of Rab3A are geranylgeranylated, and
the COOH-terminal Cys residue is carboxylmethylated
(177). Both Cys residues of the Cys-Cys structure of Rab1
are geranylgeranylated, but the COOH-terminal Cys residue is not carboxylmethylated (672). The lipid modifications of these small G proteins are necessary for their
binding to membranes and regulators and for their activation of downstream effectors as described below (89,
228, 257, 427, 701, 702, 811).
The farnesyl moiety is derived from farnesyl pyrophosphate, an intermediate product of the mevalonate
pathway which produces cholesterol from mevalonate
(231). Mevalonate is produced from 3-hydroxy-3-methyglutaryl-CoA by the action of 3-hydroxy-3-methyglutarylcoenzyme reductase. A specific inhibitor for this enzyme,
named pravachol, is used as a very effective drug for
arteriosclerosis (171, 231). The geranylgeranyl moiety is
derived from geranylgeranyl pyrophosphate, which is an
intermediate product for the synthesis of dolichol and
ubiquinone (220, 231). The palmitoyl moiety is derived
from palmitoyl CoA. The methyl moiety is derived from
S-adenosyl-methionine. The enzymes that transfer the
prenyl moieties have been isolated and characterized
(811). The farnesylation of the Cys-A-A-X structure is
catalyzed by farnesyltransferase, the geranylgeranylation
of the Cys-A-A-Leu structure is catalyzed by geranylgeranyltrasnferase I, and the prenylation of the Cys-X-Cys and
Cys-Cys structures is catalyzed by geranylgeranyltransferase II. Farnesyltransferase and geranylgeranyltransferase I consist of two subunits, and subunits, and the

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-subunits of both enzymes are identical (648). Geranylgeranyltransferase II consists of three subunits, originally termed component A but recently renamed Rab
escort protein I (Rep1), and - and -subunits (289, 645,
646, 672). Rep1 binds unprenylated Rab proteins, presents
them to the catalytic -subunits, and remains bound to
Rab proteins after the geranylgeranyl transfer reaction
(20). In cells, Rab GDP dissociation inhibitor (GDI) (see
below) may dissociate this product from Rep1, allowing
multiple cycles of catalysis. The human Rep1 gene has
been identified by positional cloning as that responsible
for choroideremia, which is an X-linked form of retinal
degeneration (131, 132, 187). Loss of Rep1 activity causes
the reduced prenylation of Ram/Rab27, which is expressed at high levels in the retinal cell layers, and the
degeneration of this protein in the progression of this
disease (647). Palmitoyltransferase that palmitoylates HaRas has been purified (406), but it is not yet known
whether this enzyme is the one that functions in vivo.
Methyltransferases that transfer the methyl moieties to
small G proteins having Cys-A-A-X, Cys-A-A-Leu, and CysX-Cys structures have not been well characterized. A
protease, Rce1, that removes the A-A-X and A-A-Leu portions of Ha-Ras, N-Ras, Ki-Ras, and Rap1B, has recently
been identified (345, 557).
B. A Role as Molecular Switches
According to the structures of small G proteins, they
have two interconvertible forms: GDP-bound inactive and
GTP-bound active forms (60, 250, 701) (Fig. 4). An upstream signal stimulates the dissociation of GDP from the
GDP-bound form, which is followed by the binding of
GTP, eventually leading to the conformational change of
the downstream effector-binding region so that this region interacts with the downstream effector(s). This interaction causes the change of the functions of the downstream effector(s). The GTP-bound form is converted by
the action of the intrinsic GTPase activity to the GDPbound form, which then releases the bound downstream
effector(s). In this way, one cycle of activation and inactivation is achieved, and small G proteins serve as molecular switches that transduce an upstream signal to a
downstream effector(s).
Thus the rate-limiting step of the GDP/GTP exchange
reaction is the dissociation of GDP from the GDP-bound
form. This reaction is extremely slow and therefore stimulated by a regulator, named GEP (also called GEF or
guanine nucleotide releasing factor), of which activity is
often regulated by an upstream signal. GEP first interacts
with the GDP-bound form and releases bound GDP to
form a binary complex of a small G protein and GEP.
Then, GEP in this complex is replaced by GTP to form the
GTP-bound form. Most GEPs, such as Son of Sevenless
(SOS), a Ras GEP, and Rab3 GEP, are specific for each

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FIG.

4. Regulation of small G protein activity.

member or subfamily of small G proteins (56, 75, 762), but

some GEPs, such as Dbl, a GEP active on Rho/Rac/Cdc42
proteins, show wider substrate specificity (259, 788). The
GDP/GTP exchange reactions of Rho/Rac/Cdc42 and Rab
proteins are furthermore regulated by another type of
regulator, named Rho GDI and Rab GDI, respectively (28,
206, 453, 629, 732). This type of regulator inhibits both the
basal and GEP-stimulated dissociation of GDP from the
GDP-bound form and keeps the small G protein in the
GDP-bound form. Rho GDI and Rab GDI show wider
substrate specificity than GEPs and GTPase-activating
proteins (GAPs) and are active on all Rho/Rac/Cdc42 and
Rab proteins, respectively (18, 275, 391, 627, 629, 732,
737). Thus the activation of Rho/Rac/Cdc42 and Rab proteins is regulated by positive and negative regulators.
Recently, Ran GDI, p10/NTF2, has also been reported
(125, 485, 563, 789), but GDIs have not been identified for
other small G proteins. The GTPase activity of each small
G protein is variable but relatively very slow and is stimulated by GAPs. Most GAPs, such as Ras GAP and Rab3
GAP, are specific for each member or subfamily of small
G proteins (56, 205, 727), but some GAPs, such as p190, a
GAP active on Rho/Rac/Cdc42 proteins, show wider substrate specificity (656).

Volume 81

pathway, such as neurons, neuroendocrine cells, and exocrine cells (140, 183, 476, 477, 625). Rab17 is detected in
epithelial cells (413). Most small G proteins are localized
either in the cytosol or on membranes. Ran is localized
either in the cytosol or in the nucleus. Each small G
protein is localized to a specific membrane. Ras proteins
are localized at the cytoplasmic face of the plasma membrane. This localization is mediated by the posttranslational modifications with lipid. The farnesyl moiety of
Ha-Ras and Ki-Ras alone is not sufficient for their binding
to the membrane (256). In the case of Ha-Ras, both the
farnesyl and palmitoyl moieties are necessary, whereas in
the case of Ki-Ras, both the farnesyl moiety and the
neighboring clustered polybasic amino acids are necessary. The farnesyl and palmitoyl moieties may interact
with the acyl moieties of the phospholipids, whereas the
polybasic amino acids may interact with the polar head
groups of the acidic phospholipids. The methyl moiety
also contributes markedly to efficient membrane association (255). Rap1 is geranylgeranylated and has clustered
polybasic amino acids. Most Rab proteins have either a
Cys-X-Cys or Cys-Cys structure of which Cys residues are
both geranylgeranylated. These small G proteins are localized at the cytoplasmic faces of distinct membrane
compartments. It has not been experimentally clarified
how Rap1 and Rab proteins exactly interact with the
membranes, but it is likely that both the prenyl moiety
and the polybasic region or two prenyl moieties are necessary. In contrast, Arf proteins have one myristoyl moiety and Sar1 has no lipid moiety, but they interact with the
cytoplasmic faces of membranes. Arf proteins interact
with membrane lipids by its myristoylated and amphipathic NH2-terminal helix (21, 47). In the case of Sar1, it
may interact with the phospholipid through only peptide
region. Small G proteins, such as Rho/Rac/Cdc42 and Rab
proteins, located on the plasma membrane and the cytosol are translocated between these two sites. Ran is also
translocated between the cytosol and the nucleus through
the nuclear pore complexes (NPCs).
III. RAS PROTEINS AS REGULATORS
OF GENE EXPRESSION

C. Localization
A. Outline
Small G proteins as well as heterotrimeric G proteins
are present only in eukaryotes from yeast to human,
although G proteins involved in protein synthesis such as
elongation factors exist in both prokaryotes and eukaryotes. Most small G proteins are widely distributed in
mammalian cells, and most cells have the Ras, Rho, Rab,
Sar1/Arf, and Ran families, although expression levels of
their members may vary from one type to another. A few
members show tissue-specific expression; for instance,
Rab3A is expressed in cells having a regulated secretion

Three Ras proteins are now known, Ha-Ras, Ki-Ras,

and N-Ras, which are capable of transforming mammalian
cells when activated by point mutations (71, 84, 182, 681).
In the yeast S. cerevisiae, there are two members of Ras
proteins, Ras1 and Ras2, that are essential for cell viability, and these yeast genes are functionally replaceable by
mammalian genes (141, 577). The downstream effector of
Ras proteins was first identified to be adenylate cyclase in
the yeast S. cerevisiae (66, 721). Mammalian adenylate

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SMALL G PROTEINS

cyclase is directly regulated by heterotrimeric G proteins,

but not by Ras proteins. Subsequently, genetic, cell biological, and biochemical studies in Caenorhabditis
elegans, Drosophila, and mammalian cells established the
mode of action of Ras proteins; they directly bind to and
activate Raf protein kinase (82, 151a, 254a, 743, 759, 767,
815), which then induces gene expression through the
mitogen-activated protein (MAP) kinase cascade in response to various extracellular signaling molecules (145,
299, 416). Other studies have clarified that Ras proteins
regulate not only cell proliferation but also differentiation
(41, 249, 523), morphology (41a, 182, 779), and apoptosis
(334). Ras proteins regulate these functions mainly
through gene expression, but it has not been established
whether the Ras protein-mediated morphological changes
are a direct effect or an indirect effect through gene
expression. Another characteristic feature of Ras proteins
is that the mutations of their genes and their regulator
genes cause human cancers (9, 42, 59, 445, 607, 756, 786).
Thus Ras proteins are crucially important molecules not
only for biology but also for human health.
B. Ras Protein Cycle: Activation/Inactivation
Ras protein activity is regulated by GEPs and GAPs,
and activation is induced by a large variety of extracellular signals, most notably signals that activate receptors
with intrinsic or associated tyrosine kinase activity (168,
207, 399, 549, 617) (Fig. 5). Phosphotyrosines serve as
docking sites for the adaptor proteins, such as GRB2 and
SHC/GRB2 complex, which then recruit SOS, the most
characterized Ras GEP, from the cytosol to produce a

159

receptor-adaptor-GEP complex. SOS recruited to the

plasma membrane then stimulates a Ras protein located
at the cytoplasmic face of the plasma membrane and
converts it from the the GDP-bound form to the GTPbound form. It is believed that GRB2 recruits SOS from
the cytosol to the plasma membrane without affecting its
GEP activity, but the possibility has not been totally excluded that GRB2 both recruits and activates SOS. Receptors not directly associated with tyrosine kinases, such as
T-cell receptors, may activate Ras proteins indirectly
through Src-like tyrosine kinases or ligand-independent
activation of receptor tyrosine kinases (243, 690, 774).
Moreover, heterotrimeric G protein-coupled receptors,
such as -adrenergic receptors, muscarinic acetylcholine
receptors, and lysophosphatidic acid, have also been
shown to activate Ras proteins (266, 285, 294). In addition,
an increase of cytoplasmic Ca2 induced by activation of
these receptors in neurons also induces activation of
another type of GEPs, p140 Ras GRF, that contains an IQ
motif regulated by calcium-bound calmodulin (178).
After the GTP-bound forms of Ras proteins accomplish their effects on downstream effector(s), they are
converted to the GDP-bound form by the action of Ras
GAPs. However, how termination of Ras protein signaling
is achieved is not fully understood. Phosphorylation of
SOS by the Raf/MAP kinase pathway (see below) may
induce the dissociation of SOS from GRB2 (281, 356), and
another possible mechanism is that Ras GAPs are activated by their binding to tyrosine-phosphorylated growth
factor receptors such as the platelet-derived growth factor (PDGF) receptor (17, 332).
Three GEPs of Ras (SOS, Cdc25, and Ras GRF) have

FIG. 5. Mode of action of Ras proteins in gene expression. Ras, Ras proteins.

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been discovered to date. The first GEP for Ras, Cdc25, has
been identified genetically in S. cerevisiae (67, 81, 605).
Cdc25 has a GEP domain that is required for its catalytic
activity, located in its COOH-terminal region. In addition,
Cdc25 has an SH3 domain in the NH2-terminal region. In
higher eukaryotes, in addition to mammalian Cdc25
(mCdc25), two different types of proteins with homology
to Cdc25 have been found. The first group includes SOS.
SOS was first identified in Drosophila. Genetic studies
have shown that SOS is downstream of Sevenless, a receptor tyrosine kinase, which is homologous to the epidermal growth factor (EGF) receptor (609, 669). One
human and two murine homologs of SOS have been
cloned. SOS has one pleckstrin homology (PH) domain
that may interact with a membrane phospholipid, phosphatidylinositol 4,5-bisphosphate (PIP2), to determine its
localization (62, 98). In addition, SOS has a GEP domain
that is required for its catalytic activity in the middle
portion and a GRB2-binding site in its COOH-terminal
region. The second GEP group includes p140 Ras GRF
that is primarily expressed in neural tissues (667). p140
Ras GRF has also a PH domain and a GEP domain. In
addition, p140 Ras GRF contains an IQ motif that is
regulated by calcium-bound calmodulin (178). Ras GRP, a
GEP also predominantly expressed in brain, binds Ca2
directly through a structure similar to the calcium binding
EF hand; moreover, Ras GRP is regulated by direct binding of diacylglycerol (166). SOS is active on Ha-Ras and
Ki-Ras but not on R-Ras, whereas Ras GRF is active on
Ha-Ras and N-Ras but not on RalA or Cdc42 (56, 75,
507, 667).
p120 Ras GAP is the first GAP to be characterized at
a molecular level (221, 727, 728, 755). p120 Ras GAP is
active on Ha-Ras, Ki-Ras, N-Ras, and R-Ras, but not on
Rho/Rac/Rab proteins (56, 221, 727). It contains a hydrophobic NH2 terminus, two Src homology-2 (SH2) domains, one Src homology-3 (SH3) domain, one PH domain, and a region similar to the calcium-dependent lipid
binding region of phospholipase A2. Although it is clear
that p120 Ras GAP acts as a negative regulator of Ras
proteins, it has long been speculated that it also has other
functions. One possible role of p120 Ras GAP is that it is
a downstream effector of Ras proteins (463, 804). However, it now seems unlikely that p120 Ras GAP plays a
major downstream role of Ras proteins. A number of
tyrosine protein kinases have been shown to stimulate
tyrosine phosphorylation of p120 Ras GAP in a variety of
cells (169, 481), although the stoichiometry of these phosphorylations is generally low, and no alteration in the
activity or localization of p120 Ras GAP has been demonstrated upon tyrosine phosphorylation. Protein tyrosine
kinases may control the activity of p120 Ras GAP by
forming a complex with it. In fact, p120 Ras GAP forms a
complex with activated PDGF receptors (332, 339). Two
other proteins have been found to associate with p120 Ras

Volume 81

GAP. One is p62 Dok, a docking protein that contains

multiple tyrosine phosphorylation sites for its binding to
the SH2 domains of p120 Ras GAP and Nck (87, 796). The
other is p190 Rho GAP; however, the precise role of the
interaction between p190 Rho GAP and p120 Ras GAP is
not fully understood (486, 656). p120 Ras GAP-deficient
mice die during embryonic development, and the ability
of endothelial cells to organize a vascular network is
severely impaired in addition to extensive neuronal cell
death in these animals (268). Another Ras GAP named
GAP1m, which shows a high degree of similarity to the
Drosophila Gap1 gene, has been also identified (426). In
addition to the GAP catalytic domain, GAP1m has two
domains with sequences closely related to those of the
phospholipid-binding domain of synaptotagmin and a region with similarity to the unique domain of Btk tyrosine
kinase.
S. cerevisiae contains two Ras GAPs, Ira1 and Ira2
(711, 712), which contain domains homologous to the
COOH terminus of p120 Ras GAP. In wild-type cells, Ras
proteins are generally inactive, but in the absence of
either the IRA gene product, they accumulate in their
GTP-bound state, becoming hyperactive and leading to
overproduction of cAMP. In yeast, at least, Ras GAPs are
therefore not the effectors of Ras proteins; rather, they
serve as negative regulators.
Neurofibromin (NF1), the human protein defective in
von Recklinghausen neurofibromatosis (a benign tumor),
contains a domain homologous to the catalytic domains
of p120 Ras GAP, Ira1, and Ira2 (445, 786). Like p120 Ras
GAP itself, neurofibromin possesses GAP activity in vitro
and can complement the loss of Ira function in the yeast
S. cerevisiae, indicating that it may be the mammalian
homolog of Ira1 and Ira2 (34). These proteins share regions of significant similarity outside the GAP-related domain and presumably perform similar functions.
Small G protein GDP dissociation stimulator (Smg
GDS) is a regulator that is entirely distinct from GEPs,
GDIs, and GAPs (328, 795). Smg GDS has two biochemical
activities on a group of small G proteins including Ki-Ras
and the Rho/Rac/Cdc42/Rap1 proteins: one is to stimulate
their GDP/GTP exchange reactions and the other is to
inhibit their binding to membranes (18, 275, 479, 480, 788).
A detailed kinetic study of Smg GDS with Ki-Ras as a
substrate has revealed that it interacts with not only
GDP-Ki-Ras and the guanine nucleotide-free Ki-Ras but
also GTP-Ki-Ras, under the conditions where a Ras GEP,
mCdc25, does not form a ternary complex with GTP-KiRas (506). This property of Smg GDS suggests that it
stimulates the GDP/GTP exchange reaction only once.
The physiological role of Smg GDS remains to be established, although it shows mitogenic and transforming activities in cooperation with Ki-Ras in fibroblasts (198).
Studies on Smg GDS-deficient mice have revealed that
mice die of heart failure shortly after birth (708). En-

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SMALL G PROTEINS

hanced apoptosis is observed in at least heart, thymus,

and neuron in Smg GDS-deficient mice. This phenotype is
apparently similar to those of Ki-Ras-deficient mice (321,
362), providing another line of evidence that Smg GDS
plays a role in Ki-Ras-mediated signaling pathway in vivo.
C. Raf Protein Kinase Activation by Ras Proteins
Ras proteins mediate their effects on cell proliferation mainly by activation of a cascade of protein kinases:
Raf protein kinase (c-Raf-1, A-Raf, and B-Raf), MEK (MAP
kinase kinases 1 and 2), and MAP kinase. Ras proteins
activate this protein kinase cascade by directly binding to
Raf proteins (743, 759, 767, 815). Raf proteins then phosophorylate and activate MEK (145, 299, 378), which then
phosphorylates and activates MAP kinase (134, 450). The
activated MAP kinase translocates to the nucleus, where
it phosphorylates and stimulates the activity of various
transcription factors, including Elk-1 (442). The recent
observation that Ras proteins interact with two distinct
NH2-terminal regions of Raf-1 suggests that Ras proteins
promote more than just membrane translocation of Raf-1
and instead may also facilitate the subsequent events that
lead to Raf-1 activation (495, 513, 759).
The initial step of the Raf-1/MEK/MAP kinase cascade is the activation of Raf-1 by direct interaction with a
GTP-Ras protein (GTP-Ras) (743, 759, 767, 815). The interaction with GTP-Ras localizes Raf-1 to the plasma
membrane (389, 687). The first described and best-characterized Ras-binding domain (RBD) is contained within
residues 51131 of Raf-1 (121, 513, 759). The association
of the RBD with the Ras effector domain is a high-affinity
interaction that is mediated primarily by residues Gln-66,
Lys-84, and Arg-89 of Raf-1 (55). The interaction between
the RBD and GTP-Ras appears to then allow for a second
RBD of Raf-1 to contact GTP-Ras (72, 159). This second
RBD (residues 139 184 of Raf-1) encompasses the conserved Cys finger motif within the Raf-1 NH2 terminus and
is referred to as the Cys-rich domain (CRD) (495). In
terms of the Ras-Raf-1 interaction, the CRD associates
with different residues of GTP-Ras than does the RBD
(159), and posttranslational modifications of Ras proteins
may be important for CRD binding (297, 376). Thus, in the
full-length molecule, the CRD is inaccessible for GTP-Ras
binding, but either mutational events or RBD binding can
unmask the CRD and allow it to interact with GTP-Ras.
Thus, for the Ras-Raf-1 interaction to result in Raf-1 activation, binding to both the RBD and the CRD appears to
be required.
Ha-Ras is localized at the cytoplasmic surface of the
plasma membrane, while mutant forms of Ha-Ras, which
lack posttranslational lipid modification, are cytosolic and
lack biological activity. These findings suggest that posttranslational lipid modifications of Ha-Ras are required

161

for both its localization and biological activity. Furthermore, lipid-modified Ras proteins have been shown to
more efficiently activate adenylate cyclase in the yeast
system (288, 376). Posttranslational modifications of KiRas are also required for MAP kinase activation in a
cell-free system (309). In addition, posttranslational modifications of Ha-Ras are required for activation of, but not
for association with, Raf protein kinase (392, 547).
The intracellular signals that couple growth factors
to MAP kinase may determine the different effects of
growth factors; for example, transient activation of MAP
kinase by EGF stimulates proliferation of PC12 cells,
whereas sustained activation of MAP kinase by nerve
growth factor (NGF) induces differentiation of PC12 cells.
Activation of MAP kinase by NGF involves two distinct
pathways: the initial activation of MAP kinase requires
Ras proteins, but its activation is sustained by Rap1 (806)
(see below). Rap1 is activated by C3G, a GEP for Rap1,
and forms a stable complex with B-Raf (806). Activation
of B-Raf by Rap1 represents a common mechanism to
induce sustained activation of the MAP kinase cascade.
D. Modifiers of the Ras Protein-Induced Raf
Protein Kinase Activation
In addition to Ras proteins, a protein named 14 3-3
seems to also interact with Raf-1 and activate it (176, 197).
14 3-3 is a specific phosphoserine-binding protein (500).
Raf-1 itself contains two phosphorylation sites that may
interact with 14 3-3. 14 3-3 may have two different roles:
first, 14 3-3 may be required for maintaining Raf-1 in an
inactive conformation, as Raf-1 that is unable to stably
interact with 14 3-3 is activated (467). In response to
signaling events and Ras protein activation, 14 3-3 may
subsequently play a second role in facilitating activation
of Raf-1 and stabilizing activated Raf-1.
The observation that Raf-1 becomes hyperphosphorylated in response to many signaling events (492) has
long suggested that phosphorylation plays a role in regulating Raf-1 activity. Mechanisms by which phosphorylation could regulate Raf-1 function include direct alteration of the intrinsic activity of Raf-1 and alteration of
critical protein interactions, such as with 14 3-3. The
rapid and transient nature of Raf-1 activation further complicates the issue, making it difficult to distinguish between activating and inactivating modifications. Nevertheless, by the use of overexpression systems and
mutational analysis, the phosphorylation of tyrosine residues 340 and 341 has been shown to enhance the catalytic activity of Raf-1 (441). The tyrosine kinases implicated in phosphorylating Raf-1, and thereby enhancing its
activity, include members of the Src kinase family (441,
562, 576).
A novel protein kinase that functions downstream of

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TAKAI, SASAKI, AND MATOZAKI

Ras proteins, kinase suppressor of Ras (KSR), has recently been identified by genetic screening as a suppressor of phenotypes caused by an activated Ras protein in
both Drosophila and C. elegans (366, 694, 717). Epistasis
analysis in Drosophila suggests that KSR functions downstream of Drosophila Ras-1 but upstream or in parallel to
Raf protein (717). Characterization of a mouse KSR homolog suggests that KSR facilitates signal transmission
between Raf proteins, MEK, and MAP kinase (718). In
addition, upon Ras protein activation, KSR translocates to
the plasma membrane, where it forms a stable complex
with Raf proteins (718). Moreover, KSR, via its kinase
domain, forms a stable complex with MEK in the cytosol
of quiescent cells (144). Therefore, in response to an
activated Ras protein, KSR might shuttle MEK from the
cytosol to activated Raf proteins at the membrane.
With the use of a screen for eye development defects in Drosophila, the connector enhancer of KSR
(CNK) protein has been identified as an enhancer of a
KSR dominant negative mutant phenotype (719). Mutation of CNK suppresses the phenotype of activated Ras
proteins or Sevenless but not Raf proteins, suggesting
that it acts upstream of Raf proteins. CNK has several
protein interaction domains. These domains include a
sterile alpha motif (SAM) domain, a PSD-95/Dlg-A/ZO-1
(PDZ) domain, two proline-rich (potential SH3-binding)
domains, and a PH domain; such domains are found in
many proteins involved in signaling and suggest further
interactions of CNK with other proteins and small molecules. In two-hybrid assays in S. cerevisiae, a COOHterminal portion of CNK that contains the PH domain
interacts with the Raf kinase domain (719). Thus the
SAM, PDZ, and novel domains might be available for
other interactions, although it is not known whether
CNK also binds other proteins in the Ras-Raf signaling
pathway. CNK has a molecular structure similar to that
of recently identified rat neuronal proteins, named
MAGUINs, that interact with the PDZ domains of PSD95/SAP90 and S-SCAM (802). MAGUINs interact with
c-Raf-1 but do not affect its enzymatic activity (803).
PSD-95/SAP90 and S-SCAM are neuronal membraneassociated guanylate kinases, and these proteins function as synaptic scaffolding proteins (264). In fact,
PSD-95/SAP90 further interacts with synGAP, which
regulates the activity of Ras proteins (346). Therefore,
MAGUINs may also bind Raf proteins and link it to
PSD-95/SAP90 and S-SCAM in synaptic junctions.
E. Other Effectors of Ras Proteins
A variety of candidate Ras protein effectors have
been reported in addition to Raf proteins. These include
Ral GDS (279, 342, 676), RIN1 (254), and phosphatidylino-

Volume 81

sitol (PI) 3-kinase (608). AF6/Canoe is also suggested to

be a binding partner of Ras proteins (373, 746), but this
result has been called into question (434). It has recently
been shown that Rap1 shows a much higher affinity to
AF6 than Ras proteins do (404). p120 Ras GAP may participate in Ras protein-mediated gene expression, although it is still unclear whether p120 Ras GAP is a
regulator, an effector, or both for Ras proteins. In contrast, activation of PI 3-kinase by Ras proteins may promote cell survival (334, 608). However, it has not been
established whether these effector molecules other than
Raf proteins really play a role in the downstream pathway
of Ras proteins.
F. Transport of Newly Synthesized Ras Proteins
From the Endoplasmic Reticulum to the
Plasma Membrane
The plasma membrane localization of Ras proteins is
crucial for their functions. The mechanism by which Ras
proteins get to the plasma membrane has not fully been
understood. It has recently been shown that Ras proteins
do not directly travel to the plasma membrane from the
cytosol, but interact with intracellular membranes (25,
114). Ras proteins first associate with the endoplasmic
reticulum and then with the Golgi apparatus. The initial
association of Ras proteins with the endoplasmic reticulum requires only the COOH-terminal Cys-A-A-X structure
and farnesylation. N-Ras and Ha-Ras seem to be transported by exocytic vesicles following association with the
endoplasmic reticulum and the Golgi apparatus. Ki-Ras
takes a faster route that may not involve the Golgi apparatus. The hypervariable domains of the three Ras proteins are necessary and sufficient to account for their
differential localizations. Inhibition of vesicle transport
with brefeldin A (BFA), an inhibitor of Arf protein GEP
(see below), blocks the transit of N-Ras to the plasma
membrane, demonstrating the importance of vesicle
transport for N-Ras function. Carboxylmethylation and
A-A-X proteolysis are also necessary for proper association with the plasma membrane (255).
G. Ras Proteins and Cancer
Mutated versions of the three human Ras genes have
been detected in 30% of all human cancers, implying an
important role for aberrant Ras protein function in carcinogenesis. For example, Ras gene mutations are highly
prevalent in pancreatic (90%) (9), lung (30%) (607), and
colorectal (50%) (59, 756) carcinomas. Because Ras proteins regulate diverse extracellular signaling pathways for
cell growth, differentiation, and apoptosis, the deregulated function of other cellular components can cause

January 2001

SMALL G PROTEINS

aberrant Ras protein function in the absence of mutations

in the Ras genes themselves. Overexpression of ErbB2 or
EGF receptor tyrosine kinase is common in breast cancers, and their transforming actions are dependent on
signaling through the loss of negative Ras protein regulators (155). Similarly, the loss of function of negative Ras
protein regulators, such as neurofibromin defective in
type 1 neurofibromatosis-associated tumors, can cause
aberrant upregulation of Ras protein function (42). Therefore, the importance of aberrant Ras protein function in
human cancers may be greater than expected and may
extend to tumors that do not harbor mutated Ras alleles.
H. Rap Proteins
The Rap subfamily consists of Rap1A, Rap1B, and
Rap2. Rap1 proteins have been independently isolated by
three laboratories by different methods: they have been
isolated as homologs of Ras proteins by hybridization
(573), they have been purified as small G proteins (smg
p21) by column chromatography (337, 534), and they have
been identified as K-Rev1 in a screen for cDNAs that
revert the morphology of Ki-Ras-transformed cells (353).
Interestingly, Rap1 proteins have an effector domain virtually identical to that of Ras proteins, suggesting that
both proteins theoretically interact with similar effectors
and show similar or antagonistic effects. The antagonistic
function of K-Rev1 on Ras-transforming activity was the
first studied (353). Rap1A binds to the two Ras-binding
regions of Raf-1 (RBD and CRD), and this binding of
Rap1A to CRD is competitive with Ras proteins (296, 297).
Rap1 does not induce Raf-1 activation in intact cells but
inhibits the Ha-Ras-induced Raf-1 activation in intact cells
when Rap1 is overexpressed (124). However, most extracellular signals that induce Raf-1 activation, such as
PDGF and EGF, activate rather than inhibit Rap1 (828).
Furthermore, a phorbol ester induces Rap1 activation in
Rat1 cells, but does not inhibit the PDGF- and EGFinduced activation of MAP kinase (828). These results
suggest that the suppression of Ras protein function by
Rap1 is simply due to the artificially competitive inhibition of the Ras protein binding to RBD or CRD.
In contrast to the role of Rap1 antagonistic to that of
Ras proteins, evidence is accumulating that Rap1 functions independently of Ras protein signaling, utilizing effectors similar or identical to those of Ras proteins, like
Raf proteins. Rap1, as well as Ki-Ras, induces DNA synthesis in Swiss 3T3 cells (11, 807). Rap1, as well as Ki-Ras,
binds and activates B-Raf in vitro (541). In intact PC12
cells in response to cAMP and NGF, Rap1 is activated and
induces B-Raf activation, causing sustained activation of
the MAP kinase cascade that is necessary for neuronal
differentiation (761, 806). Most recently, CD31, an impor-

163

tant integrin adhesion amplifier, has been shown to selectively activate Rap1, but not Ha-Ras, R-Ras, or Rap2 (587).
An activated mutant of Rap1 stimulates T lymphocyte
adhesion to intercellular adhesion molecule and vascular
cell adhesion molecule, as does C3G. Thus Rap1 regulates
ligand-induced cell adhesion, and it may play a more
general role in coordinating adhesion-dependent signals.
In contrast to Rap1, little is known about Rap2 (573).
Several distinct second messenger pathways, including those for calcium (194), diacylglycerol (462), phospholipase C- (462), and cAMP (10), and perhaps others
(497a), are able to induce Rap1 activation. Clearly, Rap1
activation is a common event, which suggests a function
that is central in signal transduction processes. C3G is a
Rap1-specific GEP containing a proline-rich domain that
interacts with the SH3 domain of members of the Crk
adaptor proteins, Crk I, Crk II, and Crk L (239, 355). In
general, this association is constitutive, but tyrosine phosphorylation of Crk may disrupt the interaction (546). The
SH2 domain of Crk binds directly to various activated
receptor tyrosine kinases and phosphotyrosine-containing adaptor proteins (355). This association of Crk-C3G
with these complexes may enhance GEP activity of C3G
(301), suggesting that complex formation and dissociation
of C3G regulate Rap1 activation by tyrosine kinases. However, in a human Jurkat T cell leukemia line, T-cell receptor-dependent induction of a Cbl-Crk L-C3G signaling
complex does not activate Rap1 (586). Therefore, more
work will be required to clarify how C3G complex formation is coupled to Rap1 regulation. Recently, two novel
GEPs specific for Rap1, named Epac/cAMP-GEFI and
nRap GEP/PDZ-GEF1/Hs-RA-GEF, have been identified
(147, 148, 335, 402, 540). Epac/cAMP-GEFI has cAMPbinding and Ras GEP domains; thus this GEP activity is
dependent on cAMP (148, 335). nRap GEP has been isolated as a binding partner of S-SCAM, that interacts with
N-methyl-D-aspartate (NMDA) receptors and neuroligin
through PSD-95/Dlg-A/ZO-1 (PDZ) domains at synaptic
junctions (540). In contrast to Epac/cAMP-GEFI, nRap
GEP/PDZ-GEF1/Hs-RA-GEF has one PDZ, one Ras association, and one Ras GEP domains as well as one COOHterminal consensus motif for binding to PDZ domains.
However, nRap GEP/PDZ-GEF1/Hs-RA-GEF has an incomplete cAMP-binding domain and its GEP activity is
independent of cAMP. SPA-1, a Rap1 GAP, has been
shown to interfere with Rap1 activation by membranetargeted C3G (729). Overexpression of SPA-1 in HeLa
cells suppresses Rap1 activation upon plating on dishes
coated with fibronectin and results in the reduced adhesion. In addition, overexpression of SPA-1 in promyelocytic 32D cells also inhibited both activation of Rap1 and
induction of cell adhesion by granulocyte colony-stimulating factor, suggesting that Rap1 is required for the cell

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TAKAI, SASAKI, AND MATOZAKI

adhesion induced by both extracellular matrix and soluble ligands (729).

I. Ral Proteins
The Ral subfamily consists of RalA and RalB (100,
101). Ral GDS, a Ral GEP, has been found to be a Ras
protein effector (279, 342, 676). Moreover, insulin and
EGF induce activation of Ral proteins, and this activation
is inhibited by a dominant negative mutant of Ras proteins, suggesting that RalA is downstream of the Ras
protein signaling pathway (783). In NIH 3T3 cells, both a
dominant active mutant of Ha-Ras and Ral GDS synergize
with Raf-1 in the induction of cell transformation and the
activation of c-fos promoter (548, 741), and a dominant
negative mutant of RalA inhibits the Ha-Ras- and Raf-1induced transformation (741). These observations suggest
that the Ral GDS-Ral protein pathway contributes to cell
transformation and gene expression. However, a dominant active mutant of RalA alone cannot efficiently induce
the oncogenic transformation or the c-fos induction compared with a dominant active mutant of Ha-Ras and Ral
GDS (548, 741), suggesting that the transformation and
the gene expression induced by Ral GDS may require
other factors in addition to Ral proteins.
Three effectors for Ral proteins are known: RalBP1,
phospholipase D, and filamin. RalBP1 contains a Rho GAP
homology domain that exhibits the GAP activity for Rac/
Cdc42 proteins, but not for Rho proteins (179). Although
Rac/Cdc42 proteins contribute to the Ha-Ras-induced oncogenic transformation (582) (see below), it is unclear
whether the association of Ral proteins with RalBP1 regulates the activity of these Rho/Rac/Cdc42 proteins.
RalBP1 has been found to interact with POB1 and Reps1
(302, 791), which have proline-rich sequences responsible
for interaction with Grb2 and Crk, and an Eps15 homology domain. Ral proteins are involved in endocytosis of
the growth factor receptors probably through RalBP1,
POB1, Eps15, and Epsin (511). Another Ral protein effector, phospholipase D, is also implicated in vesicle trafficking (179, 316). The activity of phospholipase D is induced
by Src and Ras proteins. A dominant negative mutant of
RalA inhibits both v-Src- and v-Ras-induced phospholipase D activity (316). The third effector protein of Ral is
filamin (537). Either a dominant negative mutant of RalA
or the RalA-binding domain of filamin blocks Cdc42-induced filopodium formation. A dominant active mutant of
RalA elicits actin-rich filopodia, but it does not generate
filopodia in filamin-deficient cells. Thus the Ral signaling
appears to regulate vesicle trafficking, cytoskeletal organization, gene expression, and cell transformation. The
GAP proteins for RalA were characterized and partially
purified (48, 170); however, the molecular cloning of these
proteins has not yet been achieved.

Volume 81

J. Other Ras Family Members

R-Ras has been shown to be involved in multiple
biological functions: the ability to transform NIH 3T3
cells, the promotion of cell adhesion, and the regulation of
apoptotic response in hematopoietic cells (128, 621, 695,
816). Unlike other Ras family members, R-Ras does not
activate Raf proteins or MAP kinases in cells, whereas it
stimulates PKB/Akt effectively through PI 3-kinase (444).
TC21 has a highly oncogenic potential and is found mutated in some human tumors and overexpressed in breast
cancer (37, 94, 300). As to the activation of Raf proteins by
TC21, it is controversial (242, 611). Recently, new members of the Ras family, Rit and Rin, have been identified by
an expression cloning screen (385). Rit is ubiquitously
expressed, whereas Rin is expressed only in neural tissue.
A unique feature of their structures is that they lack a
known recognition signal for COOH-terminal prenylation.
Nonetheless, both proteins localize on the plasma membrane, probably through a COOH-terminal cluster of basic
amino acids. Rin binds calmodulin through a COOH-terminal motif, suggesting that Rin may be involved in calcium-mediated signaling in neurons (385). Rad is another
member of Ras-like proteins that has originally been isolated as a gene overexpressed in the skeletal muscle of
humans with type II diabetes (594). Kir/Gem has also been
cloned as a gene that is overexpressed in cells transformed by abl tyrosine kinase (123) or cloned from mitogen-induced human peripheral blood T cells (429). Kir/
Gem and Rad constitute a new family of Ras-related
proteins. The distinct structural features of this family
include the G3 GTP-binding motif, extensive NH2- and
COOH-terminal extensions beyond the Ras-related domain, and a motif that determines membrane association
(429). Rheb, another Ras protein-related molecule, has
been isolated by differential cloning techniques to identify
genes that are rapidly induced in brain neurons by synaptic activity (790). Expression of Rheb is rapidly and
transiently induced in hippocampal granule cells by seizures and by NMDA-dependent synaptic activity (790).
The amino acid sequence of Rheb is most closely homologous to yeast Ras1 and human Rap2. In the developing
brain, Rheb mRNA is expressed at relatively high levels.
Its close homology with Ras proteins and its rapid inducibility by receptor-dependent synaptic activity suggest
that Rheb may play an important role in long-term activity-dependent neuronal responses (790). More recently,
Ras protein-like proteins, named B-Ras1 and B-Ras2,
have been identified (181). These proteins interact with
IB and IB, which are inhibitors for the nuclear transcription factor kappa B, NF-B, and decrease the rate of
degradation of IBs. In cells, B-Ras proteins are associated only with NF-B:IB complexes and therefore may
provide an explanation for the slower rate of degradation
of IB compared with IB (181).

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SMALL G PROTEINS

IV. RHO/RAC/CDC42 PROTEINS AS

REGULATORS OF BOTH CYTOSKELETAL
REORGANIZATION AND GENE EXPRESSION
A. Outline
The mammalian Rho family consists of at least 14
distinct members as shown in Table 1 and Figure 1. The
function of the Rho family was first demonstrated in yeast
(5, 45, 320). Phenotypes of the mutants that carry mutations in these genes indicated that Rho/Cdc42 proteins are
involved in the budding process, presumably through reorganization of the actin cytoskeleton (320, 798). In mammals, the function of Rac proteins was the first to be
clarified. GTP-Rac1, in addition to two other cytosolic
proteins, p47phox and p67phox, were shown to be required for the activation of NADPH oxidase of phagocytic
cells (1, 2, 18, 358, 479, 649). Then, the function of mammalian Rho proteins was elucidated by use of an exoenzyme of Clostridium botulinum, named C3, that specifically ADP-ribosylates Rho proteins (6, 343, 512). C3 ADPribosylates an amino acid (Asn-41) in the effector region
of RhoA and inhibits its function by preventing interaction
with downstream effectors (651). By the use of C3, Rho
proteins were first suggested to be involved in cytoskeletal control (97, 564). Rho proteins were subsequently
shown to regulate formation of stress fibers and focal
adhesions in fibroblasts by use of its dominant active
mutant and Rho GDI (475, 600, 601) and to regulate Ca2
sensitivity of smooth muscle contraction (276). In contrast, Rac and Cdc42 proteins regulate formation of lamellipodia and filopodia, respectively (368, 522, 602). It has
now been established that at least Rho/Rac/Cdc42 proteins regulate primarily cytoskeletal reorganization in response to extracellular signals in mammalian cells. Evidence has also accumulated that they may play additional
roles in gene expression (126, 272, 473, 567, 692, 776).
Furthermore, involvement of Rho/Rac/Cdc42 proteins in
diverse cellular events, such as cell growth (341, 420, 552,
581583, 794), membrane trafficking (4, 69, 86, 365, 380),
development (227), and axon guidance (412) and extension (277, 314, 369), have been reported. In these cellular
events, it is not known whether Rho/Rac/Cdc42 proteins
directly regulate them or indirectly regulate them through
cytoskeletal reorganization and gene expression. Many
upstream regulators and downstream effectors have been
identified for Rho/Rac/Cdc42 proteins, and although their
modes of activation and action have gradually been elucidated, our understanding remains incomplete. Posttranslational modifications of Rho proteins are also crucial for their various functions including cell shape
change, cell motility, cytoplasmic division of Xenopus
embryo, and regulation of 1,3--glucan synthase of S.
cerevisiae (305, 352, 475, 705).

165

B. Reorganization of the Actin Cytoskeleton

Reorganization of the actin cytoskeleton plays crucial roles in many cellular functions such as cell shape
change, cell motility, cell adhesion, and cytokinesis. The
actin cytoskeleton is composed of actin filaments and
many specialized actin-binding proteins (671, 688, 826).
Filamentous actin is generally organized into a number of
discrete structures (Fig. 6): 1) actin stress fibers: bundles
of actin filaments that traverse the cell and are linked to
the extracellular matrix through focal adhesions; 2) lamellipodia: thin protrusive actin sheets that dominate the
edges of cultured fibroblasts and many migrating cells;
membrane ruffles observed at the leading edge of the cell
result from lamellipodia that lift up off the substratum and
fold backward; and 3) filopodia: fingerlike protrusions
that contain a tight bundle of long actin filaments in the
direction of the protrusion. They are found primarily in
motile cells and neuronal growth cones. It is important,
therefore, that the polymerization and depolymerization
of cortical actin be tightly regulated. For the most part,
this regulation of actin polymerization is orchestrated by
Rho/Rac/Cdc42 proteins. Rho proteins regulate stress fiber formation (475, 600), while Rac proteins regulate
ruffling and lamellipodia formation (602), and Cdc42 regulates filopodium formation (368, 522).
C. Rho/Rac/Cdc42 Protein Cycle: Cyclical
Activation/Inactivation
The activation and inactivation of Rho/Rac/Cdc42
proteins are regulated by essentially the same mechanism
as Ras proteins by GEPs and GAPs, respectively. However, they are further regulated by another class of regulator, GDIs (206, 275, 280, 704, 732). In the cytosol, Rho/
Rac/Cdc42 proteins are complexed with the GDI and
maintained in the GDP-bound inactive form. The GDPbound form is first released from a GDI by a still unknown
mechanism and is converted to the GTP-bound form by
the action of a GEP. The GTP-bound form then interacts
with the downstream effector(s). Thereafter, the GTPbound form is converted to the GDP-bound form by the
action of a GAP. The GDP-bound form then forms a
complex with the GDI and returns to the cytosol.
Rho/Rac/Cdc42 proteins are posttranslationally modified with lipid as described above and therefore they have
to be in complex with GDIs to remain soluble in the
cytosol. However, it is unknown whether all the GDPbound form of Rho/Rac/Cdc42 proteins are complexed
with GDIs and remain in the cytosol. Some amount of the
GDP-bound form may be associated with membranes, and
it may be converted to the GTP-bound form and exert its
function on the membrane. In this case, GDIs would not
be essential for their cyclical activation and inactivation.

166

TAKAI, SASAKI, AND MATOZAKI

Volume 81

FIG. 6. Mode of action of Rho/Rac/Cdc42 proteins in cytoskeletal reorganization. A: mode of action of Rho proteins.
B: mode of action of Rac proteins. C: mode of action of Cdc42. Rho, Rho proteins; Rac, Rac proteins.

Many GEPs for Rho/Rac/Cdc42 proteins have been

isolated and characterized as shown in Table 2 and numbers are still increasing. Most GEPs have been isolated as
oncogenes. The GEPs thus far identified share a common
motif, designated the Dbl-homology (DH) domain, for
which the Dbl oncogene product is the prototype (93).
Biochemical analysis has confirmed that DH domains of
GEPs indeed show GEP activity on Rho/Rac/Cdc42 proteins in a cell-free assay system (259, 262, 468). In addition
to the DH domain, GEPs share a PH domain, which may
be involved in proper cellular localization presumably
through interaction with PIP2 (469, 824). Some members
of GEPs, such as Dbl and Vav1, have been shown to
exhibit exchange activity in vitro for a broad range including Rho/Rac/Cdc42 proteins, whereas others appear to be
more specific. Lbc, for example, and more recently discovered oncoproteins Lfc and Lsc are specific for Rho
proteins (226, 823), whereas FGD1 and frabin are specific
for Cdc42 (532, 738, 822). Although Vav1 is a GEP for Rac
proteins (133, 245), Vav2, a GEP closely related to Vav1,
functions preferably as a GEP for Rho proteins (642, 643).
Some GEPs, such as Dbl, prefer the lipid-modified form of
the substrate small G proteins to the lipid-unmodified
ones (788).
In addition to the PH and DH domains, many GEPs
have other domains that are commonly found in signaling
molecules, such as the SH2 domain for Vav or the SH3
domain for Vav and Dbs, suggesting that they may have

additional functions (93, 133, 642, 778). A GEP for Rho

proteins, named p115 Rho GEF, that contains the regulator of G protein (RGS) domain has recently been identified (262, 367). RGS stimulates the intrinsic GTPase activity of the -subunit of G12 and G13. p115 Rho GEF acts
as an intermediary in the regulation of Rho proteins by
G12 and G13 (260, 367). In addition, another Rho GEP
(named PDZ-Rho GEF) that contains RGS and PDZ domains has been reported (204). These findings have provided a new model for a signaling pathway for Rho proteins from membrane receptors. Recently, SHP-2, a
protein tyrosine phosphatase containing SH2 domains,
has been demonstrated to suppress the activity of Vav2
and consequently to reduce the Rhos ability to form
stress fibers and focal adhesions (360). SHP-2 thereby
positively regulates the hepatocyte growth factor (HGF)/
scatter factor (SF)-induced cell scattering. However, detailed information regarding signaling cascades coupling
the extracellular stimuli to activation of GEPs for Rac/
Cdc42 proteins is still limited. Some GEPs like Tiam1
(248) and Ras GRF (667) carry a second PH domain. For
Tiam1 and Ras GEF, this second, NH2-terminal PH domain mediates localization to cell membranes (74, 469). In
addition to the PH domain, frabin has an actin-binding
domain at its NH2-terminal region (532). The actin-binding
domain in addition to the DH and first PH domains is
essential for the filopodium formation mediated by frabin
through Cdc42 (532, 738). Frabin furthermore induces

SMALL G PROTEINS

January 2001

lamellipoidum formation through indirect activation of

Rac (553). The COOH-terminal FYVE and second PH domains, which associate with an unidentified membrane
structure, in addition to the DH and first PH domains are
necessary for this action (553).
The first GAP protein specific for Rho proteins was
biochemically purified from human spleen and bovine
adrenal gland (211, 212, 488). This protein, designated p50
Rho GAP, has GAP activity toward Rho/Rac/Cdc42 proteins in vitro (381). A number of proteins that exhibit GAP
activity for Rho/Rac/Cdc42 proteins have subsequently
been identified in mammalian cells (Table 2). These proteins all share a related GAP domain that spans 140
amino acids of the protein but bears no significant resemblance to Ras GAP. The substrate specificity of Rho GAPs
toward Rho/Rac/Cdc42 proteins varies with each GAP
protein. Although some of these proteins exhibit GAP
activity for several small G proteins in cell-free assay
systems, their substrate specificities in vivo appear to be
more restricted. For instance, the substrate spectrum of
p50 Rho GAP in vitro encompasses Rho/Rac/Cdc42 proteins; however, in vivo, it appears to be restricted to Rho
proteins only (603). Although first identified as a tyrosinephosphorylated p120 Ras GAP-associated protein in Srctransformed cells and in growth factor-treated cells (169,
657), p190 Rho GAP was later shown to possess GAP
activity for Rho proteins (656). Although the biological
function of p190 Rho GAP is not well understood, the
interaction of p190 Rho GAP with p120 Ras GAP has been
suggested to induce a conformational change in p120 Ras
TABLE

2. Regulators of Rho/Rac/Cdc42 proteins

Substrate

GEP
Dbl
Vav1
Dbs
Lbc
Lfc
Lsc
Vav2
p115 Rho GEF
PDZ-Rho GEF
Tiam-1
FGD1
Frabin
GAP
p50 Rho GAP
p190 Rho GAP
Graf
myr5
Bcr
n-Chimaerin
3BP-1
Abr
GDI
Rho GDI
D4/Ly-GDI
Rho GDI-3

Reference No.

Rho, Rac, Cdc42

Rho, Rac, Cdc42
Rho, Cdc42
Rho
Rho
Rho
Rho
Rho
Rho
Rac
Cdc42
Cdc42

259, 680
245
778
823
226
226
269
262
204
248
822
532, 738

Rho, Rac, Cdc42

Rho, Rac, Cdc42
Rho, Cdc42
Rho, Cdc42
Rac, Cdc42
Rac, Cdc42
Rac
Rac

36, 381
657
271
589
152
152
118
709

Rho, Rac, Cdc42

Rho
RhoB (not RhoA, RhoC)

206, 732
390, 635
809

167

GAP, resulting in increased accessibility of the effector

binding surface of its SH3 domain (298). A role for p190
Rho GAP in regulating Rho protein function in cells undergoing cytoskeletal rearrangements has been suggested
(95, 603), but it is not known whether this effect is induced by p190 Rho GAP as a downstream effector of Rho
proteins.
Recent studies have shown that a cycle of inactivation and activation of Rho/Rac/Cdc42 proteins is necessary for dynamic cell functions such as growth factorinduced cell scattering. Expression of dominant active
mutants of Rho/Rac/Cdc42 proteins inhibits HGF/SF-induced cell scattering (303, 331, 599), whereas C3 or Rho
GDI blocks HGF/SF-induced cell scattering (706). The
mode of action of Rho proteins in cell scattering remains
to be clarified, but the Rho protein-regulated assembly
and disassembly of stress fibers and focal adhesions have
been suggested to be, at least in part, involved in this
process (303, 331, 599, 706). It is not known how inactivation by GAPs is induced. In one case, integrin-induced
formation of stress fibers inhibits Rho protein activation
as part of a feedback inhibition system (593).
Rho GDI was originally isolated as a cytosolic protein
that preferentially associated with GDP-RhoA and GDPRhoB and thereby inhibited the dissociation of GDP (206,
732). Rho GDI requires the posttranslational lipid modifications of RhoA for its activity (286). Rho GDI prefers
GDP-RhoA and GDP-RhoB to the corresponding GTPbound forms and forms a ternary complex with the GDPbound form (628, 732). Rho GDI is also capable of inhibiting GTP hydrolysis by Rho proteins (116, 261, 628),
blocking both intrinsic and GAP-catalyzed GTPase activity. Rho GDI dissociates the GDP-bound form of prenylated RhoB from the membrane (308). Based on these
properties of Rho GDI, it has been proposed that Rho GDI
is involved not only in the regulation of the activation of
Rho proteins but also in their translocation between the
cytosol and the membrane (630, 702, 704). The GDPbound forms of Rho proteins are complexed with Rho
GDI and remain in the cytosol. When the GDP-bound form
is released from Rho GDI, it is converted to the GTPbound form by the action of Rho GEPs. The GTP-bound
form then activates its specific downstream effector(s)
until the GTP-bound form is converted to the GDP-bound
form by Rho GAPs. Once the GDP-bound form is produced on the membrane, it is captured by Rho GDI and
the complex returns to the cytosol.
Rho GDI has also been shown to be active not only
on Rho proteins but also on Rac/Cdc42 proteins (18, 275,
391). In addition to Rho GDI, at least two other isoforms,
named D4/Ly-GDI and Rho GDI-3, have been identified
(390, 635, 809). Now, the originally identified Rho GDI is
referred to as Rho GDI or Rho GDI1; D4/Ly-GDI is
named Rho GDI or Rho GDI2; and Rho GDI3 is named
Rho GDI or Rho GDI3. Recent NMR studies have shown

168

TAKAI, SASAKI, AND MATOZAKI

that Rho GDI has a pocket that masks the lipid moieties
of Rho proteins (238), consistent with biochemical analyses (286). More recently, the X-ray crystallographic
structure of the Cdc42/Rho GDI complex has revealed
two major sites of interaction between Rho GDI and
Cdc42 (280). The NH2-terminal regulatory region of Rho
GDI binds to the switch I and II domains of Cdc42, leading
to inhibition of both GDP dissociation and GTP hydrolysis. In addition, the geranylgeranyl moiety of Cdc42 inserts into a hydrophobic pocket within the immunoglobulin-like domain of the GDI molecule, keeping GDP-Cdc42
in the cytosol and permitting the dissociation of GDPCdc42 from membranes.
Although the mechanism by which Rho proteins are
released from Rho GDI has largely been unknown, it has
been shown that ERM, which consists of ezrin, radaxin,
and moesin, has the capacity to displace the GDP-bound
form of Rho proteins from Rho GDI (700). ERM has two
functional domains; the NH2-terminal plasma membranebinding and COOH-terminal F-actin-interacting domains
(30, 730). ERM is translocated to the plasma membrane
probably through the interaction with the cytoplasmic
domain of integral plasma membrane proteins such as
CD44, providing the F-actin binding sites. Rho GDI interacts with the NH2-terminal fragments of ERM, and this
interaction reduces Rho GDI activity (700). The unfolding of ERM may induce Rho GDI interaction with ERM
NH2-terminal regions, causing release of the Rho-GDP
and making it sensitive to the action of each Rho GEP
(699, 700). The interaction of full-length ERM with the
cytoplasmic fragment of CD44 does not induce the association of ERM with Rho GDI (700). However, CD44 and
Rho GDI are coimmunoprecipitated with moesin (274).
It is likely that full-length ERM unfold to permit interactions with Rho GDI and CD44 via ERM NH2-terminal
regions and F-actin via ERM COOH-terminal regions; a
signal from CD44 may trigger this process. These findings
suggest that the activation of Rho proteins is regulated in
a temporally and spatially dynamic manner and is distinct
from that of Ras proteins, which appears to be regulated
in a unidirectional manner.
Little is known about the physiological function of
Rho GDIs in vivo, but microinjection studies have shown
that Rho GDI inhibits several downstream functions of
Rho proteins (303, 352, 475, 521, 630, 705, 706). Rho
GDI-deficient mice have revealed several abnormal phenotypes (722). These mice are initially viable, but they
develop massive proteinuria mimicking nephrotic syndrome in humans, leading to death due to renal failure
within a year. Histologically, degeneration of tubular epithelial cells and dilatation of distal and collecting tubules
are readily detected in the kidneys. Rho GDI-deficient
mice are also infertile and show impaired spermatogenesis with vacuolar degeneration of seminiferous tubules. In
contrast, Rho GDI-deficient mice show normal hemato-

Volume 81

poietic differentiation but subtle defects in superoxide

production by macrophages derived from in vitro embryonal stem cell differentiation (244). Thus these two lines
of evidence suggest that Rho GDIs play physiologically
important roles.
Yeast Rom7/Bem4 is a novel type of Rho1- and
Cdc42-related molecule (273, 419). This protein appears
to be distinct from GEPs, GDIs, and GAPs, in the sense
that it interacts genetically with the Rho1 pathway but
does not show GEP, GDI, or GAP activity, and it is not a
downstream effector.
D. Mode of Action of Rho Proteins
in Cytoskeletal Reorganization
Mammalian Rho proteins are required for many actin
cytoskeleton-dependent cellular processes, such as platelet aggregation, lymphocyte and fibroblast adhesion, cell
motility, contraction, and cytokinesis (251, 704). In the
yeast S. cerevisiae, Rho proteins, including Rho1, Rho2,
Rho3, and Rho4, regulate budding and cytokinesis
through reorganization of the actin cytoskeleton (80, 119,
200, 713). The mode of action of Rho proteins was first
clarified for Rho1 in the yeast budding process. Pkc1, a
yeast homolog of mammalian protein kinase C, was first
shown to be a downstream effector of Rho1 (330, 525).
Pkc1 regulates gene expression through the MAP kinase
cascade, consisting of Bck1 (MAP kinase kinase kinase),
Mkk1/Mkk2 (MAP kinase kinase), and Mpk1 (MAP kinase) (173, 396). This MAP kinase cascade regulates expression of the genes necessary for cell wall integrity.
Bni1 is another effector of Rho1 (363). Bni1 also interacts
with GTP-Rho3, GTP-Rho4 (T. Kamei and Y. Takai, unpublished observations), and GTP-Cdc42 (175). Bni1 has
two domains, named formin homology (FH)1 and FH2
domains, which are found in a variety of proteins involved
in cytoskeletal rearrangement needed to achieve cell polarity and cytokinesis (196). Bni1 binds the actin monomer-binding protein profilin via its FH1 domain to regulate reorganization of the actin cytoskeleton (175, 363).
Furthermore, Bni1 binds Aip3 (Bud6), another actin-binding protein (15), and elongation factor 2, which is known
to stimulate actin polymerization (739), suggesting that
Bni1 is the downstream effector of Rho1 that directly
regulates reorganization of the actin cytoskeleton. More
recently, Bni1 has also been shown to participate in microtubule function, since disruption of BNI1 causes defects in spindle orientation (386), Kar9 localization (472),
and growth defect together with mutation either PAC1
and NIP100 (199), whose gene products are implicated in
microtubule function (215).
1,3--Glucan synthase is the third effector of Rho1
(157, 456, 580). This enzyme synthesizes 1,3--glucan, a
major component of the cell wall. This series of experi-

ments has established that Rho proteins regulate complicated cell functions through multiple effectors in a cooperative manner. Another protein having FH domains has
also been found in yeast and named Bnr1 (304). This
protein serves as an effector of Rho4 and binds both
profilin and Aip3 (Bud6), indicating that Rho4 regulates
the actin cytoskeleton at least though Bnr1. Rho2 has 53%
identity to Rho1, and a RHO2 null mutant does not show
any growth defect (422). Hence, it has been assumed that
Rho2 has redundant functions with Rho1, although several lines of evidence suggest Rho2-specific functions
(149, 436). Rho3 and Rho4 are implicated in polarized
growth presumably through regulating the actin cytoskeleton via their interactions with Bni1 and Bnr1 (175, 331a).
Moreover, it has been shown that Rho3 interacts with
Myo2 (type V myosin) and Exo70 (a component of the
exocyst, a multiprotein complex which is involved in
exocytosis) (606), suggesting the involvement of Rho3 in
the polarized secretion.
Numerous downstream effectors of mammalian Rho
proteins have been identified (Table 3). p160ROCK, also
named ROK/Rho kinase, is a recently identified, serine/
threonine protein kinase (307, 395, 451) and a downstream effector of Rho proteins. The activity of this protein kinase is stimulated by GTP-Rho proteins. Many
ROCK/Rho kinase substrates have been identified; these
include the myosin binding subunit of myosin light-chain
phosphatase (347), myosin light chain (13), ERM (202),
and cofilin (425). Of these substrates, myosin light-chain
phosphatase appears to be a physiological substrate
(347). Guanosine 5-O-(3-thiotriphosphate) (GTPS), a
poorly hydrolyzable GTP analog, increases the sensitivity
of smooth muscle contraction to Ca2 (354, 487, 785).
TABLE

This action of GTPS is inhibited by C3 or an exoenzyme

of Staphylococcus aureus, named EDIN, which also inhibits the function of Rho proteins, and it is mimicked by
GTPS-RhoA, but not by GDP-RhoA, indicating that Rho
proteins are involved in GTPS-induced Ca2 sensitization of smooth muscle contraction (276). Subsequently,
myosin light-chain phosphatase has been shown to be a
downstream effector of Rho proteins (232, 524). ROCK/
Rho kinase has finally been shown to phosphorylate and
to inhibit myosin light-chain phosphatase, causing the
sustained contraction of smooth muscle contraction even
after decrease in Ca2 concentrations (347). In this mode
of action, Rho proteins do not induce smooth muscle
contraction without Ca2 triggering; rather, they modify
Ca2 sensitivity of smooth muscle contraction. In this
sense, the physiological relevance of the ROCK/Rho kinase-induced direct phosphorylation of the myosin light
chain and the subsequent activation of the myosin ATPase
could not be considered (13). The physiological relevance
of the phosphorylation of other substrate proteins, including moesin, is also controversial. ROCK/Rho kinase phosphorylates moesin, which induces microvilli formation
(202), but another report indicates that ROCK/Rho kinase
is not involved in this phosphorylation (452). p140mDia, a
mammalian homolog of Bni1 and Bnr1 in the yeast S.
cerevisiae (304, 363) and that of Drosophila diaphanous
(92), has also been implicated as a downstream effector
of Rho proteins (770). mDia has FH1 and FH2 domains.
mDia as well as Bni1 and Bnr1 bind profilin via its FH1
domain to regulate reorganization of the actin cytoskeleton. Overexpression of mDia induces weak formation of
stress fibers without affecting the formation of focal adhesions (510, 769). ROCK and mDia cooperatively regu-

3. Effectors of Rho/Rac/Cdc42 proteins

Rho Proteins
Effectors

Mammal

Yeast

169

SMALL G PROTEINS

January 2001

PI 3-kinase
Phospholipase D
PI 4,5-kinase
Citron
ROK/ROCK/
Rho kinase
MBS
PKN
Rhophilin
Kinectin
Rhotekin
p140mDia
DGK
Pkc1
Fks1, 2,
(glucan synthase)
Bni1
Bnr1

PI, phosphatidylinositol.

Rac Proteins

Cdc42

Reference no.

Effectors

Reference no.

Effectors

Reference no.

812
431
112, 592
423
307, 393, 451

NADPH oxidase
PAK
PI 3-kinase
PI 4,5-kinase
IQGAP
POR1
S6-kinase
MLK2, 3
Sra-1
POSH
DGK

2, 358, 479
439
57, 724
263, 724
65, 374, 458
745
113
502
359
716
725

ACK
PI 3-kinase
PAK
WASP
S6-kinase
IQGAP
MLK2, 3
N-WASP
MRCK
MSE5
Borg

438
820
439
31, 698
113
65, 374, 458
502
470
394
76
318

Ste20
Cla4
Bni1
Skm1
Gic1, 2

819
137
175, 363
446, 652
106

347
14, 768
768
292
588
770
293
330, 525
580, 456
363
304

170

TAKAI, SASAKI, AND MATOZAKI

late the Rho protein-induced reorganization of the actin

cytoskeleton (510, 769) (Fig. 6A). ROCK is part of another
downstream cascade of Rho proteins, a Rho-ROCK-LIM
kinase pathway (425). ROCK directly phosphorylates LIM
kinase, which in turn is activated to phosphorylate cofilin.
Cofilin exhibits actin-depolymerizing activity that is inhibited as a result of its phosphorylation by LIM kinase (425,
693). Overexpression of LIM kinase induces the formation
of actin stress fibers in a ROCK-dependent manner. Thus
phosphorylation of LIM kinase by ROCK and subsequent
increased phosphorylation of cofilin by LIM kinase contribute to the Rho protein-induced reorganization of the
actin cytoskeleton.
In addition to ROCK, a variety of Rho effector proteins have recently been discovered (Table 3). These include serine/threonine protein kinase PKN (14, 768),
citron (424), rhotekin (588), and rhophilin (768). Citron
contains a protein kinase domain that is related to ROCK.
Citron kinase localizes to the cleavage furrow and midbody of cultured cells (423). Overexpression of citron
mutants results in the production of multinucleate cells,
and a kinase-active mutant causes abnormal contraction
during cytokinesis, suggesting that citron kinase is involved in the Rho protein-regulated cytokinesis (423).
E. Mode of Action of Rac/Cdc42 Proteins
in Cytoskeletal Reorganization
The function of Rac/Cdc42 proteins in cytoskeletal
reorganization was first demonstrated in the yeast. In the
yeast S. cerevisiae, Cdc42 participates in recognition of
the landmark that defines the incipient bud site (5, 320,
827). In fibroblasts, Rac proteins regulate formation of
lamellipodia and membrane ruffles and subsequent stress
fiber formation (602). In contrast, Cdc42 plays a key role
in the formation of filopodia at the cell periphery followed
by the formation of lamellipodia and membrane ruffles
(368, 522). Both Rac/Cdc42 proteins induce the assembly
of multimolecular focal complexes at the plasma membrane of fibroblasts (522). In addition, Rac/Cdc42 proteins
regulate the cadherin-based cell-cell adhesion and hence
control the HGF- and 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced cell scattering of Madin-Darby canine
kidney cells (284, 324, 331, 361, 624, 686, 707). Cdc42 may
play a role in the polymerization of both actin and microtubules toward antigen-presenting cells (689). Furthermore, the involvement of Cdc42 in the control of cytokinesis in HeLa cells and Xenopus embryos has been
reported (156, 164). In Drosophila, Rac/Cdc42 proteins
are implicated in axonal guidance, wing hair formation,
actomyosin-driven furrow canal formation, and nuclear
positioning (130, 165, 214).
To date, several potential effectors of Cdc42 and Rac
proteins have been identified (Table 3). Several of these

Volume 81

proteins are common effectors for both Rac and Cdc42

proteins. Among them, the family of serine/threonine protein kinases known as PAKs are especially interesting.
Homologs of the mammalian PAKs have been identified in
S. cerevisiae (Ste20 and Cla4) (137, 384), S. pombe (Pak1,
also called Shk1) (443, 556), Drosophila (PAK1) (258),
and C. elegans (Ste20) (108). In the yeast S. cerevisiae,
Cdc42 interacts with Ste20, which in turn associates with
Ste5 and Bem1, both of which interact with actin (387,
821). Drosophila PAK1 also plays a role in dorsal closure
(258). Thus it is likely that PAK proteins are involved in
mediating the effect of Cdc42/Rac proteins on the cytoskeleton. Most recently, Drosophila Trio, a GEP for Rac
proteins, has been shown to play an important role in
axon guidance (32, 43, 403, 515). One of the two Trio GEP
domains is critical in photoreceptor axon guidance (515).
This GEP domain activates Rac, which in turn activates
PAK. Trio interacts genetically with Rac proteins, PAK,
and DOCK, an SH2-SH3 docking protein. Thus Trio regulates Rac proteins, which subsequently activates PAK,
linking guidance receptors to the growth cone cytoskeleton (515). However, in mammalian cells, the role of PAKs
remains unclear (Fig. 6B). Expression of a mutant form of
Rac or Cdc42 protein that is unable to bind and activate
PAKs can still induce the formation of membrane ruffles
and lamellipodia (323, 379), indicating that PAKs are not
essential for the Rac protein-elicited membrane ruffling
and lamellipodium formation or for the Cdc42-triggered
filopodium formation. This does not exclude the possibility, however, that PAKs themselves may play a role in
cytoskeletal rearrangement by inducing actin reorganization independently of Rac/Cdc42 proteins. Alternatively,
PAKs may mediate effects on the cytoskeleton induced by
Rac/Cdc42 proteins, which are different from the immediate actin reorganization (437, 653).
All PAKs identified to date share a similar 18-amino
acid CRIB (Cdc42/Rac interactive binding) motif that mediates the interaction with Rac/Cdc42 proteins. Most recently, N-WASP, a ubiquitously expressed Cdc42-interacting protein (470), and the Arp2/3 complex have been
shown to participate in the downstream cascade of
Cdc42-induced actin polymerization (49, 417, 418, 610,
697, 775, 782) (Fig. 6C). Wiskott-Aldrich syndrome protein
(WASP), which is only expressed in hematopoietic cells,
was originally identified as a protein mutated in patients
with Wiskott-Aldrich syndrome (31, 698). WASP and NWASP (470) possess a PH domain that binds PIP2 and a
CRIB domain. The binding of both GTP-Cdc42 and PIP2 to
N-WASP activates N-WASP by stabilizing the active conformation of this molecule. The COOH terminus of NWASP binds the Arp2/3 complex and consequently stimulates its ability to nucleate actin polymerization in vitro
(49, 415, 417, 418, 610, 697, 775, 782). Therefore, the
interaction of N-WASP with the Arp2/3 complex is a core
mechanism that directly connects the Cdc42-mediated

January 2001

SMALL G PROTEINS

signal transduction pathway to the stimulation of actin

polymerization.
In addition to the CRIB-containing proteins, a 34-kDa
protein, POR1 (partner of Rac), interacts specifically with
GTP-Rac proteins and is implicated in mediating Rac
protein-dependent membrane ruffling (745). A dominant
active mutant of Rac1 (Rac1V12L37) that fails to bind
POR1 also fails to induce membrane ruffling (323). Arf6 is
also implicated downstream of Rac proteins and mediates
membrane ruffling through PIP2 kinase (283). Arf6 and
POR1 function on distinct signaling pathways to mediate
cytoskeletal reorganization induced by Rac proteins (Fig.
6B). Another protein with a potential role in cytoskeletal
organization is IQGAP (65, 374, 458). IQGAP interacts
with both GTP-Rac1 and GTP-Cdc42 and localizes to
membrane ruffles. IQGAP has been shown to be localized
to cell-cell adhesion sites (375). It has been suggested that
activated Cdc42 blocks the ability of IQGAP to inhibit
asse bly of a cadherin-catenin complex and thereby promotes formation of adherens junctions (375), but this
model has not yet been substantiated. In addition, no
apparent abnormality is observed in the formation of adherens junctions in the IQGAP1-deficient mice (400). Although IQGAP contains some interesting motifs found in
signaling molecules such as a WW domain, an SH3-binding domain, a calmodulin-binding domain, and, somewhat
surprisingly, a Ras GAP-like motif, its function remains
unknown. Although a number of potential Rho family
effectors have been identified, a major goal for the future
will be to determine the physiological roles of these proteins in Rho family-mediated cytoskeletal rearrangement.
F. Mode of Action of Rho/Rac/Cdc42 Proteins
in Gene Expression
Whereas Ras proteins control the activation of the
p42/44 MAPK cascade, in certain cell types, Rac/Cdc42
proteins regulate the activation of Jun NH2-terminal kinase (JNK) and p38 MAP kinase (126, 473). Expression of
a constitutively active mutant of Rac or Cdc42 protein in
HeLa, NIH 3T3, and COS cells results in a stimulation of
JNK and p38 activities (126, 473). Furthermore, the same
effects have been observed with oncogenic GEPs for
Rho/Rac/Cdc42 proteins, such as Dbl (473). The exact role
of Rho/Rac/Cdc42 proteins in MAP kinase activation is,
however, unclear, and it is still largely unknown whether
physiological activation of the JNK pathway is induced by
endogenous Rac/Cdc42 protein activity. Genetic analysis
of dorsal closure in Drosophila also supports a role for
Rac proteins in JNK regulation (227).
Rho/Rac/Cdc42 proteins have each been reported to
activate serum response factor (SRF) (272, 776). They
have also been shown to activate the transcription factor
NFB (567, 692). In addition, generation of reactive oxy-

171

gen species by Rac proteins might be the trigger for NFB

activation. This observation may be related to the fact
that Rac proteins regulate an NADPH oxidase enzyme
complex in phagocytes to produce superoxide as described earlier (1, 2, 18, 358, 479, 649). Rho/Rac/Cdc42
proteins are required for G1 cell cycle progression (420,
552, 794), but it is unclear whether this is due to their
effects on the actin cytoskeleton and integrin adhesion
complexes or, instead, to more direct effects on gene
transcription.
G. Other Functions of Rho/Rac/Cdc42 Proteins
Various studies have suggested the involvement of
Rho/Rac/Cdc42 proteins in membrane-trafficking processes. Cytoskeletal rearrangements are closely coupled
to the onset of phagocytosis. Rho/Rac/cdc42 proteins are
implicated in one or more steps of the phagocytic response (86, 129, 449). Two distinct mechanisms for the
phagocytic response have recently been defined (8, 514).
In type I phagocytosis, plasma membrane protrusions
extend to engulf the particle and drag it into the cell; this
is mediated by coordinated actions of Rac/Cdc42 proteins
(86). In type II phagocytosis, particles sink into actin-lined
investigations in the plasma membrane; here, internalization is dependent on RhoA (86). There are morphological
similarities between these processes and the invasion of
mammalian cells by certain pathogenic bacteria. Shigella
invasion starts with actin nucleation and Rho proteininduced actin polymerization (4, 496). This is followed by
continued actin polymerization around membrane-bound
protrusions that fold over the bacterium and coalesce to
engulf it. The complete inhibition of Shigella-induced
membrane folding by C3 suggests that actin polymerization is essential for the generation of the surface extensions (4, 496). Interestingly, Cdc42 has been shown to
play a direct role in Salmonella internalization (107) and
in Shigella flexneri motility (696).
Rho/Rac proteins have also been implicated in the
regulation of endocytosis. In mammalian cells, expression
of dominant active mutants of Rho/Rac proteins reduces
the efficiency of endocytosis of the transferrin receptor
(380), and a dominant active mutant of RhoA also blocks
internalization of muscarinic acetylcholine receptors
(757). Expression of a dominant active mutant of RhoA
causes an increase in clathrin-independent endocytosis in
Xenopus oocytes (639). In Swiss 3T3 cells, expression of
a dominant active mutant of Rac proteins stimulates pinocytosis (602), and the pinocytic vesicles are coated
with the Rac signaling partner PAK1 (150). However,
expression of a dominant active Rac protein has no effect
on pinocytosis in baby hamster kidney cells (398); thus
the role of Rac proteins in pinocytosis is still unclear.
Rho/Rac proteins have also been implicated in the

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regulation of secretory vesicle transport (69, 555a). In

mast cells, Rho/Rac proteins stimulate the exocytosis of
secretory granules, whereas C3 and dominant-negative
mutants of Rho/Rac proteins inhibit GTPS-induced secretion (526). In PC12 cells, RhoA, Rac1, and Rho GDI are
also involved in Ca2-dependent exocytosis at least partly
through the reorganization of actin cytoskeleton, and the
activation of RhoA or Rac1 alone is not sufficient for this
reaction (365). RhoD and RhoB are localized to endocytic
vesicles, suggesting that they might perform roles in the
regulation of intracellular trafficking (209, 498).
Rho/Rac/Cdc42 proteins have been implicated in
morphogenesis by regulation of cytoskeletal rearrangements. For instance, the Drosophila homologs of Rac1,
Rac2, and Cdc42, namely, DRac1, DRac2, and DCdc42,
respectively, are highly expressed in the nervous system
and in the mesoderm during development, respectively
(411, 412). Particularly, the inhibition of axonal outgrowth
induced by a dominant active or dominant negative mutant of DRac1 is due to different cytoskeletal defects in
developing neurons, suggesting that cyclical activation
and inactivation of DRac1 is crucial for axonal development (214, 412). In addition, expression of a dominant
active mutant of DCdc42, DCdc42V12, in the neurons of
fly embryos results in a similar but qualitatively different effect on neuronal development, compared with
DRac1V12 (412). In mammalian N1E-115 neuroblastoma
cells, Rho proteins appear to inhibit neurite outgrowth,
whereas Rac/Cdc42 proteins promote neurite outgrowth,
presumably by regulating the formation of growth cone
filopodia and lamellipodia (277, 369). In contrast, in chick
dorsal root ganglion neurons, Rho proteins induce neurite
outgrowth, while an activated mutant of Rac proteins
increases the proportion of collapsed growth cones (317).
DRac1 and DCdc42 have also been implicated in muscle
differentiation (172, 412). Rac/Cdc42 proteins also regulate muscle development, probably via their effects on
membrane fusion and the actin cytoskeleton. Finally, little is known at present about the role of Rho proteins in
neuronal and muscle development. In summary, Rho/Rac/
Cdc42 proteins have multiple essential functions during
morphogenesis.

Volume 81

teins regulate intracellular vesicle trafficking was first

obtained genetically in the yeast S. cerevisiae. Many
genes essential for secretion were isolated and named the
SEC genes (527). One of the SEC genes, the SEC4 gene,
was shown to encode a small G protein that is required for
vesicle trafficking from the Golgi apparatus to the plasma
membrane (623). The yeast YPT1 gene was first discovered as a gene located between the actin and tubulin
genes in 1983 (208) and was later identified to encode a
small G protein (641). Ypt1 was shown to be later involved in vesicle trafficking from the endoplasmic reticulum to the Golgi apparatus (33, 650). Subsequently, a large
body of evidence has accumulated in support of a role for
Rab proteins in vesicle trafficking from yeast to human
(447, 528, 529, 550, 636).
B. Vesicle Trafficking
Transmembrane proteins and secreted, soluble proteins are transported from one membrane compartment
to another by vesicles (465, 579, 616) (Fig. 7). Newly
synthesized secretory proteins are translocated into the
endoplasmic reticulum and are then transported to the
plasma membrane via the Golgi apparatus by vesicles;
some proteins are delivered to prelysosomes. In parallel,
macromolecules that are taken up from the plasma membrane are transported inward to endosomes and lysosomes by vesicles. Some cell-surface proteins including
receptors for extracellular ligands transit through a recycling endosome and are recycled back to the plasma
membrane. Thus exocytosis, endocytosis, and recycling
are performed by intracellular vesicle trafficking. There
are two exocytotic pathways: regulated and constitutive
pathways. In the regulated pathway, secretory proteins
are stored at very high concentrations in the core of

V. RAB PROTEINS AS REGULATORS

OF VESICLE TRAFFICKING
A. Outline
Rab proteins exist in all eukaryotic cells and form the
largest branch of the small G protein superfamily (103,
447, 528, 529, 550, 636). The yeast S. cerevisiae genome
sequence encodes 11 Rab proteins (383). In mammalian
cells, over 50 Rab proteins (including isoforms) are
known (447, 528, 529, 550, 636). Evidence that Rab pro-

FIG. 7. Principal mechanism of intracellular vesicle trafficking. Rab,

Rab proteins; Sar1/Arf, Sar1/Arf proteins.

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SMALL G PROTEINS

secretory granules that can be mobilized by an exocytosis

pathway in response to neural or hormonal stimuli. In
contrast, constitutive exocytosis occurs independently of
extracellular stimuli and proceeds at a steady rate. Constitutive exocytosis is responsible for the insertion of
plasma membrane proteins and secretion of extracellular
proteins, such as matrix components and plasma proteins. Vesicle trafficking is needed for various other cell
functions, such as establishment of cell polarity, cytokinesis, and cell motility.
There are at least four principal events in a given step
of intracellular vesicle transport: 1) budding of a vesicle
from the donor membrane; 2) targeting of the vesicle to
the acceptor membrane; 3) docking of the vesicle to the
acceptor membrane; and 4) fusion of the vesicle with the
acceptor membrane (Fig. 7). Rab proteins regulate these
processes (447, 528, 529, 550, 636). Substantial evidence
has accumulated that most Rab proteins regulate the
targeting/docking/fusion processes and that some of them

173

regulate the budding process, which is mainly regulated

by Sar1/Arf proteins (see sect. VI).
There are two types of Rab proteins: one type is
involved in regulated secretion, and the other type is
involved in other vesicle transport steps and is essential
for this function (447, 528, 529, 550, 636). Of the first type,
the mode of action of Rab3A has most extensively been
investigated (703). Rab3A plays a key regulatory role in
Ca2-dependent exocytosis, particularly in neurotransmitter release from nerve terminals. Studies on Rab3Adeficient mice have revealed that Rab3A is not essential
for basal neurotransmission but modulates it, thereby
contributing to synaptic plasticity (91, 216, 217, 218, 408).
In contrast, mutations of Rab proteins mediating other
vesicle transport steps can have more dramatic consequences. In the yeast S. cerevisiae (383), Rab mutants are
often characterized by a massive accumulation of the
vesicles in the respective secretory pathways (623). Moreover, some of them are essential for cell viability.

FIG. 8. Subcellular localization of Rab proteins. Plain,

localization of each Rab protein; italic, functioning site of
each Rab protein.

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C. Localization of Rab Proteins

Rab proteins contain unique, hypervariable COOHterminal domains with two Cys residues, both of which
require geranylgeranyl moieties by geranylgeranyltransferase type II as described above. This lipid modification
is essential for the membrane association of Rab proteins
(27, 322, 340, 349, 568, 765). The COOH-terminal hypervariable region is important for the correct targeting of
Rab proteins to their target membranes (64, 102), but how
it directs localization remains to be clarified.
At steady state, 10 50% of a given Rab protein is
detected in the cytosol. This pool is maintained in the
GDP-bound form through interaction with Rab GDIs.
Each Rab protein is localized at organelles and is involved
in the vesicle transport of the organelle (447, 528, 529, 550,
636) (Fig. 8). For example, Rab1, Rab2, and Rab6 are
localized at the endoplasmic reticulum and the Golgi apparatus and regulate vesicle transport along the biosynthetic/secretory pathway (104, 240, 265, 448, 575, 720);
Rab3 is localized on secretory granules including synaptic
vesicles and is involved in Ca2-dependent exocytosis
(91, 140, 183185, 216, 217, 282, 319, 408, 476, 477, 625);
Rab4 and Rab5 are present on early endosomes and regulate early steps of the endocytic process, mediating endosome-endosome fusion and receptor recycling, respectively (73, 104, 237, 747, 748). However, the sites of action
of the majority of Rab proteins have not yet been elucidated. In addition, it seems likely that some functional
specialization of Rab proteins takes place, both in different cell types and in different organisms. One example is
Rab11A and Rab11B and its two yeast homologs Ypt31p
and Ypt32p. A role for Rab11 has been documented in late
recycling of transferrin receptors (590, 736) and in transGolgi network-to-plasma membrane transport (109). On
the other hand, in yeast, Ypt31p and Ypt32p are involved
in intra-Golgi transport or protein export from the transGolgi (46).

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D. Rab Protein Cycle: Cyclical

Activation/Inactivation and Translocation
Rab proteins cycle between the GDP-bound inactive
and GTP-bound active forms and between the cytosol and
the membranes, and these cyclical activation, inactivation, and translocation are regulated by at least three
types of regulators: GEPs, GDIs, and GAPs (447, 528, 703)
(Fig. 9). In the cytosol, a Rab protein is maintained in the
GDP-bound inactive form by a GDI. The GDP-bound form
is first released from the GDI by a still unknown mechanism that is coupled to the delivery of the Rab protein to
a specific membrane compartment (673, 735); the Rab
protein is subsequently converted to the GTP-bound form
by the action of a GEP. The GTP-bound form then interacts with a downstream effector(s). Thereafter, the GTPbound form is converted to the GDP-bound form by the
action of a GAP. The GDP-bound form produced on the
membrane then complexes with the GDI and returns to
the cytosol as a Rab protein-Rab GDI complex. Thus a
characteristic of Rab proteins is that a cycle of membrane
association/dissociation is superimposed onto their GDP/
GTP cycle.
Yeast Dss4 and its mammalian counterpart, Mss4,
were first isolated and characterized as Rab GEPs (79,
497). Dss4 is active on both Sec4 and Ypt1 and Mss4 is
active on a subset of Rab proteins. Moreover, the exchange activities of both GEPs are very low compared
with the values of other GEPs for Ras and Rho/Rac/Cdc42
proteins. These proteins are unlikely to serve as physiologically relevant GEPs, and their real roles are currently
unknown. Only three physiological Rab GEPs have been
isolated and characterized: Rab3 GEP, specific for the
Rab3 subfamily members (762); Rabex-5, specific for the
Rab5 subfamily members (290); and Sec2 for Sec4 (763).
Rab3 GEP is ubiquitously expressed but abundant in
brain (762). It is active at least on Rab3A, Rab3C, and
Rab3D and is inactive on other Rab proteins, including

FIG. 9. Cyclical activation/inactivation of Rab proteins

and their translocation. Rab, Rab proteins.

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SMALL G PROTEINS

Rab2, Rab5A, Rab10, and Rab11. It prefers the lipid-modified form to the lipid-unmodified form as a substrate. A C.
elegans counterpart gene of this mammalian GEP, Aex-3,
has been identified genetically; mutation of this gene
causes a defect in synaptic transmission (310). Rabex-5
forms a tight complex with Rabaptin-5, a Rab5 effector,
and activation of Rab5 by Rabex-5 is likely coupled to
effector recruitment (290). Rabex-5 is homologous to
Vps9, a yeast protein involved in endocytosis (77). In the
yeast, Sec2, a GEP for Sec4, is genetically essential for the
Golgi-to-plasma membrane transport (763). Sec2 shares
sequence homology to Rabin-3 that specifically interacts
with Rab3A and Rab3D (68). However, Rabin-3 has no
detectable GEP activity. These data suggest that each
member or each subfamily of the Rab family could interact with a specific GEP. It is not known whether Rabex-5
and Sec2 require lipid modifications of their small G protein substrates.
Only two GAPs for Rab proteins have been isolated
and characterized in mammalian cells: Rab3 GAP, specific for the Rab3 subfamily members (205, 501) and
GAPCenA, specific for Rab6 (135). Rab3 GAP is ubiquitously expressed and enriched in the soluble synaptic
fraction in brain (205, 501, 543). Rab3 GAP consists of two
subunits, a catalytic and a noncatalytic subunit (205, 501).
It is active at least on Rab3A, Rab3B, Rab3C, and Rab3D
and is inactive on other Rab proteins, including Rab2,
Rab5A, and Rab11 (205). It prefers the lipid-modified form
to the lipid-unmodified form as a substrate. The role of the
noncatalytic subunit of Rab3 GAP is unknown, but Sar1
GAP also consists of catalytic (Sec23) and noncatalytic
(Sec24) subunits (270, 808) (see sect. VI). GAPCenA is
ubiquitously expressed (135). It is mainly cytosolic, but a
minor pool is associated with the centrosome. It forms a
complex with -tubulin and may play a role in microtubule nucleation. In the yeast S. cerevisiae, five GAPs have
been isolated and characterized: Gyp1 (GAP for Sec4 and
Ypt1), Gyp6 (GAP for Ypt6), Gyp7 (GAP for Ypt7), Mdr1p/
Gyp2p (GAP for Ypt6 and Sec4), and Msb3p/Gyp3p (GAP
for Sec4, Ypt6, Ypt51, Ypt31/ Ypt32, and Ypt1) (7, 163, 691,
760). Interestingly, Gyp proteins and GAPCenA (but not
Rab3 GAP) share significant homology to yeast cell cycle
checkpoint proteins, Bub2 and Cdc16, raising a possibility
that these Rab GAPs serve to coordinate membrane trafficking with other events taking place during mitosis, such
as the control of microtubule nucleation.
GAPs terminate the function of Rab proteins, but it is
not yet clear whether GTP hydrolysis is important for Rab
proteins to accomplish their functions. The cycle of GTP
hydrolysis of Rab5 has been shown to be uncoupled to the
Rab5-regulated endosome-endosome fusion (619). Similarly, the GTP hydrolysis of Ypt1 seems dispensable for
the Ypt1-mediated fusion events in the yeast (598). In
these cases, the GTP-bound form seems to be requisite or
even stimulatory for the progression of a vesicle toward

175

membrane fusion. In contrast, for Rab3A, the GTP-bound

form might interact with a prefusion complex, thus preventing fusion. The GTPase-deficient mutant of Rab3A
inhibits Ca2-dependent exocytosis from PC12 cells and
chromaffin cells (282, 319) (see below). In this case, Rab3
GAP plays a crucial role in the function of Rab3 proteins.
Three isoforms of Rab GDI have been isolated: Rab
GDI (Rab GDI-1), Rab GDI, and Rab GDI (Rab GDI2)(315, 453, 520, 629, 666). Southern blot analysis of
genomic DNA indicates that both mouse and rat contain
at least five Rab GDI genes (315). In contrast, there is only
one Rab GDI gene (GDI1) in the yeast S. cerevisiae, and
it is essential for cell viability (213). The properties of Rab
GDI and Rab GDI have most extensively been studied
(26, 27, 28, 315, 520, 629, 665, 666, 800). Rab GDI is
ubiquitously expressed, whereas Rab GDI is abundantly
expressed in neuronal cells (520). Despite the different
tissue distribution, both isoforms show similar biochemical properties (26, 28, 520, 629, 800). 1) Both proteins
bind the lipid-modified, GDP-bound form, but not any
other forms such as the lipid-modified, GTP-bound form,
the lipid-unmodified GDP-bound form, and the lipid-unmodified GTP-bound form. 2) Both proteins inhibit the
basal and GEP-stimulated dissociation of GDP from the
GDP-bound form. 3) Both proteins inhibit the binding of
the GDP-bound form to the membrane and keep it in the
cytosol. 4) When the GDP-bound form is produced from
the GTP-bound form on the membrane, both proteins
bind the GDP-bound form and retrieve it from the membrane. In addition to these activities, Rab GDI plays an
important role in specific delivery of Rab proteins to their
target membranes (572, 673, 703, 735). For instance, Rab5Rab GDI and Rab9-Rab GDI complexes mediate in vitro
binding of Rab5 and Rab9 to organelles where these proteins normally reside (673, 735). The GDP/GTP exchange
reaction does not seem to be a prerequisite for the Rab
protein binding to the membranes, because the GDPbound form can be transiently detected in the membranes
after delivery of Rab protein-Rab GDI complexes onto
membranes is likely to be followed by GDP/GTP exchange and the release of the GDI. A GDI dissociation
factor (GDF) has been identified that is active on Rab5
and Rab9 and devoid of GEP activity but is not yet fully
characterized (154).
Rab GDI and Rab GDI are significantly related to
the mammalian Choroideremia gene product CHM,
which delivers Rab proteins to the catalytic subunit of
geranylgeranyltransferase II as described above (645).
Choroideremia is an X-linked form of retinal degeneration (267). Recently, the crystal structure of Rab GDI has
been determined, and the sequence-conserved regions are
clustered on one face of the molecule (633). The two most
sequence-conserved regions, which form a compact structure at the apex of Rab GDI, play a crucial role in the
binding of Rab proteins. Mutations of this region in the

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TAKAI, SASAKI, AND MATOZAKI

human Rab GDI gene are responsible for X-linked nonspecific mental retardation (see sect. VG) (138).
E. SNAREs and Tethering Proteins in Vesicle
Targeting/Docking/Fusion
According to the SNARE hypothesis, there are vesicle and target membrane SNAREs (v- and t-SNAREs) and
these two types of SNAREs interact with each other,
resulting in the docking of vesicles with their target membranes (615) (Fig. 10). Recent results suggest that SNARE
pairing cannot be solely responsible for the appropriate
targeting and docking of transport vesicles. For example,
the docking of endoplasmic reticulum-derived vesicles
with Golgi membranes can occur in the absence of
SNARE components (83). Similarly, initial events leading
to the homotypic fusion of vacuoles are not dependent on
SNARE components (632, 740). Moreover, SNARE components can form promiscuous complexes in vitro (799).
These results suggest that SNAREs are not primarily involved in the specific targeting of vesicles.
Evidence is accumulating that targeting/docking
specificity is determined by the collaboration of several
factors. Tethering proteins play key roles in vesicle targeting/docking (571, 771). This group includes Uso1 (83),
TRAPP (620), p115 (505, 620), exocyst (246), and EEA1
(115), all of which bind membranes before the formation
of SNARE complexes. In S. cerevisiae, Uso1 and a large
protein complex named TRAPP are implicated in endoplasmic reticulum-to-Golgi transport (83, 620, 749); the
exocyst, a large protein complex consisting of Sec3, Sec5,
Sec6, Sec8, Sec10, Sec15, and Exo70, is involved in targeting of post-Golgi vesicles to the plasma membrane
(246). A mammalian version of the exocyst has been
identified in mammalian cells, where it may be involved in
targeting of vesicles (or docking with) the basolateral
membrane of epithelial cells (243a, 295). In mammalian
cells, a complex of p115, GM130, and giantin induces the
interaction of Golgi-derived vesicles with Golgi membranes (505, 675), and EEA1 is involved in the homotypic

Volume 81

endosome fusion process (115). EEA1 has recently been

found to be an effector of Rab5 (115) (see below). It is
likely that the events that precede stable, SNARE-dependent docking of vesicles or membranes are the result of a
network of interactions between many proteins including
tethering proteins. Although the direct interaction between Rab proteins and the tethering proteins has only
been documented for Rab5/EEA1, Sec4, and the exocyst,
Rab proteins are likely candidates for orchestrating vesicle targeting through tethering proteins as described below.
F. Mode of Action of Rab Proteins in Vesicle
Targeting/Docking/Fusion
Several Rab protein effectors have been isolated and
characterized. The first effector to be identified is Rabphilin-3 for the Rab3 subfamily members (662, 663). Rabphilin-3 is expressed in neurons and interacts preferentially
with GTP-Rab3A. Rabphilin-3 is present on the surface of
synaptic vesicles (478). It has not been established how
Rabphilin-3 localizes to synaptic vesicles: one model is
that it is recruited to the vesicles by Rab3A, analogous to
the Ras protein-Raf protein kinase system (397, 683);
another model is that it is targeted to synaptic vesicles
independently of Rab3 (460, 654, 664). The NH2-terminal
region constitutes the Rab3-binding region; the COOHterminal region contains two C2 domains that bind Ca2
and phospholipid (566, 662, 792). Expression or microinjection of either the NH2-terminal or the COOH-terminal
region of Rabphilin-3 blocks Ca2-dependent exocytosis
in several different systems (78, 117, 364). Recently, it was
shown that the abnormalities of synaptic transmission
and synaptic plasticity that are observed in Rab3A-deficient mice are not observed in Rabphilin-3-deficient mice
(638). This observation suggests that another Rab3 effector is present and compensates for loss of function of
Rabphilin-3 in these mice. Rim is another effector of Rab3
proteins (766). Rim has an NH2-terminal Rab3A-binding
domain that is homologous to that of Rabphilin-3, a cen-

FIG. 10. Possible modes of action of Rab

proteins, tethering proteins, and SNARE proteins in vesicle targeting/docking/fusion processes. Rab, Rab proteins.

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SMALL G PROTEINS

tral PDZ domain, and two COOH-terminal C2 domains

that are separated by alternatively spliced sequences. In
contrast to Rab3A and Rabphilin-3, Rim is clearly absent
from synaptic vesicles but enriched on the presynaptic
plasma membrane, especially at the active zone. Although
the precise function of Rim is still obscure, overexpression of its NH2-terminal fragment suggests that Rim is
implicated in neurotransmitter release. The localization of
Rim to the presynaptic plasma membrane is intriguing in
terms of its interaction with vesicle-associated GTPRab3A and the postulated role of Rab proteins in vesicle
docking.
Rabaptin-5 was identified in a yeast two-hybrid
screen using the GTPase-deficient mutant of Rab5 as a
bait (685). Rabaptin-5 consists of two isoforms: Rabaptin-5 and Rabaptin-5 (241). Rabaptin-5 is recruited from
the cytosol to endosomal membranes by GTP-Rab5. In
addition, its overexpression induces the formation of
large endosomes, a phenotype that is also observed upon
overexpression of a GTPase-deficient mutant of Rab5.
Rabaptin-5 has an NH2-terminal coiled-coil region that
serves as a self-association determinant and a COOHterminal Rab5-binding domain. The NH2 terminus of
Rabaptin-5 has recently been found to also interact with
the GTPase-deficient mutant of Rab4, which is involved in
the receptor recycling process (753). Rabaptin-5 is part of
a large complex required for membrane docking/fusion
processes (457). EEA1, another effector of Rab5, is also
included in this complex (115, 457, 670). EEA1 may represent the core component of this complex and serve as a
tethering protein, because it alone could replace the requirement for cytosol in an early endosome fusion assay
(115). EEA1 contains two spatially separate Rab5-binding
domains and a phosphatidylinositol 3,4,5-trisphosphate
(PIP3) binding, FYVE finger at its COOH terminus (670).
Rabaptin-5 also interacts with the NH2-terminal region of
Rabphilin-3, and this interaction is inhibited by GTPRab3A (542). Because endocytosis is often coupled with
exocytosis, particularly at nerve terminals, this Rabphilin3-Rabaptin-5 interaction may contribute to the coupling of
these two events.
A Rab8-interacting protein (Rab8ip) is a serine/threonine protein kinase, closely related to the GC kinase
found in lymphoid germinal centers (591). Like Rab8, this
kinase localizes to the Golgi apparatus where it may
regulate vesicle transport to the cell surface in response
to Rab8. Rab8ip shares sequence homology to PAKs, protein kinases that are effectors of Rac/Cdc42 proteins
(439). An effector of Rab9, p40, is associated with endosomes and shows synergy with Rab9 in its ability to
stimulate transport of the mannose 6-phosphate receptor
from endosome to the trans-Golgi network in an in vitro
transport assay (151). p40 is comprised entirely of six
internally repeated sequences, named kelch repeats,
which were first identified in the Drosophila kelch protein

177

(787), although the function of this domain remains unknown. Rabkinesin-6 is an effector of Rab6, and a member
of the kinesin family (167). This protein displays a conventional kinesin structure with an NH2-terminal motor
domain, followed by a region predicted to form an helical coiled-coil stalk and a tail domain. Rab6 may regulate microtubule-dependent retrograde transport from
the Golgi apparatus through Rabkinesin-6 (777).
Rab11BP/Rabphilin-11, an effector of Rab11, is also involved in microtubule-based vesicle transport (432, 668,
810), although it is not a motor protein. This protein
contains a proline-rich domain within its NH2-terminal
half and WD40 repeats, important for the protein-protein
interactions, within its COOH-terminal half.
Thus several effectors have been identified within the
past 5 years. In addition, the recent determination of the
X-ray crystal structure of Rab3A complexed to the Rab3binding domain of Rabphilin-3 has enabled the identification of complementarity-determining regions, which are
potentially involved in the interactions of Rab proteins
with their effectors (555). These regions exhibit a high
degree of sequence variability among Rab proteins and
might enable them to interact with a wide variety of
effectors (482).
Although it has not been fully elucidated how each
Rab protein regulates vesicle targeting/docking/fusion
processes through their specific effectors, one probable
mechanism is to regulate or facilitate the assembly of
SNARE complexes (Fig. 10). Rab proteins are not core
components of SNARE complexes. However, genetic
studies in yeast have shown that functions of Rab and
SNARE proteins are linked: the effects of a mutation in
the effector domain of Sec4 is suppressed by overexpression of Sec9, the SNAP-25-like protein (63), and Ypt1 is
involved in the priming of t-SNARE (401). In this process,
Rab proteins may function to recruit tethering proteins
onto membranes and coordinate loose membrane tethering to induce the SNARE complex-mediated, tighter and
stable docking process.
GTP-Sec4 interacts with Sec15, a component of the
exocyst (246). Then, a chain of protein-protein interactions leads to the assembly of the exocyst and its binding
to Sec3, which marks the specific site of exocytosis at the
plasma membrane. After Rab5 is activated by Rabex-5
complexed with Rabaptin-5, Rab5 recruits EEA1, which
interacts with PIP3 to early endosome (115, 670). PIP3
might serve to stabilize EEA1 on membranes through
interaction with its FYVE domain. Uso1 is needed to allow
the assembly of SNARE complex and Uso1-regulated tethering is helped by Ypt1 (83). Although it is not clear how
Rab proteins and tethering proteins induce the assembly
of SNARE complexes, they may induce the dissociation of
the Sec1/Munc18 family members which impair the association of t-SNARE with v-SNARE (Fig. 10). Consistent
with this possibility, Vac1, a mammalian homolog of

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EEA1, interacts not only with an effector of Vps21 (a

yeast Rab protein), but also with Vps45, a member of the
Sec1/Munc18 family (569). In addition, other docking
complexes regulated by Rab proteins have been characterized. For example, Rabphilin-3 and Rim, two effectors
of Rab3 proteins, may be part of a complex that links
synaptic vesicles to the presynaptic plasma membrane in
neurons.
G. Rab3A in Ca2-Dependent Exocytosis
Transient overexpression or microinjection of a dominant active mutant of Rab3A inhibits Ca2-dependent
secretion from bovine chromaffin cells and PC12 cells as
described above (282, 319). In contrast, Rab3B stimulates
secretion: in antisense RNA experiments performed in
anterior pituitary cells, reduction in Rab3B mRNA inhibited Ca2-dependent secretion from these cells (407), and
when a dominant active mutant of Rab3B was stably
expressed in PC12 cells, where endogenous Rab3B is
absent, Ca2-dependent norepinephrine secretion was
markedly stimulated (772). The explanation for these apparently opposite effects of Rab3A and Rab3B on Ca2dependent exocytosis is currently unknown.
Studies on Rab3A-deficient mice have revealed an
important insight into Rab3A function. Synaptic depletion
is much faster in the hippocampal CA1 region of these
mice, although two forms of short-term synaptic plasticity, paired-pulse facilitation and posttetanic potentiation,
are unaffected (216). These findings suggest that Rab3A
plays a role in the recruitment of synaptic vesicles for
exocytosis. In Rab3A-deficient, cultured hippocampal
cells, Rab3A is involved in a late step in vesicle fusion
(217). Moreover, Rab3A-deficient mice show reduced formation of postsynaptic long-term potentiation in the CA3
region (91). Thus Rab3A is not essential for basal synaptic
transmission; rather, it modulates synaptic vesicle trafficking, thereby contributing to synaptic plasticity.
Recent genetic analysis of X-linked, nonspecific,
mental retardation has revealed that mutations or deletions of the Rab GDI gene can cause this disease (138).
Notably, Rab GDI is localized to the distal part of chromosome Xq28. Mutations in this region have recently
been described to cause a syndromic form of X-linked
nonspecific mental retardation that comprises epileptic
seizures (29). Consistent with these findings, analysis of
Rab GDI-deficient mice has shown that synaptic potentials remain elevated and show reduced depression during repetitive stimulation in the hippocampal CA1 region
(306). In addition, the mice have increased propensity for
seizures. These observations suggest that Rab GDI is
essentially important for vesicle trafficking in neural cells,
and its function may be needed to suppress hyperexcitability, likely through Rab3 proteins.

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H. Rab Proteins and Cytoskeleton

Recently, a kinesin-like protein, termed Rabkinesin-6,
has been identified as a downstream effector of Rab6, for
the first time linking Rab proteins to the microtubule
cytoskeleton (167). Rab6 is associated with highly dynamic tubular structures that move along microtubules
from the Golgi apparatus to the cell periphery (777). Rab6
may facilitate the transport of these structures to their
acceptor compartment, probably the endoplasmic reticulum, either by recruiting Rabkinesin-6 onto membranes or
by changing the biochemical properties of the motor.
Similarly, Rab5 stimulates both association of early endosomes with microtubules and early-endosome motility
toward the minus ends of microtubules (516). Rab11 is
colocalized with its effector, Rab11BP/Rabphilin-11, along
microtubules (88, 432, 668, 810). Rab11 and its effector
protein are involved in cell migration, probably via regulation of the vesicle recycling (303, 432). Sec4 promotes
polarized transport of post-Golgi vesicles along the actin
cytoskeleton through an interaction with its GEP, Sec2
(763). Rab proteins may also serve to link vesicles and
target membranes to the cytoskeleton at docking sites.
For example, Rabphilin-3, an effector of Rab3 proteins,
interacts with the actin-bundling protein -actinin (333).
This interaction may induce the local organization of the
actin cytoskeleton for the fusion of synaptic vesicles with
the plasma membrane (233).
I. Rab Proteins in Vesicle Budding
There are several lines of evidence that Rab proteins
may also play a role in the budding process (see below).
Depletion of Ypt31 and Ypt32 in the S. cerevisiae leads to
the accumulation of stacks of membranes that resemble
the typical Golgi cisternae of mammalian cells, suggesting
a role for these proteins in intra-Golgi transport and in the
formation of transport vesicles at the exit face of the
Golgi apparatus (46). Dominant negative mutants of Rab1
and Rab9 appear to block the budding of the vesicles from
the endoplasmic reticulum and late endosomes, respectively (530, 604). A complex of Rab5 and Rab GDI appears
to be required for coat invagination and receptor sequestration in an in vitro assay for clathrin-coated pit assembly
(461). Similarly, clathrin-coated vesicles can recruit Rab5
(287). However, it has not yet been demonstrated that Rab
proteins regulate the assembly of coat proteins as described for Sar1/Arf proteins (see below). Currently, the
possibility cannot be excluded that the effects of Rab
proteins in the budding process may be an unknown,
secondary effect in the targeting/docking process, because after or during the bud formation, coat proteins are
disassembled to produce uncoated vesicles. Before, during, or after this uncoating process Rab proteins may

January 2001

SMALL G PROTEINS

be associated with the vesicles (Fig. 7). Consistently,

Rab11BP/Rabphilin-11 (432, 668, 810), a downstream effector of Rab11 mainly implicated in vesicle recycling
(590, 736), directly interacts with mammalian Sec13 (433).
In yeast, Sec13 counterpart is involved in Sar1-induced
vesicle coat assembly (714) (see below). This result provides a physical connection between a Rab protein and a
structural component necessary for vesicle budding.
VI. SAR1/ARF PROTEINS AS REGULATORS
OF VESICLE BUDDING
A. Outline
The SAR1 gene was originally isolated as a multiple
copy suppressor of a ts mutant of the SEC12 gene (Sar1
GEP gene) that had genetically been identified to be required for transport from the endoplasmic reticulum to
the Golgi apparatus in the yeast S. cerevisiae (508). Subsequently, the SAR1 gene was also shown to be involved
in transport from the endoplasmic reticulum (509, 545).
There is only one SAR1 gene in yeast. Two Sar1 proteins
(Sar1a and Sar1b) have been found in mammals (372).
Arf proteins are homologous to Sar1 with 35% identity. Six Arf proteins and some Arf-like proteins (ARLs
and ARD) have thus far been identified in mammals, and
three members exist in S. cerevisiae (493). Arf proteins
were originally purified from rabbit liver and bovine brain
membranes based on their ability to stimulate in vitro the
cholera toxin-catalyzed ADP-ribosylation of Gs (326, 327).
The cDNAs of bovine and yeast Arf1 were then isolated
(658). It was shown that Arf1 is localized to the Golgi
apparatus in some cell lines and that Arf1 mutations cause
secretion defects in S. cerevisiae (684). Subsequently,
small G proteins involved in vesicle formation from the
Golgi apparatus were identified to be Arf1 and another Arf
protein (655). It is now established that Sar1 and Arf
proteins are involved in the budding of vesicles from yeast
to human (103, 493, 494, 634).
Mammalian Arf proteins are structurally grouped
into three classes: class I including Arf1, Arf2, and Arf3;
class II including Arf4 and Arf5; and class III including
Arf6 (103, 494). Of these Arf proteins, Arf1 has been the
most extensively characterized and is established to be
involved in the budding of vesicles from the Golgi. Earlier
studies implicated Arf6 in the formation of vesicles from
the plasma membrane (12, 161, 162, 585, 814). However,
recent studies indicate that it is also involved in remodeling of the actin cytoskeleton and cell motility downstream of Rac1 (584).
B. Coat Proteins and Vesicle Budding
Vesicle budding requires the assembly of specific
proteins coating the cytoplasmic face of a donor mem-

179

brane (634). The assembly of coat proteins is thought to

serve at least two functions: it provides the mechanical
force to pull the membrane into a bud, and it helps to
capture specific membrane receptors and their bound
cargo molecules. Three classes of coated vesicles have
been well-characterized to date: clathrin-, COP (coat protein) I-, and COPII-coated vesicles (634, 640). Clathrin
coats contain clathrin and heterotetrameric adaptor protein (AP) complexes, AP-1, AP-2, AP-3, or AP-4 (278, 350).
Two different clathrin-coated vesicles containing AP-1
and AP-2 carry selected proteins from the trans-Golgi
network to endosomes and lysosomes and from the cell
surface to endosomes, respectively. Another AP complex,
AP-3, is involved in the biogenesis of specialized organelles such as pigmented granules and synaptic vesicles
and in an alternative Golgi-to-vacuolar pathway in yeast.
AP-4 has recently been identified as a novel complex
related to other AP complexes through searches in EST
data, but its function remains to be determined. COPIIcoated vesicles have been shown to be involved exclusively in the export of cargo molecules from the endoplasmic reticulum (35, 38, 634). COPI-coated vesicles
seem to be involved in membrane trafficking between the
endoplasmic reticulum and the Golgi apparatus, in intraGolgi transport, and possibly in endosomes. However, it is
currently a matter of debate as to how many transport
steps COPI is involved in, and whether it functions in
transport in the anterograde or retrograde direction or
both (35, 405, 410). Sar1 and Arf proteins play crucial
roles in the membrane recruitment of the COPII and COPI
components, respectively (38, 405). Arf proteins are also
involved in the recruitment of AP-1 and AP-3 components
to the membranes (278). In contrast, neither of these
small G proteins is involved in the clathrin/AP-2-coated
vesicle formation from the cell surface.
C. Arf Protein Cycle: Cyclical
Activation/Inactivation and Translocation
Arf proteins cycle between GDP-bound, inactive and
GTP-bound, active forms, and the cycling is regulated by
specific GEPs and GAPs (312, 494) (Fig. 11). This cycling
is not regulated by GDIs. The GDP-bound form of Arf
proteins is present in the cytosol and is converted to the
GTP-bound form by the action of a GEP, inducing the
conformational change that allows the myristoylated NH2terminal amphipathic helix of the Arf proteins to interact
with phospholipid bilayers. Nucleotide exchange also induces a conformational change in the switch I and II
regions, allowing Arf proteins to interact with downstream effector (110, 229, 613).
Certain recent studies indicate that the myristate is
exposed and can interact with the phospholipid bilayer
when Arf proteins are still in the GDP-bound form (21).

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Volume 81

FIG. 11. Cyclical activation/inactivation of Arf proteins and their translocation. Arf, Arf proteins.

This weak but measurable membrane association is completely abolished if the myristate is removed (190). Arf
protein-membrane interaction permits the activation of
Arf proteins by a GEP. Membrane association is stabilized
in the GTP-bound form by a conformational change of the
NH2-terminal helix that exposes several hydrophobic
amino acid residues, including Leu-8 and Phe-9 (which are
buried inside the Arf-GDP), and permits their insertion
into the membrane (21, 47). A cytosolic GAP is recruited
to the membrane after vesicle budding to stimulate the
hydrolysis of bound GTP to GDP, resulting in the release
of the Arf protein and in turn the COPI coat.
Many GEPs for Arf proteins have been isolated and
characterized in yeast and mammals (312, 494, 613). Although they show different substrate specificities, they
have a conserved 200-amino acid region, called as the
Sec7 domain, that shows strong similarity to yeast Sec7.
Sec7 was initially identified as a protein required for
transport at different stages of the yeast secretory pathway and is localized to the Golgi apparatus (195). The
Sec7 domain itself was subsequently shown to be a minimum GEP unit capable of binding to and activating Arf
proteins (99, 229, 440, 489, 570, 631).
Members of the Arf GEP family can be grouped into
two major subfamilies on the basis of their sequence
similarities and functional differences (312, 614). The
high-molecular-weight Arf GEP subfamily includes yeast
Sec7, Gea1, and Gea2, and mammalian BIG1/p200, BIG2,
and GBF1, which all consist of 1,400 2,000 amino acid
residues. Plants also have related Arf GEPs. There is a
Sec7 domain in the middle of these polypeptides, as well
as some additional, homologous regions. It is a notable
feature of these Arf GEPs that all but one (GBF1) are
sensitive to a fungal metabolite, BFA (490, 570, 631, 723),
which inhibits their Arf GEP activity in an uncompetitive
manner (440, 570). BIG1/p200 has been shown to be also
a Golgi-localized protein in mammalian cells (440). GBF1
is colocalized with COPI to the Golgi apparatus, and,
when overexpressed in cells, confers BFA resistance, suggesting that it may be involved in the formation of COPIcoated vesicles through activation of Arf proteins (122).
The second subfamily (low-molecular-weight type) in-

cludes mammalian ARNO, cytohesin-1, GRP1, and cytohesin-4 (312, 614). No homologs exist in yeast. These
proteins consist of 400 amino acids and have in common an NH2-terminal coiled-coil region, a central Sec7
domain, and a COOH-terminal PH domain. Furthermore,
all are insensitive to BFA. EFA6, a GEP specific for Arf6
(191), can also be grouped into this subfamily because it
is BFA insensitive and has all the structural features,
despite a slightly larger size. Although earlier studies
reported that ARNO, cytohesin-1, and GRP1 showed preferential GEP activity on class I and II Arf proteins, recent
studies have revealed that these small GEPs are colocalized with Arf6 at the cell periphery and cause elevation of
GTP-Arf6 (192, 193, 382).
ARNO prefers myristoylated Arf proteins and its activity is enhanced by PIP2 (99). In contrast, GRP1 binds to
PIP3 but poorly to PIP2 through its PH domain (357). It is
therefore likely that these small GEPs use the phosphoinositide-PH domain for their recruitment to a specialized
membrane subdomain, where the GEPs activate myristoylated GDP-Arf proteins. Supporting this model are the
recent observations that translocation of these small
GEPs to the plasma membrane is stimulated by growth
factors and is blocked by inhibitors of PI 3-kinases (382,
503, 751).
The crystal structure of GDP-Arf1 has revealed a
patch of positively charged amino acids on its surface (16). Thus Arf1 may also interact with negatively
charged phospholipids on membranes via this patch.
Crystal structure analyses of 5-guanylylimidodiphosphate [Gpp(NH)p]-bound Arf1 and of nucleotide-free Arf1
complexed with the Sec7 domain of Gea2 have provided
insight into the activation mechanism of Arf1 (229). Arf1
in its inactive conformation cannot fit into the recognition
surface of the Sec7 domain. Taken together with the
biochemical data that a low-affinity interaction of myristoylated GDP-Arf1 with the phospholipid membrane is
required for subsequent nucleotide exchange by GEPs
(21, 47, 190), it is likely that phospholipids act on Arf1 to
induce the conformational change that permits the binding of Arf1 to the Sec7 domain, followed by Arf1 activation.

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SMALL G PROTEINS

Many Arf GAPs have been identified in yeast and

mammals (494, 614). These GAPs each contain a zinc
fingerlike Arf GAP domain, which has been shown to be a
minimum unit for the GAP activity. However, their substrate specificities have not yet been systematically characterized. An Arf GAP purified from rat spleen cytosol is
activated by PIP2 and shows broad substrate specificity,
being active on Arf1, Arf3, Arf5, and Arf6 (153). This
activity does not require Arf myristoylation. Another Arf
GAP (Arf1 GAP), the first GAP identified in mammals
(136), is localized to the Golgi apparatus and is stimulated
by diacylglycerol, which may be produced from phosphatidylcholine by the sequential actions of phospholipase D (a downstream effector of Arf proteins; see below), and phosphatidic acid phosphohydrolase (22).
Recruitment of cytosolic Arf1 GAP to the vesicle membranes is enhanced by cargo receptors such as the KDEL
receptor, which recognizes a Lys-Asp-Glu-Leu (KDEL)
COOH-terminal motif on certain soluble proteins destined
for Golgi-to-endoplasmic reticulum retrieval (23, 24). Centaurin- and PIP3-BP, two closely related proteins, have
two PH domains that are involved in binding to PIP3 (253,
710). ASAP1/DEF-1 and PAP, also close relatives, have a
PH domain, three ankyrin repeats, a proline-rich region,
and an SH3 domain and have been shown to interact with
Src and Pyk2, respectively (19, 70, 348). Both enhance the
GTPase activity of Arf1 and Arf5 but act only weakly on
Arf6. PKL and GIT are closely related and contain three
ankyrin repeats and a PH domain (120, 528, 731). PKL was
identified as a protein that binds to the paxillin LD4 motif
and serves as a connector between paxillin and molecules
of the Rac1/Cdc42 pathway, PAK and PIX (731). GIT1 was
identified as a protein that interacts with a G proteincoupled receptor kinase; it regulates endocytosis of various G protein-coupled receptors (120, 578). To date, the
crystal structures of the GAP domain of PAP and that of
Arf1 GAP complexed with Arf1 have been determined
(230, 435). A unique chimera protein, ARD1, contains both
Arf protein and Arf GAP structures (474). The GAP domain is specific for the Arf domain and shows GDI activity
that slows the dissociation of GDP from the Arf domain
(754).
D. Arf Proteins in Vesicle Budding
Although it is currently unclear at which stage Arf
GEP is activated or how its activation is controlled, GTPArf proteins, once produced by the action of Arf GEP,
associate with the Golgi apparatus and recruit coatomer,
a seven subunit complex (410, 780), or a clathrin-adaptor
complex, AP-1 or AP-3 (278). In the case of COPI-coated
vesicle formation, GTP-Arf1 has been shown to interact
with -COP and -COP (780, 817, 818). However, it is

181

currently unknown which subunits of the AP complexes

interact with GTP-Arf proteins. Acidic phospholipids,
such as phosphatidic acid, enhance Arf1-induced binding
of coatomer (370, 614). Assembled COPI captures cargo
molecules through binding to cargo receptors, such as
KDEL receptor and the p24 family of proteins (410, 780).
In the latter case, COPI recognizes the phenylalanine or
dilysine motif (or both) within the cytoplasmic tail of the
p24 family members. Thus vesicle formation and cargo
selection are coupled through a bivalent interaction of
coatomer with Arf1 and a putative cargo receptor. It is
speculated that coat protein assembly induces local membrane deformation into a nascent vesicle bud.
The entire process of the formation of COPI-coated
and clathrin/AP-1-coated vesicles can be reproduced with
pure components in vitro (677, 825). The simplest systems
consist of GTP-Arf1 and either coatomer, or clathrin and
AP-1, added to liposomes of defined composition that
support the formation of small coated vesicles (40 70 nm
diameter for COPI and 60 80 nm diameter for clathrin/
AP-1) (677, 825). The results indicate that Arf1 and
coatomer are sufficient to pinch off vesicles. Although it
has been proposed that production of phosphatidic acid
by Arf1-induced activation of phospholipase D triggers
binding of coat proteins to membranes and subsequent
vesicle formation, the physiological relevance of these
events is unclear (612). On the other hand, a recent study
has suggested that production of phosphatidic acid by
acylation of lysophosphatidic acid may play a role in
vesicle budding from the Golgi apparatus (773).
Either concomitant with or after vesicle budding,
GTP-Arf1 is converted to the GDP-bound form by the
action of a GAP. Recent studies have shown that
coatomer and a transmembrane cargo receptor control
membrane association of Arf1 GAP and its activity (23, 24,
230). KDEL receptor enhances the recruitment of Arf1
GAP to the membranes of the Golgi apparatus (23, 24).
Arf1 GAP activity may then be stimulated by coatomer on
the membranes (230). Consistent with this observation,
the crystal structure of GTP-Arf1 complexed with the
minimal GAP domain of Arf1 GAP indicates that, unlike
most G proteins, the GAP binding site on Arf1 does not
overlap with its effector binding site (230). Thus Arf1 may
simultaneously interact with Arf1 GAP and coatomer, and
both coatomer and cargo receptors may regulate the termination of the Arf activity.
E. Arf6 in Endocytic Recycling
and Cytoskeletal Reorganization
The subcellular localization of Arf6 is quite different
from that of class I and class II Arf proteins. Various
studies have shown that Arf6 is involved in recycling of

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endosomal vesicles and regulates receptor-mediated endocytosis (12, 161, 162, 585, 814). However, Arf6 is also
implicated in remodeling of the cytoskeleton underlying
the plasma membrane. Arf6 is localized to the plasma
membrane, especially to membrane ruffles in spreading
cells. Activation of Arf6 induces remodeling of the actin
cytoskeleton and cell spreading, and expression of a dominant negative mutant of Arf6 blocks cell spreading (160,
585, 674). Furthermore, ARNO and EFA6, two low-molecular-weight Arf GEPs, have been reported to modulate
growth factor- and protein kinase C-mediated cytoskeletal reorganization through activation of Arf6 (191, 193).
In addition, a dominant negative mutant of ARNO inhibits
both Arf6 translocation and cortical actin formation induced by growth factors (750). In this context, it is noteworthy that recent studies have suggested that there is
cross-talk between the Arf6 and Rac1 pathways in actin
remodeling. First, EFA6-induced cytoskeletal remodeling
is blocked by coexpression of a dominant negative mutant
of Arf6 or Rac1 (191). Second, a dominant negative mutant of Arf6 inhibits growth factor- and Rac1-mediated
membrane ruffling (584, 813). Third, a deletion mutant of
POR1/arfaptin-2, which interacts with both Rac and Arf
proteins, but not a dominant negative mutant of Rac1,
inhibits Arf6-mediated cytoskeletal rearrangements (160).
Taken together, these results indicate that Arf6 functions
either downstream of or in concert with Rac1. Further
evidence suggesting a connection between Arf6 and Rac1
pathways is the finding that PKL, an Arf GAP, serves as a
connector between paxillin and the Rac/Cdc42 pathway
components PAK and PIX (731). An effector of Arf6 that
is implicated in membrane ruffling is phosphatidylinositol
4-phophate 5-kinase (283). This kinase translocates to
ruffling membranes and produces PIP2 synergistically
with Arf6 and phosphatidic acid, the production of which
is catalyzed by phospholipase D. Because phospholipase
D itself is an effector of Arf proteins and PIP2 is an
activator of Arf proteins, it is tempting to speculate that
the local phospholipid metabolism that is regulated by
Arf6 may play a crucial role in membrane ruffle formation.
F. Sar1 as a Regulator of Vesicle Budding
The function of Sar1 in vesicle budding has been
extensively characterized in the yeast S. cerevisiae but in
less detail in mammals (38, 634). Sar1 does not undergo
any known lipid modification, but it is associated with the
endoplasmic reticulum and is involved in the formation of
COPII-coated transport vesicles from the endoplasmic
reticulum (39, 518, 544). The Sar1 cycle is regulated by a
GEP (Sec12) and a GAP (Sec23) (40, 44, 371, 808). Sec12
is an integral endoplasmic reticulum membrane protein
involved in activation of Sar1 (40, 142, 143). However, its
mammalian homolog has not yet been identified. Sec23

Volume 81

accelerates the GTPase activity of Sar1 and is tightly

associated with Sec24 (the Sec23 complex) (808). Sec23
homologs are present in mammalian cells, and they localize to compartments between the endoplasmic reticulum
and the Golgi apparatus (554, 558). A Sec24 homolog is
also present in mammalian cells, and as expected, it interacts with mammalian Sec23 (559, 715). Thus the Sar1/
COPII system is well conserved between yeast and mammalian cells.
COPII assembly and disassembly are regulated by the
Sar1 cycle (44, 371, 679). Sec12-induced activation of Sar1
promotes association of the Sec23 complex to the budding site at the endoplasmic reticulum. Then, the Sec13
complex comprised of Sec13 and Sec31 is recruited onto
the Sec23 complex, and this binding and the subsequent
polymerization of the Sec13 complex are proposed to lead
to the concentration of cargo proteins and the deformation of the membrane into a coated bud. Thus vesicles
coated with GTP-Sar1, the Sec23 complex, and the Sec13
complex, are produced. Sec23-induced GTP hydrolysis of
Sar1 leads to dissociation of these proteins from the
vesicle, although hydrolysis appears not to be a prerequisite for the vesicle budding. Two other factors have
been implicated in COPII-vesicle budding. Sec16 interacts
directly with the Sec23 complex and may serve as a
scaffold for COPII coat assembly (174, 225). Sed4 is highly
homologous to Sec12 in its cytoplasmic domain, although
it does not seem to show GEP activity on Sar1. However, unlike Sec12, Sed4 interacts directly with Sec16
(224). It is thus possible that Sar1, along with Sed4 and
Sec16, may be involved in a distinct vesicle formation
process.
Proteins necessary for the targeting and docking processes, such as v-SNARE and Rab proteins, must be incorporated into transport vesicles. However, the mechanism for this is currently unknown. Two facts are
established. 1) The Sec23 complex interacts simultaneously with GTP-Sar1 and two v-SNAREs, Bet1 and
Bos1, in a cooperative manner during the formation of
endoplasmic reticulum-to-Golgi COPII transport vesicles
(678), and 2) that Rab11BP/Rabphilin-11 (432, 668, 810), a
downstream effector of Rab11 mainly implicated in vesicle recycling (590, 736), interacts directly with mammalian Sec13 (433).
VII. RAN FUNCTION IN NUCLEOCYTOPLASMIC
TRANSPORT AND MICROTUBULE
ORGANIZATION
A. Outline
There is only one Ran gene in many cell types (including human and Schizosaccharomyces pombe), although cells of other species (S. cerevisiae and tomato)

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183

SMALL G PROTEINS

contain two or more closely related Ran genes (483). Ran

protein (Ras-related nuclear protein: Ran) was originally
cloned on the basis of its homology to Ras proteins (158).
The first evidence for the involvement of Ran in nuclear
transport was obtained in 1993 by showing that Ran is
essential for the nuclear import in permeabilized cells of
a reporter construct containing the nuclear localization
signal (NLS) of the simian virus 40 T antigen (PKKKRKV)
(464, 484). It is now clear that Ran plays a central role in
nucleocytoplasmic transport. Recent studies have uncovered another role for Ran in microtubule organization
during the M phase of the cell cycle (85, 325, 329, 351, 504,
533, 781).

export receptors, called exportins (734). Importins and

exportins recognize NLS and nuclear export signal (NES)
of cargos, respectively, bind them, and transport bound
cargos through the NPCs (3, 483). In some cases, transport receptors do not directly bind cargos and indirectly
bind them through adaptors that can recognize their NLS.
For instance, importin- and importin- are well-known
adaptor and receptor molecules, respectively. Transport
receptors constitute a superfamily of proteins that share a
binding domain of GTP-Ran (189, 235). In the yeast S.
cerevisiae, 14 members of importin- superfamily are
predicted from the genome sequence (3). Fewer receptors
have been identified in mammals, but more receptors will
be found in the future.

B. Nucleocytoplamsic Transport
Macromolecules are transported back and forth between the cytoplasm and the nucleus through NPCs. The
NPCs contain more than 50 different proteins (454, 517,
536, 565). Movement of macromolecules through the
NPCs occurs by at least two distinct mechanisms: passive
diffusion and active transport (180). Small molecules diffuse quickly through the NPCs in either direction,
whereas those larger than 50 60 kDa, including proteins
and RNAs, are actively and selectively transported. The
transport of cargo proteins requires at least three types of
soluble factors: transport receptor molecules, adaptor
molecules, and Ran and its binding proteins (139, 234,
454). Transport receptors shuttle continuously between
the nucleus and the cytoplasm, interact with NPCs, bind
cargo molecules, and facilitate cargo translocation
through the NPCs. There are two types of transport receptors: nuclear import receptors, called importins, and

C. Ran Cycle: Cyclical Activation/Inactivation

and Translocation
Like other small G proteins, Ran is cyclically activated and inactivated, but the most characteristic feature
of the Ran cycle is that the GTP-bound form and the
GDP-bound form are asymmetrically distributed in the
nucleus and the cytoplasm, respectively. This asymmetric
distribution of the two forms are due to asymmetric distributions of the regulators, a GEP and a GAP (Fig. 12).
Only one Ran GEP, regulator of chromosome condensation (RCC1) (54, 538), and only one Ran GAP, Ran GAP1
(51, 52), have been identified in mammals. RCC1 was
originally identified as a regulator of chromatin condensation (538). RCC1 is localized exclusively in the nucleus
where it is associated with the chromatin (539) and converts the GDP-bound form to the GTP-bound form (54). In
contrast to RCC1, Ran GAP1 is localized exclusively in the

FIG. 12. Cyclical activation/inactivation of

Ran and its translocation.

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TAKAI, SASAKI, AND MATOZAKI

cytoplasm (455, 596) but some can be detected in the

nucleus (726). In vertebrate cells, a significant portion of
Ran GAP1 acquires a 121-amino acid ubiquitin-like addition (called SUMO-1) (430, 455). This modification does
not target this protein for degradation but instead targets
it specifically to the filaments at the cytoplasmic face of
the NPCs. Moreover, two GTP-Ran-binding proteins, Ranbinding protein 1 (RanBP1) and Ran-binding protein 2
(RanBP2/NUP380), have been isolated (53, 784, 805).
RanBP1 is localized in the cytoplasm and enhances the
Ran GAP1 activity (53, 127, 597, 637), whereas RanBP2 is
a component of the NPCs at the cytoplasmic face that
recruits SUMO-Ran GAP1 and enhances its GAP activity
(430, 455). Once GTP-Ran is produced in the nucleus, an
exportin binds both GTP-Ran and its cargo in a cooperative manner, and this ternary complex is exported to the
cytoplasm through the NPCs. At the cytoplasmic face of
the NPCs, GTP-Ran in this complex is attacked by SUMORan GAP1 and Ran BP2 (430, 455, 622), and in the cytoplasm it is attacked by Ran GAP1 and RanBP1, leading to
conversion to GDP-Ran (50, 186, 409). Inactivation of
GTP-Ran induces the dissociation of the complex in the
cytoplasm. GDP-Ran then forms a complex with another
regulator, named p10/NTF2, which transports GDP-Ran
back to the nucleus through the NPCs (595). p10/NTF2
has a GDI activity, keeping Ran in the GDP-bound form
(789). GDP-Ran thus returned to the nucleus and is then
reutilized for the next cycle of transport. Thus the Ran
cycle is regulated by many factors. The most important

Volume 81

aspect of this cycle is that GTP-Ran forms a gradient

across the nuclear envelope and that this gradient plays a
key role in controlling the directionality of nuclocytoplasmic transport (236, 311). Ran does not undergo any posttranslational lipid modification and probably, as a result,
does not bind to membranes inside the cell or require
lipids for its activity (618).
D. Mode of Action of Ran in Nucleocytoplasmic
Transport
As described above, one role of Ran is to export
cargos via exportins from the nucleus to the cytoplasm.
The most well-characterized exportin is CRM1 (also
called exportin 1), which recognizes a specific, leucinerich type of NES (188, 203, 682). In the presence of
GTP-Ran, both GTP-Ran and a cargo bind to CRM1 in a
cooperative manner to form a GTP-Ran-CRM1-cargo complex (Fig. 13A). This complex is exported to the cytoplasm, where GTP-Ran is converted to GDP-Ran, causing
the cargo to dissociate from CRM1. CRM1 is then reimported to the nucleus in the empty state. In this example,
CRM1 functions without an adaptor. In some cases, NEScontaining proteins bind RNAs that are destined for nuclear export. In these cases, the NES proteins serve as
adaptors and the cargo is the RNA.
Ran has another role in nuclear import (536). In the
absence of GTP-Ran, importin- forms a complex with

FIG. 13. Mode of action of Ran in nucleocytoplasmic transport. A: mode of action of Ran in nuclear export. B: mode
of action of Ran in nuclear import.

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SMALL G PROTEINS

importin- that serves as an adaptor, binding NLS-containing proteins (Fig. 13B). This complex is imported into
the nucleus through the NPCs, where it encounters GTPRan. GTP-Ran binds to importin-, causing dissociation of
the complex into each component. Empty importin-,
probably still bound to GTP-Ran, is then reexported to the
cytoplasm. Recycling of importin- requires a specific
exportin, named CAS, that is analogous to CRM1 (377).
The binding of importin- to CAS requires cooperative
GTP-Ran binding, and CAS prefers importin-, which is
not associated with a cargo. Thus empty importin- is
exported again by CAS. After inactivation of GTP-Ran to
GDP-Ran in the cytoplasm, importin- dissociates, and
CAS returns to the nucleus for reutilization.
During nuclear transport, the nucleotide-bound state
of Ran acts as a simple switch to delineate the direction of
movement, and the energy of GTP hydrolysis is not
strictly required. Moreover, the binding of GTP-Ran to
importins induces conformational changes so that they
dissociate cargos. GTP-Ran is not essential for the transportation of importins through the NPCs, because the
NPC-interacting region of importins alone can be transported through the NPCs in the absence of GTP-Ran (3).

185

The precise mode of action of Ran and RanBPM in spindle

formation is not understood, but in the absence of a
nucleus, chromatin-bound RCC1 and cytoplasmic Ran
GAP1 produce a natural gradient of GTP-Ran that may be
most concentrated at the chromosome surface (325).
Consequently, the GTP-Ran-RanBPM complex would
stimulate microtubule assembly adjacent to the chromosomes. In this case again, GTP hydrolysis is not essential
for microtubule assembly but may have some secondary
role in the elongation of previously nucleated microtubules. Other cellular factors, including the chromosomeassociated, kinesin-like protein, XKLP1 (752, 764), and a
complex of cytoplasmic dynein, dynactin and NuMA (466,
801), are also involved in organizing these microtubules
into spindles. Thus the observation that Ran and RanBPM
stimulate microtubule polymerization offers a significant
insight into the process by which chromosomes drive
spindle assembly.
VIII. SMALL G PROTEIN CASCADES
AND CROSS-TALKS
A. Small G Protein Cascades

E. A Role for Ran in Microtubule Organization

During G1, S, and G2 phases of the cell cycle, nucleocytoplasmic transport is active, but during the M phase,
the nuclear envelope is broken down and nucleocytoplasmic transport stops. At the onset of mitosis in eukaryotes,
the nuclear envelope and interphase microtubule array
disassemble, and the duplicated centrosomes nucleate
microtubules that interact with chromosomes, forming a
bipolar spindle. Sister chromosomes balanced at the spindle equator segregate and move to opposite spindle poles
during anaphase. How chromosomes influence spindle
assembly in the absence of microtubule-nucleating organelles has been a long-standing mystery. Recent studies
have revealed that Ran and its novel effector, named
Ran-binding molecule (RanBPM) (504), regulate aster formation and spindle assembly.
An initial hint for Ran involvement in microtubule
regulation was derived from the observation that, in the
yeast S. cerevisiae, overexpression of Prp20, the yeast
RCC1 homolog, suppresses the toxic effect of certain
hyperstable -tubulin mutants (351). More direct evidence has recently been obtained by the identification of
a novel mammalian GTP-Ran-binding protein, RanBPM
(504). RanBPM associates with centrosomes, the microtubule-nucleating centers of mammalian cells. Overexpression of RanBPM in cultured cells or addition of GTPRan or GTPS-Ran to a cell extract induces aster
formation. Conversely, inhibition of RanBPM or Ran activity prevents in vitro aster formation (85, 329, 533, 781).

Multiple small G proteins form a signal cascade and

thereby transduce their signals to downstream effectors.
For instance, in the yeast S. cerevisiae, Rsr1, a member of
the Ras family, is thought to be first activated by an
unknown cue that is produced by the previous budding
site (45, 96) (Fig. 14). In the next step, GTP-Rsr1 binds to
Cdc24, a GEP for Cdc42, which in turn binds GDP-Cdc42
to activate it (821). The activation of Cdc42 induces not
only reorganization of the actin cytoskeleton but also the
recruitment of multiple small G proteins, including Rho1
and Sec4, to the future budding site. Sec4 would supply
the bud with vesicles necessary for bud enlargement (527,
623), while Rho1 would induce synthesis of the new cell
wall component, 1,3--glucan, by directly activating 1,3-glucan synthase and stimulating expression of the
genes necessary for this synthesis (80, 157, 456, 580).
Rho1 would also regulate reorganization of the actin cytoskeleton necessary for the budding processes (200, 304,
363, 739).
Another typical example of small G protein cascade
includes Rho/Rac/Cdc42 proteins in mammalian cells
(420, 744) (Fig. 15). Rho proteins regulate formation of
stress fibers and focal adhesions (564, 600, 601), Rac
proteins regulate formation of lamellipodia and membrane ruffles (521, 522, 602), and Cdc42 proteins regulate
formation of filopodia (368, 522). The sequential activation of these three small G proteins by extracellular agonists has been best characterized in quiescent Swiss 3T3
fibroblasts (522, 600, 602). Agonists like bradykinin activate Cdc42 proteins in these cells to produce filopodia or

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TAKAI, SASAKI, AND MATOZAKI

Volume 81

FIG. 14. Small G protein cascade in control of polarized

cell growth in the yeast S. cerevisiae.

microspikes. Activation of Cdc42 proteins leads to localized activation of Rac proteins; hence, filopodia are often
associated with lamellipodia, which are produced by Rac
proteins. In fact, it is hard to see filopodium formation
unless Rac activity is first inhibited. Quiescent Swiss 3T3
cells have no detectable stress fibers, and activation of
Rac proteins under these conditions leads to weak and
delayed activation of Rho proteins that produces stress
fibers. This typical cascade of Rho/Rac/Cdc42 proteins
observed in Swiss 3T3 fibroblasts has not always been
seen in other cell lines. For instance, it has been shown
that Rho proteins are inhibited by Rac/Cdc42 proteins in
cultured cells such as neuroblastoma cells, N1E-115 cells,
and NIH 3T3 cells (277, 388). In addition, Rho proteins
may in turn inhibit the activity of Rac/Cdc42 proteins in
N1E-115 cells (624).
B. Cross-talk Between Small G Proteins
In addition to the sequential cascade of multiple
small G proteins, distinct families of small G proteins

regulate various cellular functions in a cooperative manner. Although a dominant active mutant of Rho proteins
does not cause transformation of fibroblasts, dominant
negative mutants of Rho proteins inhibit Ras proteininduced transformation (341, 583). Similarly, a dominant
negative mutant of Rac/Cdc42 proteins inhibits Ras protein-induced transformation, while a dominant active mutant of these Rac/Cdc42 proteins enhances oncogenic Ras
protein-triggered, morphologic transformation (581, 582).
Thus Rho/Rac/Cdc42 proteins are involved in a cooperative manner in Ras protein-induced transformation.
It has recently been shown that Rho/Rab proteins
coordinately regulate cell adhesion and migration of cultured MDCK cells (303). TPA induces cell scattering of
MDCK cells and induces early disassembly of stress fibers
and focal adhesions followed by their reassembly in
MDCK cells. Expression of a dominant active mutant of
RhoA inhibits the TPA-induced disassembly of stress fibers and focal adhesions, while microinjection of C3
blocks their reassembly. In addition, microinjection of C3
or a dominant active mutant of RhoA inhibits the TPA-

FIG. 15. Small G protein cascade in control of cytoskeletal reorganization in Swiss 3T3 fibroblasts. Rho,
Rho proteins; Rac, Rac proteins; LPA, lysophosphatidic
acid.

January 2001

SMALL G PROTEINS

induced cell scattering. In contrast, expression of Rab

GDI or a dominant negative mutant of Rab5 attenuates
either the TPA-induced reassembly of stress fibers and
focal adhesions and subsequent cell scattering of MDCK
cells (303). Expression of a dominant active mutant of
Rac1 or Cdc42 inhibits the HGF/SF- or TPA-induced cell
scattering (331, 361). Therefore, during cell migration at
least, Rho/Rac/Cdc42 proteins may regulate the reorganization of the actin cytoskeleton, while Rab5 may control
the recycling of adhesion molecules such as integrins by
intracellular vesicle trafficking. The temporal and spatial
activation and inactivation of these two families of small
G proteins appear to be required for reorganization of the
actin cytoskeleton and cell migration.
IX. CONCLUSIONS AND PERSPECTIVES
A. Roles in Two Types of Cell Regulation
The mechanism of Gs-induced activation of adenylate
cyclase is different from that of EF-Tu-induced elongation
of polypeptide. Once GTP-Gs binds to adenylate cyclase, it
continuously activates it until it is converted to the GDPbound form. In contrast, GTP-EF-Tu elongates a single
amino acid, and further elongation requires the inactivation and reactivation of EF-Tu (60) (Fig. 16). GTPS-Gs
continuously activates adenylate cyclase, whereas GTPSEF-Tu stimulates just one cycle of peptide elongation and
stops further elongation. In this sense, Ras-induced Raf
protein kinase activation and Rho/Rac/Cdc42 protein-in-

187

duced activation of their specific downstream effector

protein kinases are similar to the Gs-induced adenylate
cyclase activation, whereas reorganization of the actin
cytoskeleton regulated by Rho/Rac/Cdc42 proteins, vesicle trafficking regulated by Rab and Sar1/Arf proteins, and
nucleocytoplasmic transport regulated by Ran are similar
to the EF-Tu-induced elongation of polypeptide and require cyclical activation and inactivation of the small G
proteins. Consistent with this mode of action of Gs and
Ras proteins, the ADP-riboyslation of Gs by cholera toxin
continuously activates adenylate cyclase, thereby causing
diarrhea (733), and point-mutated activated forms of Ras
proteins frequently found in a variety of cancers continuously activate Raf protein kinases that cause stimulated
cell proliferation (82). In contrast, no point-mutated activated forms of other small G proteins have been identified, and this may be due to the necessity for the cyclical
activation and inactivation of these small G proteins for
their cell functions. Instead, many GEPs for Rho/Rac/
Cdc42 proteins have been identified as oncogenes (93,
251, 744). Oncogenic GEPs increase relative ratios of
small G protein GTP-bound forms to GDP-bound forms
and show oncogenic phenotypes. From this point of view,
we should carefully interpret data obtained using dominant active mutants of small G proteins.
B. A Role as Biotimers Rather Than as Molecular
Switches
All downstream effectors of small G proteins have at
least two functionally distinct domains: a small G protein-

FIG. 16. Two types of cell regulation by G proteins. A: mode of action of EF-Tu in protein sysnthesis. G, EF-Tu; aa
tRNA, aminoacyl-tRNA. B: mode of action of heterotrimeric G protein. G, heterotrimeric G protein.

188

TAKAI, SASAKI, AND MATOZAKI

binding domain and a catalytic domain (in the case of

enzymes) or a protein-interacting domain (in the case of
nonenzymes). In the case of enzymes, the binding of the
small G protein induces conformational change, resulting
in its activation or inactivation. The activation (or inactivation) continues until the small G protein is converted to
the GDP-bound form by the action of GAPs, causing its
release from the enzyme. In the case of nonenzymes, the
binding of the small G protein induces conformational
change in the effector, resulting in the interaction with or
the dissociation from a further downstream effector. The
interaction or the dissociation continues until the small G
protein is converted to the GDP-bound form. In both
cases, the small G protein-binding region directly or indirectly masks the catalytic domain or the other proteinbinding domain, and the binding of the small G protein
induces unfolding of this functional domain. In this sense,
small G proteins work as more than molecular switches,
that activate or inactivate the downstream effectors, and
work as biological timers (biotimers), that induce activation or inactivation and determine the periods of the
functioning time (Fig. 17). It would be, therefore, of crucial importance to understand the mechanisms when and
how small G proteins are activated by GEPs and inactivated by GAPs.
C. A Role as Spatial Determinants
In addition to temporal regulation of small G proteins, spatial regulation is also essential for them to regulate various cell functions, particularly very dynamic cell
functions, such as cell migration and adhesion. The sites
of the activation of some small G proteins, such as Ras,
Sar1, and Ran proteins, are determined by their GEPs.
Upon stimulation of membrane receptors, Ras GEPs, such
as SOS, are recruited to the cytoplasmic region of the
plasma membrane receptors through adaptors, such as
GRB2/SHC, where Ras proteins are activated (75, 399,

FIG. 17. A role of small G proteins as biotimers rather than as

molecular switches. G, small G protein.

Volume 81

549, 617). A Sar1 GEP, Sec12, is a transmembrane protein

of the endoplasmic reticulum and interacts with GDPSar1 and activates it at the cytoplasmic surface of the
endoplasmic reticulum (40, 142, 143). A Ran GEP, RCC1,
is associated with chromatin and activates it in the nucleus (54, 539). In these cases, however, it remains unknown how activities of these GEPs are regulated. Chromatin structure may regulate RCC1 activity and cargo
proteins may regulate Sec12 activity. There is another
possibility that they are always active without being regulated and attack their substrates when they are available.
An Arf1 GEP, ARNO, binds to PIP2 and/or PIP3 and is
associated with a specialized membrane subdomain,
where it interacts with GDP-Arf1 and converts it to the
GTP-bound form (99). Thus lipid metabolism is crucial for
the localization of Arf1.
In contrast to these small G proteins, Rab proteins
stay in the cytosol in the GDP-bound form complexed
with Rab GDIs and they are translocated to their specific
membrane compartments where they are activated (572,
701703). GEPs may also be determinants for their localization, but Rab3 GEP is mostly found in the cytosol, and
there is no evidence that Rab3 GEP is first associated with
membranes and then recruits Rab3 proteins (543, 762).
Another possible mechanism is that the downstream effector(s) may serve as determinants. A downstream effector of Rab3 proteins, Rabphilin-3, is associated with synaptic vesicles in a Rab3A-independent manner, and upon
its activation, it is associated with the vesicles through
Rabphilin-3 (460, 654, 664). However, there is a contradictory result that the association of Rabphilin-3 with synaptic vesicles is dependent on GTP-Rab3A (397, 683). Rab5
proteins are also reported to be first associated with
endosomes, followed by recruitment of their downstream
effector, Rabaptin-5, and the COOH-terminal regions of
Rab5 proteins are essential for this specific localization
(685). These mechanisms are similar to the Ras protein
system in which the Ras proteins activated at the cytoplasmic surface of the plasma membrane recruit their
downstream effector, Raf protein kinases (389, 687).
However, it is unknown how these activated Rab proteins
determine their specific localizations. They could be physically associated with membrane phospholipid through
their COOH-terminal prenyl moieties (27, 322, 340, 349,
568, 765), but lipid-lipid interactions alone presumably do
not determine their specific localizations. It also remains
unknown how these GEP activities are regulated.
The situation is similar for Rho/Rac/Cdc42 proteins.
Most Rho/Rac/Cdc42 proteins stay in the cytosol in the
GDP-bound form complexed with Rho GDIs (702, 704).
Upon activation by their specific GEPs, they are translocated to their functioning sites. Some GEPs, such as
Tiam1 for Rac proteins and frabin for Cdc42, require
specific localizations through the membrane or F-actinbinding domains for their physiological functions (469,

January 2001

SMALL G PROTEINS

738). When dominant active mutants of Rac or Cdc42

proteins are overexpressed, they could be artificially distributed in the cell, and some of them are located at the
real functioning site and show similar effects there. Under
physiological conditions, small amounts of Rac or Cdc42
proteins activated by their GEPs at specific sites are
involved in their functions. In many literatures, overexpressed dominant active mutants of these proteins show
a variety of phenotypes, but we should be careful for their
interpretations. It is of crucial importance to understand
the mechanisms of the temporal and spatial activation
and inactivation of these small G proteins.

14.

15.

16.

17.

18.

We thank Drs. Yoshito Kajiro, Kenichi Kariya, Tohru

Kataoka, Akira Kikuchi, Akihiko Nakano, Kazuhisa Nakayama,
Takeharu Nishimoto, Fuyuhiko Tamanoi, Kazuma Tanaka, and
Yoshihiro Yoneda for their valuable comments on the manuscript.
Address for reprint requests and other correspondence: Y.
Takai, Dept. of Molecular Biology and Biochemistry, Osaka
University Graduate School of Medicine/Faculty of Medicine,
Suita 565 0871, Japan (E-mail: ytakai@molbio.med.osaka-u.ac.jp).

19.

20.

21.

REFERENCES
1. ABO A, BOYHAN A, WEST I, THRASHER AJ, AND SEGAL AW. Reconstitution of neutrophil NADPH oxidase activity in the cell-free system
by four components: p67-phox, p47-phox, p21rac1, and cytochrome
b-245. J Biol Chem 267: 1676716770, 1992.
2. ABO A, PICK E, HALL A, TOTTY N, TEAHAN CG, AND SEGAL AW.
Activation of the NADPH oxidase involves the small GTP-binding
protein p21rac1. Nature 353: 668 670, 1991.
3. ADAM SA. Transport pathways of macromolecules between the
nucleus and the cytoplasm. Curr Opin Cell Biol 11: 402 406, 1999.
4. ADAM T, GIRY M, BOQUET P, AND SANSONETTI P. Rho-dependent
membrane folding causes Shigella entry into epithelial cells. EMBO
J 15: 33153321, 1996.
5. ADAMS AE, JOHNSON DI, LONGNECKER RM, SLOAT BF, AND PRINGLE JR.
CDC42 and CDC43, two additional genes involved in budding and
the establishment of cell polarity in the yeast Saccharomyces cerevisiae. J Cell Biol 111: 131142, 1990.
6. AKTORIES K, ROSENER S, BLASCHKE U, AND CHHATWAL GS. Botulinum
ADP-ribosyltransferase C3. Purification of the enzyme and characterization of the ADP-ribosylation reaction in platelet membranes.
Eur J Biochem 172: 445 450, 1988.
7. ALBERT S AND GALLWITZ D. Two new members of a family of Ypt/Rab
GTPase activating proteins. Promiscuity of substrate recognition.
J Biol Chem 274: 33186 33189, 1999.
8. ALLEN LA AND ADEREM A. Molecular definition of distinct cytoskeletal structures involved in complement- and Fc receptor-mediated
phagocytosis in macrophages. J Exp Med 184: 627 637, 1996.
9. ALMOGUERA C, SHIBATA D, FORRESTER K, MARTIN J, ARNHEIM N, AND
PERUCHO M. Most human carcinomas of the exocrine pancreas
contain mutant c-K-ras genes. Cell 53: 549 554, 1988.
10. ALTSCHULER D AND LAPETINA EG. Mutational analysis of the cAMPdependent protein kinase-mediated phosphorylation site of Rap1b.
J Biol Chem 268: 75277531, 1993.
11. ALTSCHULER DL AND RIBEIRO-NETO F. Mitogenic and oncogenic properties of the small G protein Rap1b. Proc Natl Acad Sci USA 95:
74757479, 1998.
12. ALTSCHULER Y, LIU S, KATZ L, TANG K, HARDY S, BRODSKY F, APODACA
G, AND MOSTOV K. ADP-ribosylation factor 6 and endocytosis at the
apical surface of Madin-Darby canine kidney cells. J Cell Biol 147:
712, 1999.
13. AMANO M, ITO M, KIMURA K, FUKATA Y, CHIHARA K, NAKANO T, MATSUURA Y, AND KAIBUCHI K. Phosphorylation and activation of myosin

22.

23.

24.

25.

26.

27.

28.

29.

30.

31.

32.

33.

189

by Rho-associated kinase (Rho-kinase). J Biol Chem 271: 20246

20249, 1996.
AMANO M, MUKAI H, ONO Y, CHIHARA K, MATSUI T, HAMAJIMA Y, OKAWA
K, IWAMATSU A, AND KAIBUCHI K. Identification of a putative target for
Rho as the serine-threonine kinase protein kinase N. Science 271:
648 650, 1996.
AMBERG DC, ZAHNER JE, MULHOLLAND JW, PRINGLE JR, AND BOTSTEIN
D. Aip3p/Bud6p, a yeast actin-interacting protein that is involved in
morphogenesis and the selection of bipolar budding sites. Mol Biol
Cell 8: 729 753, 1997.
AMOR JC, HARRISON DH, KAHN RA, AND RINGE D. Structure of the
human ADP-ribosylation factor 1 complexed with GDP. Nature
372: 704 708, 1994.
ANDERSON D, KOCH CA, GREY L, ELLIS C, MORAN MF, AND PAWSON T.
Binding of SH2 domains of phospholipase C-1, GAP, and Src to
activated growth factor receptors. Science 250: 979 982, 1990.
ANDO S, KAIBUCHI K, SASAKI T, HIRAOKA K, NISHIYAMA T, MIZUNO T,
ASADA M, NUNOI H, MATSUDA I, AND MATSUURA Y. Posttranslational
processing of rac p21s is important both for their interaction with
the GDP/GTP exchange proteins and for their activation of NADPH
oxidase. J Biol Chem 267: 25709 25713, 1992.
ANDREEV J, SIMON JP, SABATINI DD, KAM J, PLOWMAN G, RANDAZZO PA,
AND SCHLESSINGER J. Identification of a new Pyk2 target protein with
Arf-GAP activity. Mol Cell Biol 19: 2338 2350, 1999.
ANDRES DA, SEABRA MC, BROWN MS, ARMSTRONG SA, SMELAND TE,
CREMERS FP, AND GOLDSTEIN JL. cDNA cloning of component A of
Rab geranylgeranyl transferase and demonstration of its role as a
Rab escort protein. Cell 73: 10911099, 1993.
ANTONNY B, BERAUD-DUFOUR S, CHARDIN P, AND CHABRE M. N-terminal
hydrophobic residues of the G-protein ADP-ribosylation factor-1
insert into membrane phospholipids upon GDP to GTP exchange.
Biochemistry 36: 4675 4684, 1997.
ANTONNY B, HUBER I, PARIS S, CHABRE M, AND CASSEL D. Activation of
ADP-ribosylation factor 1 GTPase-activating protein by phosphatidylcholine-derived diacylglycerols. J Biol Chem 272: 30848 30851,
1997.
AOE T, CUKIERMAN E, LEE A, CASSEL D, PETERS PJ, AND HSU VW. The
KDEL receptor, ERD2, regulates intracellular traffic by recruiting a
GTPase-activating protein for ARF1. EMBO J 16: 73057316, 1997.
AOE T, LEE AJ, VAN DONSELAAR E, PETERS PJ, AND HSU VW. Modulation of intracellular transport by transported proteins: insight from
regulation of COPI-mediated transport. Proc Natl Acad Sci USA 95:
1624 1629, 1998.
APOLLONI A, PRIOR IA, LINDSAY M, PARTON RG, AND HANCOCK JF. H-ras
but not K-ras traffics to the plasma membrane through the exocytic
pathway. Mol Cell Biol 20: 24752487, 2000.
ARAKI K, NAKANISHI H, HIRANO H, KATO M, SASAKI T, AND TAKAI Y.
Purification and characterization of Rab GDI from rat brain. Biochem Biophys Res Commun 211: 296 305, 1995.
ARAKI S, KAIBUCHI K, SASAKI T, HATA Y, AND TAKAI Y. Role of the
C-terminal region of smg p25A in its interaction with membranes
and the GDP/GTP exchange protein. Mol Cell Biol 11: 1438 1447,
1991.
ARAKI S, KIKUCHI A, HATA Y, ISOMURA M, AND TAKAI Y. Regulation of
reversible binding of smg p25A, a ras p21-like GTP-binding protein,
to synaptic plasma membranes and vesicles by its specific regulatory protein, GDP dissociation inhibitor. J Biol Chem 265: 13007
13015, 1990.
ARMFIELD K, NELSON R, LUBS HA, HA NE B, SCHROER RJ, ARENA F,
SCHWARTZ CE, AND STEVENSON RE. X-linked mental retardation syndrome with short stature, small hands and feet, seizures, cleft
palate, and glaucoma is linked to Xq28. Am J Med Genet 85:
236 242, 1999.
ARPIN M, ALGRAIN M, AND LOUVARD D. Membrane-actin microfilament
connections: an increasing diversity of players related to band 4.1.
Curr Opin Cell Biol 6: 136 141, 1994.
ASPENSTROM P, LINDBERG U, AND HALL A. Two GTPases, Cdc42 and
Rac, bind directly to a protein implicated in the immunodeficiency
disorder Wiskott-Aldrich syndrome. Curr Biol 6: 70 75, 1996.
AWASAKI T, SAITO M, SONE M, SUZUKI E, SAKAI R, ITO K, AND HAMA C.
The Drosophila trio plays an essential role in patterning of axons
by regulating their directional extension. Neuron 26: 119 131, 2000.
BAKER D, WUESTEHUBE L, SCHEKMAN R, BOTSTEIN D, AND SEGEV N.

190

TAKAI, SASAKI, AND MATOZAKI

GTP-binding Ypt1 protein and Ca2 function independently in a

cell-free protein transport reaction. Proc Natl Acad Sci USA 87:
355359, 1990.
34. BALLESTER R, MARCHUK D, BOGUSKI M, SAULINO A, LETCHER R, WIGLER
M, AND COLLINS F. The NF1 locus encodes a protein functionally
related to mammalian GAP and yeast IRA proteins. Cell 63: 851
859, 1990.
35. BANNYKH SI AND BALCH WE. Selective transport of cargo between
the endoplasmic reticulum and Golgi compartments. Histochem
Cell Biol 109: 463 475, 1998.
36. BARFOD ET, ZHENG Y, KUANG WJ, HART MJ, EVANS T, CERIONE RA, AND
ASHKENAZI A. Cloning and expression of a human CDC42 GTPaseactivating protein reveals a functional SH3-binding domain. J Biol
Chem 268: 26059 26062, 1993.
37. BARKER KT AND CROMPTON MR. Ras-related TC21 is activated by
mutation in a breast cancer cell line, but infrequently in breast
carcinomas in vivo. Br J Cancer 78: 296 300, 1998.
38. BARLOWE C. COPII and selective export from the endoplasmic
reticulum. Biochim Biophys Acta 1404: 6776, 1998.
39. BARLOWE C, ORCI L, YEUNG T, HOSOBUCHI M, HAMAMOTO S, SALAMA N,
REXACH MF, RAVAZZOLA M, AMHERDT M, AND SCHEKMAN R. COPII: a
membrane coat formed by Sec proteins that drive vesicle budding
from the endoplasmic reticulum. Cell 77: 895907, 1994.
40. BARLOWE C AND SCHEKMAN R. SEC12 encodes a guanine-nucleotideexchange factor essential for transport vesicle budding from the
ER. Nature 365: 347349, 1993.
41. BAR-SAGI D AND FERAMISCO JR. Microinjection of the ras oncogene
protein into PC12 cells induces morphological differentiation. Cell
42: 841 848, 1985.
41a.BAR-SAGI D AND FERAMISCO JR. Induction of membrane ruffling and
fluid-phase pinocytosis in quiescent fibroblasts by ras proteins.
Science 233: 10611068, 1986.
42. BASU TN, GUTMANN DH, FLETCHER JA, GLOVER TW, COLLINS FS, AND
DOWNWARD J. Aberrant regulation of ras proteins in malignant tumour cells from type 1 neurofibromatosis patients. Nature 356:
713715, 1992.
43. BATEMAN J, SHU H, AND VAN VACTOR D. The guanine nucleotide
exchange factor trio mediates axonal development in the Drosophila embryo. Neuron 26: 93106, 2000.
44. BEDNAREK SY, ORCI L, AND SCHEKMAN R. Traffic COPs and the formation of vesicle coats. Trends Cell Biol 6: 468 473, 1996.
45. BENDER A AND PRINGLE JR. Multicopy suppression of the cdc24
budding defect in yeast by CDC42 and three newly identified genes
including the ras-related gene RSR1. Proc Natl Acad Sci USA 86:
9976 9980, 1989.
46. BENLI M, DORING F, ROBINSON DG, YANG X, AND GALLWITZ D. Two
GTPase isoforms, Ypt31p and Ypt32p, are essential for Golgi function in yeast. EMBO J 15: 6460 6475, 1996.
47. BERAUD-DUFOUR S, PARIS S, CHABRE M, AND ANTONNY B. Dual interaction of ADP ribosylation factor 1 with Sec7 domain and with lipid
membranes during catalysis of guanine nucleotide exchange. J Biol
Chem 274: 37629 37636, 1999.
48. BHULLAR RP AND SENEVIRATNE HD. Characterization of human platelet GTPase activating protein for the Ral GTP-binding protein.
Biochim Biophys Acta 1311: 181188, 1996.
49. BI E AND ZIGMOND SH. Actin polymerization: where the WASP
stings. Curr Biol 9: R160 R163, 1999.
50. BISCHOFF FR AND GORLICH D. RanBP1 is crucial for the release of
RanGTP from importin -related nuclear transport factors. FEBS
Lett 419: 249 254, 1997.
51. BISCHOFF FR, KLEBE C, KRETSCHMER J, WITTINGHOFER A, AND PONSTINGL H. RanGAP1 induces GTPase activity of nuclear Ras-related
Ran. Proc Natl Acad Sci USA 91: 25872591, 1994.
52. BISCHOFF FR, KREBBER H, KEMPF T, HERMES I, AND PONSTINGL H.
Human RanGTPase-activating protein RanGAP1 is a homologue of
yeast Rna1p involved in mRNA processing and transport. Proc Natl
Acad Sci USA 92: 1749 1753, 1995.
53. BISCHOFF FR, KREBBER H, SMIRNOVA E, DONG W, AND PONSTINGL H.
Co-activation of RanGTPase and inhibition of GTP dissociation by
Ran-GTP binding protein RanBP1. EMBO J 14: 705715, 1995.
54. BISCHOFF FR AND PONSTINGL H. Catalysis of guanine nucleotide
exchange on Ran by the mitotic regulator RCC1. Nature 354:
80 82, 1991.

Volume 81

55. BLOCK C, JANKNECHT R, HERRMANN C, NASSAR N, AND WITTINGHOFER A.

Quantitative structure-activity analysis correlating Ras/Raf interaction in vitro to Raf activation in vivo. Nat Struct Biol 3: 244 251,
1996.
56. BOGUSKI MS AND MCCORMICK F. Proteins regulating Ras and its
relatives. Nature 366: 643 654, 1993.
57. BOKOCH GM, VLAHOS CJ, WANG Y, KNAUS UG, AND TRAYNOR-KAPLAN
AE. Rac GTPase interacts specifically with phosphatidylinositol
3-kinase. Biochem J 315: 775779, 1996.
58. BOS JL. All in the family? New insights and questions regarding
interconnectivity of Ras, Rap1 and Ral. EMBO J 17: 6776 6782,
1998.
59. BOS JL, FEARON ER, HAMILTON SR, VERLAAN-DE VRIES M, VAN BOOM JH,
VAN DER EB AJ, AND VOGELSTEIN B. Prevalence of ras gene mutations
in human colorectal cancers. Nature 327: 293297, 1987.
60. BOURNE HR, SANDERS DA, AND MCCORMICK F. The GTPase superfamily: a conserved switch for diverse cell functions. Nature 348:
125132, 1990.
61. BOURNE HR, SANDERS DA, AND MCCORMICK F. The GTPase superfamily: conserved structure and molecular mechanism. Nature 349:
117127, 1991.
62. BOWTELL D, FU P, SIMON M, AND SENIOR P. Identification of murine
homologues of the Drosophila son of sevenless gene: potential
activators of ras. Proc Natl Acad Sci USA 89: 6511 6515, 1992.
63. BRENNWALD P, KEARNS B, CHAMPION K, KERANEN S, BANKAITIS V, AND
NOVICK P. Sec9 is a SNAP-25-like component of a yeast SNARE
complex that may be the effector of Sec4 function in exocytosis.
Cell 79: 245258, 1994.
64. BRENNWALD P AND NOVICK P. Interactions of three domains distinguishing the Ras-related GTP-binding proteins Ypt1 and Sec4. Nature 362: 560 563, 1993.
65. BRILL S, LI S, LYMAN CW, CHURCH DM, WASMUTH JJ, WEISSBACH L,
BERNARDS A, AND SNIJDERS AJ. The Ras GTPase-activating-proteinrelated human protein IQGAP2 harbors a potential actin binding
domain and interacts with calmodulin and Rho family GTPases.
Mol Cell Biol 16: 4869 4878, 1996.
66. BROEK D, SAMIY N, FASANO O, FUJIYAMA A, TAMANOI F, NORTHUP J, AND
WIGLER M. Differential activation of yeast adenylate cyclase by
wild-type and mutant RAS proteins. Cell 41: 763769, 1985.
67. BROEK D, TODA T, MICHAELI T, LEVIN L, BIRCHMEIER C, ZOLLER M,
POWERS S, AND WIGLER M. The S. cerevisiae CDC25 gene product
regulates the RAS/adenylate cyclase pathway. Cell 48: 789 799,
1987.
68. BRONDYK WH, MCKIERNAN CJ, FORTNER KA, STABILA P, HOLZ RW, AND
MACARA IG. Interaction cloning of Rabin3, a novel protein that
associates with the Ras-like GTPase Rab3A. Mol Cell Biol 15:
11371143, 1995.
69. BROWN AM, OSULLIVAN AJ, AND GOMPERTS BD. Induction of exocytosis from permeabilized mast cells by the guanosine triphosphatases Rac and Cdc42. Mol Biol Cell 9: 10531063, 1998.
70. BROWN MT, ANDRADE J, RADHAKRISHNA H, DONALDSON JG, COOPER JA,
AND RANDAZZO PA. ASAP1, a phospholipid-dependent arf GTPaseactivating protein that associates with and is phosphorylated by
Src. Mol Cell Biol 18: 7038 7051, 1998.
71. BROWN R, MARSHALL CJ, PENNIE SG, AND HALL A. Mechanism of
activation of an N-ras gene in the human fibrosarcoma cell line
HT1080. EMBO J 3: 13211326, 1984.
72. BRTVA TR, DRUGAN JK, GHOSH S, TERRELL RS, CAMPBELL-BURK S, BELL
RM, AND DER CJ. Two distinct Raf domains mediate interaction
with Ras. J Biol Chem 270: 9809 9812, 1995.
73. BUCCI C, PARTON RG, MATHER IH, STUNNENBERG H, SIMONS K, HOFLACK
B, AND ZERIAL M. The small GTPase rab5 functions as a regulatory
factor in the early endocytic pathway. Cell 70: 715728, 1992.
74. BUCHSBAUM R, TELLIEZ JB, GOONESEKERA S, AND FEIG LA. The Nterminal pleckstrin, coiled-coil, and IQ domains of the exchange
factor Ras-GRF act cooperatively to facilitate activation by calcium. Mol Cell Biol 16: 4888 4896, 1996.
75. BUDAY L AND DOWNWARD J. Epidermal growth factor regulates
p21ras through the formation of a complex of receptor, Grb2
adapter protein, and Sos nucleotide exchange factor. Cell 73: 611
620, 1993.
76. BURBELO PD, SNOW DM, BAHOU W, AND SPIEGEL S. MSE55, a Cdc42

January 2001

77.

78.

79.

80.

81.

82.

83.

84.

85.

86.

87.

88.

89.
90.
91.

92.

93.
94.

95.

96.

97.

98.

SMALL G PROTEINS

effector protein, induces long cellular extensions in fibroblasts.

Proc Natl Acad Sci USA 96: 90839088, 1999.
BURD CG, MUSTOL PA, SCHU PV, AND EMR SD. A yeast protein related
to a mammalian Ras-binding protein, Vps9p, is required for localization of vacuolar proteins. Mol Cell Biol 16: 2369 2377, 1996.
BURNS ME, SASAKI T, TAKAI Y, AND AUGUSTINE GJ. Rabphilin-3A: a
multifunctional regulator of synaptic vesicle traffic. J Gen Physiol
111: 243255, 1998.
BURTON J, ROBERTS D, MONTALDI M, NOVICK P, AND DE CAMILLI P. A
mammalian guanine-nucleotide-releasing protein enhances function of yeast secretory protein Sec4. Nature 361: 464 467, 1993.
CABIB E, DRGONOVA J, AND DRGON T. Role of small G proteins in yeast
cell polarization and wall biosynthesis. Annu Rev Biochem 67:
307333, 1998.
CAMONIS JH, KALEKINE M, GONDRE B, GARREAU H, BOY-MARCOTTE E,
AND JACQUET M. Characterization, cloning and sequence analysis of
the CDC25 gene which controls the cyclic AMP level of Saccharomyces cerevisiae. EMBO J 5: 375380, 1986.
CAMPBELL SL, KHOSRAVI-FAR R, ROSSMAN KL, CLARK GJ, AND DER CJ.
Increasing complexity of Ras signaling. Oncogene 17: 13951413,
1998.
CAO X, BALLEW N, AND BARLOWE C. Initial docking of ER-derived
vesicles requires Uso1p and Ypt1p but is independent of SNARE
proteins. EMBO J 17: 2156 2165, 1998.
CAPON DJ, SEEBURG PH, MCGRATH JP, HAYFLICK JS, EDMAN U, LEVINSON AD, AND GOEDDEL DV. Activation of Ki-ras2 gene in human
colon and lung carcinomas by two different point mutations. Nature 304: 507513, 1983.
CARAZO-SALAS RE, GUARGUAGLINI G, GRUSS OJ, SEGREF A, KARSENTI E,
AND MATTAJ IW. Generation of GTP-bound Ran by RCC1 is required
for chromatin-induced mitotic spindle formation. Nature 400: 178
181, 1999.
CARON E AND HALL A. Identification of two distinct mechanisms of
phagocytosis controlled by different Rho GTPases. Science 282:
17171721, 1998.
CARPINO N, WISNIEWSKI D, STRIFE A, MARSHAK D, KOBAYASHI R, STILLdok
MAN B, AND CLARKSON B. p62
: a constitutively tyrosine-phosphorylated, GAP-associated protein in chronic myelogenous leukemia
progenitor cells. Cell 88: 197204, 1997.
CASANOVA JE, WANG X, KUMAR R, BHARTUR SG, NAVARRE J, WOODRUM
JE, ALTSCHULER Y, RAY GS, AND GOLDENRING JR. Association of
Rab25 and Rab11a with the apical recycling system of polarized
Madin-Darby canine kidney cells. Mol Biol Cell 10: 47 61, 1999.
CASEY PJ AND SEABRA MC. Protein prenyltransferases. J Biol Chem
271: 5289 5292, 1996.
CASEY PJ, SOLSKI PA, DER CJ, AND BUSS JE. p21ras is modified by a
farnesyl isoprenoid. Proc Natl Acad Sci USA 86: 8323 8327, 1989.
CASTILLO PE, JANZ R, SUDHOF TC, TZOUNOPOULOS T, MALENKA RC, AND
NICOLL RA. Rab3A is essential for mossy fibre long-term potentiation in the hippocampus. Nature 388: 590 593, 1997.
CASTRILLON DH AND WASSERMAN SA. Diaphanous is required for
cytokinesis in Drosophila and shares domains of similarity with the
products of the limb deformity gene. Development 120: 33673377,
1994.
CERIONE RA AND ZHENG Y. The Dbl family of oncogenes. Curr Opin
Cell Biol 8: 216 222, 1996.
CHAN AM, MIKI T, MEYERS KA, AND AARONSON SA. A human oncogene
of the RAS superfamily unmasked by expression cDNA cloning.
Proc Natl Acad Sci USA 91: 7558 7562, 1994.
CHANG JH, GILL S, SETTLEMAN J, AND PARSONS SJ. c-Src regulates the
simultaneous rearrangement of actin cytoskeleton, p190RhoGAP,
and p120RasGAP following epidermal growth factor stimulation.
J Cell Biol 130: 355368, 1995.
CHANT J, CORRADO K, PRINGLE JR, AND HERSKOWITZ I. Yeast BUD5,
encoding a putative GDP-GTP exchange factor, is necessary for
bud site selection and interacts with bud formation gene BEM1.
Cell 65: 12131224, 1991.
CHARDIN P, BOQUET P, MADAULE P, POPOFF MR, RUBIN EJ, AND GILL
DM. The mammalian G protein rhoC is ADP-ribosylated by Clostridium botulinum exoenzyme C3 and affects actin microfilaments in Vero cells. EMBO J 8: 10871092, 1989.
CHARDIN P, CAMONIS JH, GALE NW, VAN AELST L, SCHLESSINGER J,
WIGLER MH, AND BAR-SAGI D. Human Sos1: a guanine nucleotide

99.

100.

101.
102.

103.
104.

105.

106.

107.

108.

109.

110.

111.

112.

113.

114.

115.

116.

117.

118.

119.

120.

191

exchange factor for Ras that binds to GRB2. Science 260: 1338
1343, 1993.
CHARDIN P, PARIS S, ANTONNY B, ROBINEAU S, BERAUD-DUFOUR S,
JACKSON CL, AND CHABRE M. A human exchange factor for ARF
contains Sec7- and pleckstrin-homology domains. Nature 384: 481
484, 1996.
CHARDIN P AND TAVITIAN A. The ral gene: a new ras related gene
isolated by the use of a synthetic probe. EMBO J 5: 22032208,
1986.
CHARDIN P AND TAVITIAN A. Coding sequences of human ralA and
ralB cDNAs. Nucleic Acids Res 17: 4380, 1989.
CHAVRIER P, GORVEL JP, STELZER E, SIMONS K, GRUENBERG J, AND
ZERIAL M. Hypervariable C-terminal domain of rab proteins acts as
a targeting signal. Nature 353: 769 772, 1991.
CHAVRIER P AND GOUD B. The role of ARF and Rab GTPases in
membrane transport. Curr Opin Cell Biol 11: 466 475, 1999.
CHAVRIER P, PARTON RG, HAURI HP, SIMONS K, AND ZERIAL M. Localization of low molecular weight GTP binding proteins to exocytic
and endocytic compartments. Cell 62: 317329, 1990.
CHAVRIER P, VINGRON M, SANDER C, SIMONS K, AND ZERIAL M. Molecular cloning of YPT1/SEC4-related cDNAs from an epithelial cell
line. Mol Cell Biol 10: 6578 6585, 1990.
CHEN GC, KIM YJ, AND CHAN CS. The Cdc42 GTPase-associated
proteins Gic1 and Gic2 are required for polarized cell growth in
Saccharomyces cerevisiae. Genes Dev 11: 2958 2971, 1997.
CHEN LM, HOBBIE S, AND GALAN JE. Requirement of CDC42 for
Salmonella-induced cytoskeletal and nuclear responses. Science
274: 21152118, 1996.
CHEN W, CHEN S, YAP SF, AND LIM L. The Caenorhabditis elegans
p21-activated kinase (CePAK) colocalizes with CeRac1 and
CDC42Ce at hypodermal cell boundaries during embryo elongation. J Biol Chem 271: 2636226368, 1996.
CHEN W, FENG Y, CHEN D, AND WANDINGER-NESS A. Rab11 is required
for trans-Golgi network-to-plasma membrane transport and a preferential target for GDP dissociation inhibitor. Mol Biol Cell 9:
32413257, 1998.
CHERFILS J AND CHARDIN P. GEFs: structural basis for their activation
of small GTP-binding proteins. Trends Biochem Sci 24: 306 311,
1999.
CHIEN UH, LAI M, SHIH TY, VERMA IM, SCOLNICK EM, ROY-BURMAN P,
AND DAVIDSON N. Heteroduplex analysis of the sequence relationships between the genomes of Kirsten and Harvey sarcoma viruses,
their respective parental murine leukemia viruses, and the rat
endogenous 30S RNA. J Virol 31: 752760, 1979.
CHONG LD, TRAYNOR-KAPLAN A, BOKOCH GM, AND SCHWARTZ MA. The
small GTP-binding protein Rho regulates a phosphatidylinositol
4-phosphate 5-kinase in mammalian cells. Cell 79: 507513, 1994.
CHOU MM AND BLENIS J. The 70 kDa S6 kinase complexes with and
is activated by the Rho family G proteins Cdc42 and Rac1. Cell 85:
573583, 1996.
CHOY E, CHIU VK, SILLETTI J, FEOKTISTOV M, MORIMOTO T, MICHAELSON
D, IVANOV IE, AND PHILIPS MR. Endomembrane trafficking of ras: the
CAAX motif targets proteins to the ER and Golgi. Cell 98: 69 80,
1999.
CHRISTOFORIDIS S, MCBRIDE HM, BURGOYNE RD, AND ZERIAL M. The
Rab5 effector EEA1 is a core component of endosome docking.
Nature 397: 621 625, 1999.
CHUANG TH, XU X, KNAUS UG, HART MJ, AND BOKOCH GM. GDP
dissociation inhibitor prevents intrinsic and GTPase activating protein-stimulated GTP hydrolysis by the Rac GTP-binding protein.
J Biol Chem 268: 775778, 1993.
CHUNG SH, TAKAI Y, AND HOLZ RW. Evidence that the Rab3a-binding
protein, rabphilin3a, enhances regulated secretion. Studies in adrenal chromaffin cells. J Biol Chem 270: 16714 16718, 1995.
CICCHETTI P, MAYER BJ, THIEL G, AND BALTIMORE D. Identification of
a protein that binds to the SH3 region of Abl and is similar to Bcr
and GAP-rho. Science 257: 803 806, 1992.
CID VJ, DURAN A, DEL REY F, SNYDER MP, NOMBELA C, AND SANCHEZ M.
Molecular basis of cell integrity and morphogenesis in Saccharomyces cerevisiae. Microbiol Rev 59: 345386, 1995.
CLAING A, PERRY SJ, ACHIRILOAIE M, WALKER JK, ALBANESI JP, LEFKOWITZ RJ, AND PREMONT RT. Multiple endocytic pathways of G protein-

192

121.

122.

123.

124.

125.

126.

127.

128.

129.

130.

131.

132.

133.

134.

135.

136.

137.

138.

139.
140.

TAKAI, SASAKI, AND MATOZAKI

coupled receptors delineated by GIT1 sensitivity. Proc Natl Acad
Sci USA 97: 1119 1124, 2000.
CLARK GJ, DRUGAN JK, TERRELL RS, BRADHAM C, DER CJ, BELL RM,
AND CAMPBELL S. Peptides containing a consensus Ras binding
sequence from Raf-1 and the GTPase activating protein NF1 inhibit
Ras function. Proc Natl Acad Sci USA 93: 15771581, 1996.
CLAUDE A, ZHAO BP, KUZIEMSKY CE, DAHAN S, BERGER SJ, YAN JP,
ARMOLD AD, SULLIVAN EM, AND MELANCON P. GBF1: a novel Golgiassociated BFA-resistant guanine nucleotide exchange factor that
displays specificity for ADP-ribosylation factor 5. J Cell Biol 146:
71 84, 1999.
COHEN L, MOHR R, CHEN YY, HUANG M, KATO R, DORIN D, TAMANOI F,
GOGA A, AFAR D, AND ROSENBERG N. Transcriptional activation of a
ras-like gene (kir) by oncogenic tyrosine kinases. Proc Natl Acad
Sci USA 91: 12448 12452, 1994.
COOK SJ, RUBINFELD B, ALBERT I, AND MCCORMICK F. RapV12 antagonizes Ras-dependent activation of ERK1 and ERK2 by LPA and
EGF in Rat-1 fibroblasts. EMBO J 12: 34753485, 1993.
CORBETT AH AND SILVER PA. The NTF2 gene encodes an essential,
highly conserved protein that functions in nuclear transport in vivo.
J Biol Chem 271: 1847718484, 1996.
COSO OA, CHIARIELLO M, YU JC, TERAMOTO H, CRESPO P, XU N, MIKI T,
AND GUTKIND JS. The small GTP-binding proteins Rac1 and Cdc42
regulate the activity of the JNK/SAPK signaling pathway. Cell 81:
11371146, 1995.
COUTAVAS E, REN M, OPPENHEIM JD, DEUSTACHIO P, AND RUSH MG.
Characterization of proteins that interact with the cell-cycle regulatory protein Ran/TC4. Nature 366: 585587, 1993.
COX AD, BRTVA TR, LOWE DG, AND DER CJ. R-Ras induces malignant,
but not morphologic, transformation of NIH3T3 cells. Oncogene 9:
32813288, 1994.
COX D, CHANG P, ZHANG Q, REDDY PG, BOKOCH GM, AND GREENBERG
S. Requirements for both Rac1 and Cdc42 in membrane ruffling and
phagocytosis in leukocytes. J Exp Med 186: 14871494, 1997.
CRAWFORD JM, HARDEN N, LEUNG T, LIM L, AND KIEHART DP. Cellularization in Drosophila melanogaster is disrupted by the inhibition
of rho activity and the activation of Cdc42 function. Dev Biol 204:
151164, 1998.
CREMERS FP, MOLLOY CM, VAN DE POL DJ, VAN DEN HURK JA, BACH I,
GEURTS VAN KESSEL AH, AND ROPERS HH. An autosomal homologue
of the choroideremia gene colocalizes with the Usher syndrome
type II locus on the distal part of chromosome 1q. Hum Mol Genet
1: 7175, 1992.
CREMERS FP, VAN DE POL DJ, VAN KERKHOFF LP, WIERINGA B, AND
ROPERS HH. Cloning of a gene that is rearranged in patients with
choroideraemia. Nature 347: 674 677, 1990.
CRESPO P, SCHUEBEL KE, OSTROM AA, GUTKIND JS, AND BUSTELO XR.
Phosphotyrosine-dependent activation of Rac-1 GDP/GTP exchange by the vav proto-oncogene product. Nature 385: 169 172,
1997.
CREWS CM, ALESSANDRINI A, AND ERIKSON RL. The primary structure
of MEK, a protein kinase that phosphorylates the ERK gene product. Science 258: 478 480, 1992.
CUIF MH, POSSMAYER F, ZANDER H, BORDES N, JOLLIVET F, COUEDELCOURTEILLE A, JANOUEIX-LEROSEY I, LANGSLEY G, BORNENS M, AND
GOUD B. Characterization of GAPCenA, a GTPase activating protein
for Rab6, part of which associates with the centrosome. EMBO J
18: 17721782, 1999.
CUKIERMAN E, HUBER I, ROTMAN M, AND CASSEL D. The ARF1 GTPaseactivating protein: zinc finger motif and Golgi complex localization.
Science 270: 1999 2002, 1995.
CVRCKOVA F, DE VIRGILIO C, MANSER E, PRINGLE JR, AND NASMYTH K.
Ste20-like protein kinases are required for normal localization of
cell growth and for cytokinesis in budding yeast. Genes Dev 9:
18171830, 1995.
DADAMO P, MENEGON A, LO NIGRO C, GRASSO M, GULISANO M, TAMANINI F, BIENVENU T, GEDEON AK, OOSTRA B, WU SK, TANDON A,
VALTORTA F, BALCH WE, CHELLY J, AND TONIOLO D. Mutations in GDI1
are responsible for X-linked non-specific mental retardation. Nat
Genet 19: 134 139, 1998.
DAHLBERG JE AND LUND E. Functions of the GTPase Ran in RNA
export from the nucleus. Curr Opin Cell Biol 10: 400 408, 1998.
DARCHEN F, ZAHRAOUI A, HAMMEL F, MONTEILS MP, TAVITIAN A, AND

Volume 81

SCHERMAN D. Association of the GTP-binding protein Rab3A with

bovine adrenal chromaffin granules. Proc Natl Acad Sci USA 87:
56925696, 1990.
141. DEFEO-JONES D, SCOLNICK EM, KOLLER R, AND DHAR R. ras-Related
gene sequences identified and isolated from Saccharomyces cerevisiae. Nature 306: 707709, 1983.
142. DENFERT C, BARLOWE C, NISHIKAWA S, NAKANO A, AND SCHEKMAN R.
Structural and functional dissection of a membrane glycoprotein
required for vesicle budding from the endoplasmic reticulum. Mol
Cell Biol 11: 57275734, 1991.
143. DENFERT C, WUESTEHUBE LJ, LILA T, AND SCHEKMAN R. Sec12pdependent membrane binding of the small GTP-binding protein
Sar1p promotes formation of transport vesicles from the ER. J Cell
Biol 114: 663 670, 1991.
144. DENOUEL-GALY A, DOUVILLE EM, WARNE PH, PAPIN C, LAUGIER D,
CALOTHY G, DOWNWARD J, AND EYCHENE A. Murine Ksr interacts with
MEK and inhibits Ras-induced transformation. Curr Biol 8: 46 55,
1998.
145. DENT P, HASER W, HAYSTEAD TA, VINCENT LA, ROBERTS TM, AND
STURGILL TW. Activation of mitogen-activated protein kinase kinase
by v-Raf in NIH 3T3 cells and in vitro. Science 257: 1404 1407, 1992.
146. DER CJ, KRONTIRIS TG, AND COOPER GM. Transforming genes of
human bladder and lung carcinoma cell lines are homologous to
the ras genes of Harvey and Kirsten sarcoma viruses. Proc Natl
Acad Sci USA 79: 36373640, 1982.
147. DE ROOIJ J, BOENINK NM, VAN TRIEST M, COOL RH, WITTINGHOFER A,
AND BOS JL. PDZ-GEF1, a guanine nucleotide exchange factor specific for Rap1 and Rap2. J Biol Chem 274: 3812538130, 1999.
148. DE ROOIJ J, ZWARTKRUIS FJ, VERHEIJEN MH, COOL RH, NIJMAN SM,
WITTINGHOFER A, AND BOS JL. Epac is a Rap1 guanine-nucleotideexchange factor directly activated by cyclic AMP. Nature 396:
474 477, 1998.
149. DESRIVIERES S, COOKE FT, PARKER PJ, AND HALL MN. MSS4, a phosphatidylinositol-4-phosphate 5-kinase required for organization of
the actin cytoskeleton in Saccharomyces cerevisiae. J Biol Chem
273: 1578715793, 1998.
150. DHARMAWARDHANE S, SANDERS LC, MARTIN SS, DANIELS RH, AND
BOKOCH GM. Localization of p21-activated kinase 1 (PAK1) to pinocytic vesicles and cortical actin structures in stimulated cells.
J Cell Biol 138: 12651278, 1997.
151. DIAZ E, SCHIMMOLLER F, AND PFEFFER SR. A novel Rab9 effector
required for endosome-to-TGN transport. J Cell Biol 138: 283290,
1997.
151a.DICKSON B, SPRENGER F, MORRISON D, AND HAFEN E. Raf functions
downstream of Ras1 in the Sevenless signal transduction pathway.
Nature 360: 600 603, 1992.
152. DIEKMANN D, BRILL S, GARRETT MD, TOTTY N, HSUAN J, MONFRIES C,
HALL C, LIM L, AND HALL A. Bcr encodes a GTPase-activating protein
for p21rac. Nature 351: 400 402, 1991.
153. DING M, VITALE N, TSAI SC, ADAMIK R, MOSS J, AND VAUGHAN M.
Characterization of a GTPase-activating protein that stimulates
GTP hydrolysis by both ADP-ribosylation factor (ARF) and ARFlike proteins. Comparison to the ARD1 gap domain. J Biol Chem
271: 2400524009, 1996.
154. DIRAC-SVEJSTRUP AB, SUMIZAWA T, AND PFEFFER SR. Identification of
a GDI displacement factor that releases endosomal Rab GTPases
from Rab-GDI. EMBO J. 16: 465 472, 1997.
155. DOWNWARD J. Control of ras activation. Cancer Surv 27: 87100,
1996.
156. DRECHSEL DN, HYMAN AA, HALL A, AND GLOTZER M. A requirement for
Rho and Cdc42 during cytokinesis in Xenopus embryos. Curr Biol
7: 1223, 1997.
157. DRGONOVA J, DRGON T, TANAKA K, KOLLAR R, CHEN GC, FORD RA,
CHAN CS, TAKAI Y, AND CABIB E. Rho1p, a yeast protein at the
interface between cell polarization and morphogenesis. Science
272: 277279, 1996.
158. DRIVAS GT, SHIH A, COUTAVAS E, RUSH MG, AND DEUSTACHIO P.
Characterization of four novel ras-like genes expressed in a human
teratocarcinoma cell line. Mol Cell Biol 10: 17931798, 1990.
159. DRUGAN JK, KHOSRAVI-FAR R, WHITE MA, DER CJ, SUNG YJ, HWANG
YW, AND CAMPBELL SL. Ras interaction with two distinct binding
domains in Raf-1 may be required for Ras transformation. J Biol
Chem 271: 233237, 1996.

January 2001

SMALL G PROTEINS

160. DSOUZA-SCHOREY C, BOSHANS RL, MCDONOUGH M, STAHL PD, AND VAN

AELST L. A role for POR1, a Rac1-interacting protein, in ARF6mediated cytoskeletal rearrangements. EMBO J 16: 54455454,
1997.
161. DSOUZA-SCHOREY C, LI G, COLOMBO MI, AND STAHL PD. A regulatory
role for ARF6 in receptor-mediated endocytosis. Science 267: 1175
1178, 1995.
162. DSOUZA-SCHOREY C, VAN DONSELAAR E, HSU VW, YANG C, STAHL PD,
AND PETERS PJ. ARF6 targets recycling vesicles to the plasma membrane: insights from an ultrastructural investigation. J Cell Biol 140:
603 616, 1998.
163. DU LL, COLLINS RN, AND NOVICK PJ. Identification of a Sec4p GTPase-activating protein (GAP) as a novel member of a Rab GAP
family. J Biol Chem 273: 32533256, 1998.
164. DUTARTRE H, DAVOUST J, GORVEL JP, AND CHAVRIER P. Cytokinesis
arrest and redistribution of actin-cytoskeleton regulatory components in cells expressing the Rho GTPase CDC42Hs. J Cell Sci 109:
367377, 1996.
165. EATON S, WEPF R, AND SIMONS K. Roles for Rac1 and Cdc42 in planar
polarization and hair outgrowth in the wing of Drosophila. J Cell
Biol 135: 12771289, 1996.
166. EBINU JO, BOTTORFF DA, CHAN EY, STANG SL, DUNN RJ, AND STONE JC.
RasGRP, a Ras guanyl nucleotide-releasing protein with calciumand diacylglycerol-binding motifs. Science 280: 10821086, 1998.
167. ECHARD A, JOLLIVET F, MARTINEZ O, LACAPERE JJ, ROUSSELET A, JANOUEIX-LEROSEY I, AND GOUD B. Interaction of a Golgi-associated
kinesin-like protein with Rab6. Science 279: 580 585, 1998.
168. EGAN SE, GIDDINGS BW, BROOKS MW, BUDAY L, SIZELAND AM, AND
WEINBERG RA. Association of Sos Ras exchange protein with Grb2
is implicated in tyrosine kinase signal transduction and transformation. Nature 363: 4551, 1993.
169. ELLIS C, MORAN M, MCCORMICK F, AND PAWSON T. Phosphorylation of
GAP and GAP-associated proteins by transforming and mitogenic
tyrosine kinases. Nature 343: 377381, 1990.
170. EMKEY R, FREEDMAN S, AND FEIG LA. Characterization of a GTPaseactivating protein for the Ras-related Ral protein. J Biol Chem 266:
97039706, 1991.
171. ENDO A. Chemistry, biochemistry, and pharmacology of HMG-CoA
reductase inhibitors. Klin Wochenschr 66: 421 427, 1988.
172. ERICKSON MR, GALLETTA BJ, AND ABMAYR SM. Drosophila myoblast
city encodes a conserved protein that is essential for myoblast
fusion, dorsal closure, and cytoskeletal organization. J Cell Biol
138: 589 603, 1997.
173. ERREDE B, GARTNER A, ZHOU Z, NASMYTH K, AND AMMERER G. MAP
kinase-related FUS3 from S. cerevisiae is activated by STE7 in
vitro. Nature 362: 261264, 1993.
174. ESPENSHADE P, GIMENO RE, HOLZMACHER E, TEUNG P, AND KAISER CA.
Yeast SEC16 gene encodes a multidomain vesicle coat protein that
interacts with Sec23p. J Cell Biol 131: 311324, 1995.
175. EVANGELISTA M, BLUNDELL K, LONGTINE MS, CHOW CJ, ADAMES N,
PRINGLE JR, PETER M, AND BOONE C. Bni1p, a yeast formin linking
cdc42p and the actin cytoskeleton during polarized morphogenesis.
Science 276: 118 122, 1997.
176. FANTL WJ, MUSLIN AJ, KIKUCHI A, MARTIN JA, MACNICOL AM, GROSS
RW, AND WILLIAMS LT. Activation of Raf-1 by 14 3-3 proteins. Nature 371: 612 614, 1994.
177. FARNSWORTH CC, KAWATA M, YOSHIDA Y, TAKAI Y, GELB MH, AND
GLOMSET JA. C terminus of the small GTP-binding protein smg p25A
contains two geranylgeranylated cysteine residues and a methyl
ester. Proc Natl Acad Sci USA 88: 6196 6200, 1991.
178. FARNSWORTH CL, FRESHNEY NW, ROSEN LB, GHOSH A, GREENBERG ME,
AND FEIG LA. Calcium activation of Ras mediated by neuronal
exchange factor Ras-GRF. Nature 376: 524 527, 1995.
179. FEIG LA, URANO T, AND CANTOR S. Evidence for a Ras/Ral signaling
cascade. Trends Biochem Sci 21: 438 441, 1996.
180. FELDHERR CM AND AKIN D. Role of nuclear trafficking in regulating
cellular activity. Int Rev Cytol 151: 183228, 1994.
181. FENWICK C, NA SY, VOLL RE, ZHONG H, IM SY, LEE JW, AND GHOSH S.
A subclass of ras proteins that regulate the degradation of ikappaB.
Science 287: 869 873, 2000.
182. FERAMISCO JR, GROSS M, KAMATA T, ROSENBERG M, AND SWEET RW.
Microinjection of the oncogene form of the human H-ras (T-24)

183.

184.

185.

186.

187.
188.

189.

190.

191.

192.

193.

194.
195.

196.
197.

198.

199.

200.

201.

202.

203.

193

protein results in rapid proliferation of quiescent cells. Cell 38:

109 117, 1984.
FISCHER VON MOLLARD G, MIGNERY GA, BAUMERT M, PERIN MS, HANSON
TJ, BURGER PM, JAHN R, AND SUDHOF TC. rab3 is a small GTP-binding
protein exclusively localized to synaptic vesicles. Proc Natl Acad
Sci USA 87: 1988 1992, 1990.
FISCHER VON MOLLARD G, STAHL B, KHOKHLATCHEV A, SUDHOF TC, AND
JAHN R. Rab3C is a synaptic vesicle protein that dissociates from
synaptic vesicles after stimulation of exocytosis. J Biol Chem 269:
1097110974, 1994.
FISCHER VON MOLLARD G, SUDHOF TC, AND JAHN R. A small GTPbinding protein dissociates from synaptic vesicles during exocytosis. Nature 349: 79 81, 1991.
FLOER M, BLOBEL G, AND REXACH M. Disassembly of RanGTP-karyopherin complex, an intermediate in nuclear protein import. J Biol
Chem 272: 19538 19546, 1997.
FODOR E, LEE RT, AND ODONNELL JJ. Analysis of choroideraemia
gene. Nature 351: 614, 1991.
FORNEROD M, OHNO M, YOSHIDA M, AND MATTAJ IW. CRM1 is an
export receptor for leucine-rich nuclear export signal. Cell 90:
10511060, 1997.
FORNEROD M, VAN DEURSEN J, VAN BAAL S, REYNOLDS A, DAVIS D, MURTI
KG, FRANSEN J, AND GROSVELD G. The human homologue of yeast
CRM1 is in a dynamic subcomplex with CAN/Nup214 and a novel
nuclear pore component Nup88. EMBO J 16: 807 816, 1997.
FRANCO M, CHARDIN P, CHABRE M, AND PARIS S. Myristoylation of
ADP-ribosylation factor 1 facilitates nucleotide exchange at physiological Mg2 levels. J Biol Chem 270: 13371341, 1995.
FRANCO M, PETERS PJ, BORETTO J, VAN DONSELAAR E, NERI A, DSOUZASCHOREY C, AND CHAVRIER P. EFA6, a sec7 domain-containing exchange factor for ARF6, coordinates membrane recycling and actin
cytoskeleton organization. EMBO J 18: 1480 1491, 1999.
FRANK S, UPENDER S, HANSEN SH, AND CASANOVA JE. ARNO is a
guanine nucleotide exchange factor for ADP-ribosylation factor 6.
J Biol Chem 273: 2327, 1998.
FRANK SR, HATFIELD JC, AND CASANOVA JE. Remodeling of the actin
cytoskeleton is coordinately regulated by protein kinase C and the
ADP-ribosylation factor nucleotide exchange factor ARNO. Mol
Biol Cell 9: 31333146, 1998.
FRANKE B, AKKERMAN JW, AND BOS JL. Rapid Ca2-mediated activation of Rap1 in human platelets. EMBO J 16: 252259, 1997.
FRANZUSOFF A, REDDING K, CROSBY J, FULLER RS, AND SCHEKMAN R.
Localization of components involved in protein transport and processing through the yeast Golgi apparatus. J Cell Biol 112: 2737,
1991.
FRAZIER JA AND FIELD CM. Actin cytoskeleton: are FH proteins local
organizers? Curr Biol 7: R414 R417, 1997.
FREED E, SYMONS M, MACDONALD SG, MCCORMICK F, AND RUGGIERI R.
Binding of 14 3-3 proteins to the protein kinase Raf and effects on
its activation. Science 265: 17131716, 1994.
FUJIOKA H, KAIBUCHI K, KISHI K, YAMAMOTO T, KAWAMURA M, SAKODA
T, MIZUNO T, AND TAKAI Y. Transforming and c-fos promoter/enhancer-stimulating activities of a stimulatory GDP/GTP exchange protein for small GTP-binding proteins. J Biol Chem 267: 926 930,
1992.
FUJIWARA T, TANAKA K, INOUE E, KIKYO M, AND TAKAI Y. Bni1p
regulates microtubule-dependent nuclear migration through the
actin cytoskeleton in Saccharomyces cerevisiae. Mol Cell Biol 19:
8016 8027, 1999.
FUJIWARA T, TANAKA K, MINO A, KIKYO M, TAKAHASHI K, SHIMIZU K, AND
TAKAI Y. Rho1p-Bni1p-Spa2p interactions: implication in localization of Bni1p at the bud site and regulation of the actin cytoskeleton in Saccharomyces cerevisiae. Mol Biol Cell 9: 12211233, 1998.
FUJIYAMA A AND TAMANOI F. RAS2 protein of Saccharomyces cerevisiae undergoes removal of methionine at N terminus and removal
of three amino acids at C terminus. J Biol Chem 265: 33623368,
1990.
FUKATA Y, KIMURA K, OSHIRO N, SAYA H, MATSUURA Y, AND KAIBUCHI K.
Association of the myosin-binding subunit of myosin phosphatase
and moesin: dual regulation of moesin phosphorylation by Rhoassociated kinase and myosin phosphatase. J Cell Biol 141: 409
418, 1998.
FUKUDA M, ASANO S, NAKAMURA T, ADACHI M, YOSHIDA M, YANAGIDA M,

194

204.

205.

206.

207.

208.

209.

210.

211.

212.

213.

214.

215.

216.

217.

218.

219.

220.
221.

222.

223.
224.

225.

226.

TAKAI, SASAKI, AND MATOZAKI

AND NISHIDA E. CRM1 is responsible for intracellular transport
mediated by the nuclear export signal. Nature 390: 308 311, 1997.
FUKUHARA S, MURGA C, ZOHAR M, IGISHI T, AND GUTKIND JS. A novel
PDZ domain containing guanine nucleotide exchange factor links
heterotrimeric G proteins to Rho. J Biol Chem 274: 5868 5879,
1999.
FUKUI K, SASAKI T, IMAZUMI K, MATSUURA Y, NAKANISHI H, AND TAKAI Y.
Isolation and characterization of a GTPase activating protein specific for the Rab3 subfamily of small G proteins. J Biol Chem 272:
4655 4658, 1997.
FUKUMOTO Y, KAIBUCHI K, HORI Y, FUJIOKA H, ARAKI S, UEDA T, KIKUCHI
A, AND TAKAI Y. Molecular cloning and characterization of a novel
type of regulatory protein (GDI) for the rho proteins, ras p21-like
small GTP-binding proteins. Oncogene 5: 13211328, 1990.
GALE NW, KAPLAN S, LOWENSTEIN EJ, SCHLESSINGER J, AND BAR-SAGI D.
Grb2 mediates the EGF-dependent activation of guanine nucleotide
exchange on Ras. Nature 363: 88 92, 1993.
GALLWITZ D, DONATH C, AND SANDER C. A yeast gene encoding a
protein homologous to the human c-has/bas proto-oncogene product. Nature 306: 704 707, 1983.
GAMPEL A, PARKER PJ, AND MELLOR H. Regulation of epidermal
growth factor receptor traffic by the small GTPase rhoB. Curr Biol
9: 955958, 1999.
GARCIA-RANEA JA AND VALENCIA A. Distribution and functional diversification of the ras superfamily in Saccharomyces cerevisiae.
FEBS Lett 434: 219 225, 1998.
GARRETT MD, MAJOR GN, TOTTY N, AND HALL A. Purification and
N-terminal sequence of the p21rho GTPase-activating protein, rho
GAP. Biochem J 276: 833 836, 1991.
GARRETT MD, SELF AJ, VAN OERS C, AND HALL A. Identification of
distinct cytoplasmic targets for ras/R-ras and rho regulatory proteins. J Biol Chem 264: 10 13, 1989.
GARRETT MD, ZAHNER JE, CHENEY CM, AND NOVICK PJ. GDI1 encodes
a GDP dissociation inhibitor that plays an essential role in the yeast
secretory pathway. EMBO J 13: 1718 1728, 1994.
GARRITY PA, RAO Y, SALECKER I, MCGLADE J, PAWSON T, AND ZIPURSKY
SL. Drosophila photoreceptor axon guidance and targeting requires the dreadlocks SH2/SH3 adapter protein. Cell 85: 639 650,
1996.
GEISER JR, SCHOTT EJ, KINGSBURY TJ, COLE NB, TOTIS LJ, BHATTACHARYYA G, HE L, AND HOYT MA. Saccharomyces cerevisiae genes
required in the absence of the CIN8-encoded spindle motor act in
functionally diverse mitotic pathways. Mol Biol Cell 8: 10351050,
1997.
GEPPERT M, BOLSHAKOV VY, SIEGELBAUM SA, TAKEI K, DE CAMILLI P,
HAMMER RE, AND SUDHOF TC. The role of Rab3A in neurotransmitter
release. Nature 369: 493 497, 1994.
GEPPERT M, GODA Y, STEVENS CF, AND SUDHOF TC. The small GTPbinding protein Rab3A regulates a late step in synaptic vesicle
fusion. Nature 387: 810 814, 1997.
GEPPERT M AND SUDHOF TC. RAB3 and synaptotagmin: the yin and
yang of synaptic membrane fusion. Annu Rev Neurosci 21: 7595,
1998.
GEYER M AND WITTINGHOFER A. GEFs, GAPs, GDIs and effectors:
taking a closer (3D) look at the regulation of Ras-related GTPbinding proteins. Curr Opin Struct Biol 7: 786 792, 1997.
GIBBS JB. Ras C-terminal processing enzymesnew drug targets?
Cell 65: 1 4, 1991.
GIBBS JB, SCHABER MD, ALLARD WJ, SIGAL IS, AND SCOLNICK EM.
Purification of ras GTPase activating protein from bovine brain.
Proc Natl Acad Sci USA 85: 5026 5030, 1988.
GIBBS JB, SIGAL IS, POE M, AND SCOLNICK EM. Intrinsic GTPase
activity distinguishes normal and oncogenic ras p21 molecules.
Proc Natl Acad Sci USA 81: 5704 5708, 1984.
GILMAN AG. G proteins: transducers of receptor-generated signals.
Annu Rev Biochem 56: 615 649, 1987.
GIMENO RE, ESPENSHADE P, AND KAISER CA. SED4 encodes a yeast
endoplasmic reticulum protein that binds Sec16p and participates
in vesicle formation. J Cell Biol 131: 325338, 1995.
GIMENO RE, ESPENSHADE P, AND KAISER CA. COPII coat subunit
interactions: Sec24p and Sec23p bind to adjacent regions of
Sec16p. Mol Biol Cell 7: 18151823, 1996.
GLAVEN JA, WHITEHEAD IP, NOMANBHOY T, KAY R, AND CERIONE RA.

Volume 81

Lfc and Lsc oncoproteins represent two new guanine nucleotide

exchange factors for the Rho GTP-binding protein. J Biol Chem
271: 27374 27381, 1996.
227. GLISE B AND NOSELLI S. Coupling of Jun amino-terminal kinase and
Decapentaplegic signaling pathways in Drosophila morphogenesis.
Genes Dev 11: 1738 1747, 1997.
228. GLOMSET JA AND FARNSWORTH CC. Role of protein modification
reactions in programming interactions between ras-related GTPases and cell membranes. Annu Rev Cell Biol 10: 181205, 1994.
229. GOLDBERG J. Structural basis for activation of ARF GTPase: mechanisms of guanine nucleotide exchange and GTP-myristoyl switching. Cell 95: 237248, 1998.
230. GOLDBERG J. Structural and functional analysis of the ARF1-ARFGAP complex reveals a role for coatomer in GTP hydrolysis. Cell
96: 893902, 1999.
231. GOLDSTEIN JL AND BROWN MS. Regulation of the mevalonate pathway. Nature 343: 425 430, 1990.
232. GONG MC, IIZUKA K, NIXON G, BROWNE JP, HALL A, ECCLESTON JF,
SUGAI M, KOBAYASHI S, SOMLYO AV, AND SOMLYO AP. Role of guanine
nucleotide-binding proteins-ras-family or trimeric proteins or both
in Ca2 sensitization of smooth muscle. Proc Natl Acad Sci USA 93:
1340 1345, 1996.
233. GONZALEZ L JR AND SCHELLER RH. Regulation of membrane trafficking: structural insights from a Rab/effector complex. Cell 96: 755
758, 1999.
234. GORLICH D. Transport into and out of the cell nucleus. EMBO J 17:
27212727, 1998.
235. GORLICH D, DABROWSKI M, BISCHOFF FR, KUTAY U, BORK P, HARTMANN
E, PREHN S, AND IZAURRALDE E. A novel class of RanGTP binding
proteins. J Cell Biol 138: 65 80, 1997.
236. GORLICH D, PANTE N, KUTAY U, AEBI U, AND BISCHOFF FR. Identification of different roles for RanGDP and RanGTP in nuclear protein
import. EMBO J 15: 5584 5594, 1996.
237. GORVEL JP, CHAVRIER P, ZERIAL M, AND GRUENBERG J. rab5 controls
early endosome fusion in vitro. Cell 64: 915925, 1991.
238. GOSSER YQ, NOMANBHOY TK, AGHAZADEH B, MANOR D, COMBS C,
CERIONE RA, AND ROSEN MK. C-terminal binding domain of Rho
GDP-dissociation inhibitor directs N-terminal inhibitory peptide to
GTPases. Nature 387: 814 819, 1997.
239. GOTOH T, HATTORI S, NAKAMURA S, KITAYAMA H, NODA M, TAKAI Y,
KAIBUCHI K, MATSUI H, HATASE O, AND TAKAHASHI H. Identification of
Rap1 as a target for the Crk SH3 domain-binding guanine nucleotide-releasing factor C3G. Mol Cell Biol 15: 6746 6753, 1995.
240. GOUD B, ZAHRAOUI A, TAVITIAN A, AND SARASTE J. Small GTP-binding
protein associated with Golgi cisternae. Nature 345: 553556, 1990.
241. GOURNIER H, STENMARK H, RYBIN V, LIPPE R, AND ZERIAL M. Two
distinct effectors of the small GTPase Rab5 cooperate in endocytic
membrane fusion. EMBO J 17: 1930 1940, 1998.
242. GRAHAM SM, VOJTEK AB, HUFF SY, COX AD, CLARK GJ, COOPER JA,
AND DER CJ. TC21 causes transformation by Raf-independent signaling pathways. Mol Cell Biol 16: 6132 6140, 1996.
243. GRAVES JD, DOWNWARD J, IZQUIERDO-PASTOR M, RAYTER S, WARNE PH,
AND CANTRELL DA. The growth factor IL-2 activates p21ras proteins
in normal human T lymphocytes. J Immunol 148: 24172422, 1992.
243a.GRINDSTAFF KK, YEAMAN C, ANANDASABAPATHY N, HSU SC, RODRIGUEZBOULAN E, SCHELLER RH, AND NELSON WJ. Sec6/8 complex is recruited to cell-cell contacts and specifies transport vesicle delivery
to the basal-lateral membrane in epithelial cells. Cell 93: 731740,
1998.
244. GUILLEMOT JC, KRUSKAL BA, ADRA CN, ZHU S, KO JL, BURCH P, NOCKA
K, SEETOO K, SIMONS E, AND LIM B. Targeted disruption of guanosine diphosphate-dissociation inhibitor for Rho-related proteins,
GDID4: normal hematopoietic differentiation but subtle defect in
superoxide production by macrophages derived from in vitro embryonal stem cell differentiation. Blood 88: 27222731, 1996.
245. GULBINS E, COGGESHALL KM, BAIER G, KATZAV S, BURN P, AND ALTMAN
A. Tyrosine kinase-stimulated guanine nucleotide exchange activity
of Vav in T cell activation. Science 260: 822 825, 1993.
246. GUO W, ROTH D, WALCH-SOLIMENA C, AND NOVICK P. The exocyst is an
effector for Sec4p, targeting secretory vesicles to sites of exocytosis. EMBO J 18: 10711080, 1999.
247. GUTIERREZ L, MAGEE AI, MARSHALL CJ, AND HANCOCK JF. Posttranslational processing of p21ras is two-step and involves carboxyl-

January 2001

SMALL G PROTEINS

methylation and carboxy-terminal proteolysis. EMBO J 8: 1093

1098, 1989.
248. HABETS GG, SCHOLTES EH, ZUYDGEEST D, VAN DER KAMMEN RA, STAM
JC, BERNS A, AND COLLARD JG. Identification of an invasion-inducing
gene, Tiam-1, that encodes a protein with homology to GDP-GTP
exchangers for Rho-like proteins. Cell 77: 537549, 1994.
249. HAGAG N, HALEGOUA S, AND VIOLA M. Inhibition of growth factorinduced differentiation of PC12 cells by microinjection of antibody
to ras p21. Nature 319: 680 682, 1986.
250. HALL A. The cellular functions of small GTP-binding proteins. Science 249: 635 640, 1990.
251. HALL A. Rho GTPases and the actin cytoskeleton. Science 279:
509 514, 1998.
251a.HALL A. GTPases. Oxford, UK: Oxford Univ. Press, 2000.
252. HALL A, MARSHALL CJ, SPURR NK, AND WEISS RA. Identification of
transforming gene in two human sarcoma cell lines as a new
member of the ras gene family located on chromosome 1. Nature
303: 396 400, 1983.
253. HAMMONDS-ODIE LP, JACKSON TR, PROFIT AA, BLADER IJ, TURCK CW,
PRESTWICH GD, AND THEIBERT AB. Identification and cloning of centaurin-alpha. A novel phosphatidylinositol 3,4,5-trisphosphate-binding protein from rat brain. J Biol Chem 271: 18859 18868, 1996.
254. HAN L AND COLICELLI J. A human protein selected for interference
with Ras function interacts directly with Ras and competes with
Raf1. Mol Cell Biol 15: 1318 1323, 1995.
254a.HAN M, GOLDEN A, HAN Y, AND STERNBERG PW. C. elegans lin-45 raf
gene participates in let-60 ras-stimulated vulval differentiation. Nature 363: 133140, 1993.
255. HANCOCK JF, CADWALLADER K, AND MARSHALL CJ. Methylation and
proteolysis are essential for efficient membrane binding of prenylated p21K-ras(B). EMBO J 10: 641 646, 1991.
256. HANCOCK JF, MAGEE AI, CHILDS JE, AND MARSHALL CJ. All ras proteins
are polyisoprenylated but only some are palmitoylated. Cell 57:
11671177, 1989.
257. HANCOCK JF, PATERSON H, AND MARSHALL CJ. A polybasic domain or
palmitoylation is required in addition to the CAAX motif to localize
p21ras to the plasma membrane. Cell 63: 133139, 1990.
258. HARDEN N, LEE J, LOH HY, ONG YM, TAN I, LEUNG T, MANSER E, AND
LIM L. A Drosophila homolog of the Rac- and Cdc42-activated
serine/threonine kinase PAK is a potential focal adhesion and focal
complex protein that colocalizes with dynamic actin structures.
Mol Cell Biol 16: 1896 1908, 1996.
259. HART MJ, EVA A, EVANS T, AARONSON SA, AND CERIONE RA. Catalysis
of guanine nucleotide exchange on the CDC42Hs protein by the dbl
oncogene product. Nature 354: 311314, 1991.
260. HART MJ, JIANG X, KOZASA T, ROSCOE W, SINGER WD, GILMAN AG,
STERNWEIS PC, AND BOLLAG G. Direct stimulation of the guanine
nucleotide exchange activity of p115 RhoGEF by G13. Science 280:
21122114, 1998.
261. HART MJ, MARU Y, LEONARD D, WITTE ON, EVANS T, AND CERIONE RA.
A GDP dissociation inhibitor that serves as a GTPase inhibitor for
the Ras-like protein CDC42Hs. Science 258: 812 815, 1992.
262. HART MJ, SHARMA S, ELMASRY N, QIU RG, MCCABE P, POLAKIS P, AND
BOLLAG G. Identification of a novel guanine nucleotide exchange
factor for the Rho GTPase. J Biol Chem 271: 2545225458, 1996.
263. HARTWIG JH, BOKOCH GM, CARPENTER CL, JANMEY PA, TAYLOR LA,
TOKER A, AND STOSSEL TP. Thrombin receptor ligation and activated
Rac uncap actin filament barbed ends through phosphoinositide
synthesis in permeabilized human platelets. Cell 82: 643 653, 1995.
264. HATA Y, NAKANISHI H, AND TAKAI Y. Synaptic PDZ domain-containing
proteins. Neurosci Res 32: 17, 1998.
265. HAUBRUCK H, PRANGE R, VORGIAS C, AND GALLWITZ D. The ras-related
mouse ypt1 protein can functionally replace the YPT1 gene product
in yeast. EMBO J 8: 14271432, 1989.
266. HAWES BE, VAN BIESEN T, KOCH WJ, LUTTRELL LM, AND LEFKOWITZ RJ.
Distinct pathways of Gi- and Gq-mediated mitogen-activated protein kinase activation. J Biol Chem 270: 17148 17153, 1995.
267. HECKENLIVELY JR AND BIRD AC. Choroideremia. In: Retinitis Pigmentosa, edited by Heckenlivelys JR. New York: Lippincott, 1988.
268. HENKEMEYER M, ROSSI DJ, HOLMYARD DP, PURI MC, MBAMALU G,
HARPAL K, SHIH TS, JACKS T, AND PAWSON T. Vascular system defects
and neuronal apoptosis in mice lacking ras GTPase-activating protein. Nature 377: 695701, 1995.

195

269. HENSKE EP, SHORT MP, JOZWIAK S, BOVEY CM, RAMLAKHAN S, HAINES
JL, AND KWIATKOWSKI DJ. Identification of VAV2 on 9q34 and its
exclusion as the tuberous sclerosis gene TSC1. Ann Hum Genet 59:
2537, 1995.
270. HICKE L, YOSHIHISA T, AND SCHEKMAN R. Sec23p and a novel 105-kDa
protein function as a multimeric complex to promote vesicle budding and protein transport from the endoplasmic reticulum. Mol
Biol Cell 3: 667 676, 1992.
271. HILDEBRAND JD, TAYLOR JM, AND PARSONS JT. An SH3 domain-containing GTPase-activating protein for Rho and Cdc42 associates
with focal adhesion kinase. Mol Cell Biol 16: 3169 3178, 1996.
272. HILL CS, WYNNE J, AND TREISMAN R. The Rho family GTPases RhoA,
Rac1, and CDC42Hs regulate transcriptional activation by SRF. Cell
81: 1159 1170, 1995.
273. HIRANO H, TANAKA K, OZAKI K, IMAMURA H, KOHNO H, HIHARA T,
KAMEYAMA T, HOTTA K, ARISAWA M, WATANABE T, QADOTA H, OHYA Y,
AND TAKAI Y. ROM7/BEM4 encodes a novel protein that interacts
with the Rho1p small GTP-binding protein in Saccharomyces cerevisiae. Mol Cell Biol 16: 4396 4403, 1996.
274. HIRAO M, SATO N, KONDO T, YONEMURA S, MONDEN M, SASAKI T, TAKAI
Y, TSUKITA S, AND TSUKITA S. Regulation mechanism of ERM (ezrin/
radixin/moesin) protein/plasma membrane association: possible involvement of phosphatidylinositol turnover and Rho-dependent
signaling pathway. J Cell Biol 135: 3751, 1996.
275. HIRAOKA K, KAIBUCHI K, ANDO S, MUSHA T, TAKAISHI K, MIZUNO T,
ASADA M, MENARD L, TOMHAVE E, DIDSBURY J, SNYDERMAN R, AND
TAKAI Y. Both stimulatory and inhibitory GDP/GTP exchange proteins, smg GDS and rho GDI, are active on multiple small GTPbinding proteins. Biochem Biophys Res Commun 182: 921930,
1992.
276. HIRATA K, KIKUCHI A, SASAKI T, KURODA S, KAIBUCHI K, MATSUURA Y,
SEKI H, SAIDA K, AND TAKAI Y. Involvement of rho p21 in the GTPenhanced calcium ion sensitivity of smooth muscle contraction.
J Biol Chem 267: 8719 8722, 1992.
277. HIROSE M, ISHIZAKI T, WATANABE N, UEHATA M, KRANENBURG O,
MOOLENAAR WH, MATSUMURA F, MAEKAWA M, BITO H, AND NARUMIYA S.
Molecular dissection of the Rho-associated protein kinase
(p160ROCK)-regulated neurite remodeling in neuroblastoma N1E115 cells. J Cell Biol 141: 16251636, 1998.
278. HIRST J AND ROBINSON MS. Clathrin and adaptors. Biochim Biophys
Acta 1404: 173193, 1998.
279. HOFER F, FIELDS S, SCHNEIDER C, AND MARTIN GS. Activated Ras
interacts with the Ral guanine nucleotide dissociation stimulator.
Proc Natl Acad Sci USA 91: 11089 11093, 1994.
280. HOFFMAN GR, NASSAR N, AND CERIONE RA. Structure of the Rho
family GTP-binding protein Cdc42 in complex with the multifunctional regulator RhoGDI. Cell 100: 345356, 2000.
281. HOLT KH, KASSON BG, AND PESSIN JE. Insulin stimulation of a MEKdependent but ERK-independent SOS protein kinase. Mol Cell Biol
16: 577583, 1996.
282. HOLZ RW, BRONDYK WH, SENTER RA, KUIZON L, AND MACARA IG.
Evidence for the involvement of Rab3A in Ca2-dependent exocytosis from adrenal chromaffin cells. J Biol Chem 269: 10229 10234,
1994.
283. HONDA A, NOGAMI M, YOKOZEKI T, YAMAZAKI M, NAKAMURA H, WATANABE H, KAWAMOTO K, NAKAYAMA K, MORRIS AJ, FROHMAN MA, AND
KANAHO Y. Phosphatidylinositol 4-phosphate 5-kinase is a downstream effector of the small G protein ARF6 in membrane ruffle
formation. Cell 99: 521532, 1999.
284. HORDIJK PL, TEN KLOOSTER JP, VAN DER KAMMEN RA, MICHIELS F,
OOMEN LC, AND COLLARD JG. Inhibition of invasion of epithelial cells
by Tiam1-Rac signaling. Science 278: 1464 1466, 1997.
285. HORDIJK PL, VERLAAN I, VAN CORVEN EJ, AND MOOLENAAR WH. Protein
tyrosine phosphorylation induced by lysophosphatidic acid in Rat-1
fibroblasts. Evidence that phosphorylation of map kinase is mediated by the Gi-p21ras pathway. J Biol Chem 269: 645 651, 1994.
286. HORI Y, KIKUCHI A, ISOMURA M, KATAYAMA M, MIURA Y, FUJIOKA H,
KAIBUCHI K, AND TAKAI Y. Posttranslational modifications of the
C-terminal region of the rho protein are important for its interaction with membranes and the stimulatory and inhibitory GDP/GTP
exchange proteins. Oncogene 6: 515522, 1991.
287. HORIUCHI H, GINER A, HOFLACK B, AND ZERIAL M. A GDP/GTP exchange-stimulatory activity for the Rab5-RabGDI complex on clath-

196

288.

289.

290.

291.

292.

293.

294.

295.

296.

297.

298.

299.

300.

301.

302.

303.

304.

305.

306.

TAKAI, SASAKI, AND MATOZAKI

rin-coated vesicles from bovine brain. J Biol Chem 270: 11257
11262, 1995.
HORIUCHI H, KAIBUCHI K, KAWAMURA M, MATSUURA Y, SUZUKI N, KURODA Y, KATAOKA T, AND TAKAI Y. The posttranslational processing of
ras p21 is critical for its stimulation of yeast adenylate cyclase. Mol
Cell Biol 12: 4515 4520, 1992.
HORIUCHI H, KAWATA M, KATAYAMA M, YOSHIDA Y, MUSHA T, ANDO S,
AND TAKAI Y. A novel prenyltransferase for a small GTP-binding
protein having a C-terminal Cys-Ala-Cys structure. J Biol Chem
266: 1698116984, 1991.
HORIUCHI H, LIPPE R, MCBRIDE HM, RUBINO M, WOODMAN P, STENMARK
H, RYBIN V, WILM M, ASHMAN K, MANN M, AND ZERIAL M. A novel Rab5
GDP/GTP exchange factor complexed to Rabaptin-5 links nucleotide exchange to effector recruitment and function. Cell 90: 1149
1159, 1997.
HOSHIJIMA M, KONDO J, KIKUCHI A, YAMAMOTO K, AND TAKAI Y. Purification and characterization from bovine brain membranes of a
GTP-binding protein with a Mr of 21,000, ADP-ribosylated by an
ADP-ribosyltransferase contaminated in botulinum toxin type C1identification as the rhoA gene product. Brain Res Mol Brain Res
7: 9 16, 1990.
HOTTA K, TANAKA K, MINO A, KOHNO H, AND TAKAI Y. Interaction of
the Rho family small G proteins with kinectin, an anchoring protein
of kinesin motor. Biochem Biophys Res Commun 225: 69 74, 1996.
HOUSSA B, DE WIDT J, KRANENBURG O, MOOLENAAR WH, AND VAN
BLITTERSWIJK WJ. Diacylglycerol kinase theta binds to and is negatively regulated by active RhoA. J Biol Chem 274: 6820 6822, 1999.
HOWE LR AND MARSHALL CJ. Lysophosphatidic acid stimulates mitogen-activated protein kinase activation via a G-protein-coupled
pathway requiring p21ras and p74raf-1. J Biol Chem 268: 20717
20720, 1993.
HSU SC, HAZUKA CD, FOLETTI DL, AND SCHELLER RH. Targeting vesicles to specific sites on the plasma membrane: the role of the sec6/8
complex. Trends Cell Biol 9: 150 153, 1999.
HU CD, KARIYA K, KOTANI G, SHIROUZU M, YOKOYAMA S, AND KATAOKA
T. Coassociation of Rap1A and Ha-Ras with Raf-1 N-terminal region
interferes with ras-dependent activation of Raf-1. J Biol Chem 272:
1170211705, 1997.
HU CD, KARIYA K, TAMADA M, AKASAKA K, SHIROUZU M, YOKOYAMA S,
AND KATAOKA T. Cysteine-rich region of Raf-1 interacts with activator domain of posttranslationally modified Ha-Ras. J Biol Chem
270: 30274 30277, 1995.
HU KQ AND SETTLEMAN J. Tandem SH2 binding sites mediate the
RasGAP-RhoGAP interaction: a conformational mechanism for
SH3 domain regulation. EMBO J 16: 473 483, 1997.
HUANG W, ALESSANDRINI A, CREWS CM, AND ERIKSON RL. Raf-1 forms
a stable complex with Mek1 and activates Mek1 by serine phosphorylation. Proc Natl Acad Sci USA 90: 1094710951, 1993.
HUANG Y, SAEZ R, CHAO L, SANTOS E, AARONSON SA, AND CHAN AM. A
novel insertional mutation in the TC21 gene activates its transforming activity in a human leiomyosarcoma cell line. Oncogene 11:
12551260, 1995.
ICHIBA T, KURAISHI Y, SAKAI O, NAGATA S, GROFFEN J, KURATA T,
HATTORI S, AND MATSUDA M. Enhancement of guanine-nucleotide
exchange activity of C3G for Rap1 by the expression of Crk, CrkL,
and Grb2. J Biol Chem 272: 2221522220, 1997.
IKEDA M, ISHIDA O, HINOI T, KISHIDA S, AND KIKUCHI A. Identification
and characterization of a novel protein interacting with Ral-binding
protein 1, a putative effector protein of Ral. J Biol Chem 273:
814 821, 1998.
IMAMURA H, TAKAISHI K, NAKANO K, KODAMA A, OISHI H, SHIOZAKI H,
MONDEN M, SASAKI T, AND TAKAI Y. Rho and Rab small G proteins
coordinately reorganize stress fibers and focal adhesions in MDCK
cells. Mol Biol Cell 9: 25612575, 1998.
IMAMURA H, TANAKA K, HIHARA T, UMIKAWA M, KAMEI T, TAKAHASHI K,
SASAKI T, AND TAKAI Y. Bni1p and Bnr1p: downstream targets of the
Rho family small G-proteins which interact with profilin and regulate actin cytoskeleton in Saccharomyces cerevisiae. EMBO J 16:
27452755, 1997.
INOUE SB, QADOTA H, ARISAWA M, WATANABE T, AND OHYA Y. Prenylation of Rho1p is required for activation of yeast 1,3--glucan
synthase. J Biol Chem 274: 38119 38124, 1999.
ISHIZAKI H, MIYOSHI J, TOGAWA A, TANAKA M, SASAKI T, KAMIYA H,

307.

308.

309.

310.

311.

312.

313.

314.

315.

316.

317.
318.

319.

320.

321.

322.

323.

324.

325.

326.

Volume 81

OZAWA S, ENDO K, MIZOGUCHI A, AND TAKAI Y. Role of Rab GDP

dissociation inhibitor in regulating plasticity of hippocampal
neurotransmission. Proc Natl Acad Sci USA 97: 1158711592, 2000.
ISHIZAKI T, MAEKAWA M, FUJISAWA K, OKAWA K, IWAMATSU A, FUJITA A,
WATANABE N, SAITO Y, KAKIZUKA A, MORII N, AND NARUMIYA S. The
small GTP-binding protein Rho binds to and activates a 160 kDa
Ser/Thr protein kinase homologous to myotonic dystrophy kinase.
EMBO J 15: 18851893, 1996.
ISOMURA M, KIKUCHI A, OHGA N, AND TAKAI Y. Regulation of binding
of rhoB p20 to membranes by its specific regulatory protein, GDP
dissociation inhibitor. Oncogene 6: 119 124, 1991.
ITOH T, KAIBUCHI K, MASUDA T, YAMAMOTO T, MATSUURA Y, MAEDA A,
SHIMIZU K, AND TAKAI Y. The posttranslational processing of ras p21
is critical for its stimulation of mitogen-activated protein kinase.
J Biol Chem 268: 30253028, 1993.
IWASAKI K, STAUNTON J, SAIFEE O, NONET M, AND THOMAS JH. aex-3
encodes a novel regulator of presynaptic activity in C. elegans.
Neuron 18: 613 622, 1997.
IZAURRALDE E, KUTAY U, VON KOBBE C, MATTAJ IW, AND GORLICH D.
The asymmetric distribution of the constituents of the Ran system
is essential for transport into and out of the nucleus. EMBO J 16:
6535 6547, 1997.
JACKSON CL AND CASANOVA JE. Turning on ARF: the Sec7 family of
guanine-nucleotide-exchange factors. Trends Cell Biol 10: 60 67,
2000.
JACKSON JH, COCHRANE CG, BOURNE JR, SOLSKI PA, BUSS JE, AND DER
CJ. Farnesol modification of Kirsten-ras exon 4B protein is essential for transformation. Proc Natl Acad Sci USA 87: 30423046,
1990.
JALINK K, VAN CORVEN EJ, HENGEVELD T, MORII N, NARUMIYA S, AND
MOOLENAAR WH. Inhibition of lysophosphatidate- and thrombininduced neurite retraction and neuronal cell rounding by ADP
ribosylation of the small GTP-binding protein Rho. J Cell Biol 126:
801 810, 1994.
JANOUEIX-LEROSEY I, JOLLIVET F, CAMONIS J, MARCHE PN, AND GOUD B.
Two-hybrid system screen with the small GTP-binding protein
Rab6. Identification of a novel mouse GDP dissociation inhibitor
isoform and two other potential partners of Rab6. J Biol Chem 270:
1480114808, 1995.
JIANG H, LUO JQ, URANO T, FRANKEL P, LU Z, FOSTER DA, AND FEIG LA.
Involvement of Ral GTPase in v-Src-induced phospholipase D activation. Nature 378: 409 412, 1995.
JIN Z AND STRITTMATTER SM. Rac1 mediates collapsin-1-induced
growth cone collapse. J Neurosci 17: 6256 6263, 1997.
JOBERTY G, PERLUNGHER RR, AND MACARA IG. The Borgs, a new
family of Cdc42 and TC10 GTPase-interacting proteins. Mol Cell
Biol 19: 6585 6597, 1999.
JOHANNES L, LLEDO PM, ROA M, VINCENT JD, HENRY JP, AND DARCHEN
F. The GTPase Rab3a negatively controls calcium-dependent exocytosis in neuroendocrine cells. EMBO J 13: 2029 2037, 1994.
JOHNSON DI AND PRINGLE JR. Molecular characterization of CDC42,
a Saccharomyces cerevisiae gene involved in the development of
cell polarity. J Cell Biol 111: 143152, 1990.
JOHNSON L, GREENBAUM D, CICHOWSKI K, MERCER K, MURPHY E,
SCHMITT E, BRONSON RT, UMANOFF H, EDELMANN W, KUCHERLAPATI R,
AND JACKS T. K-ras is an essential gene in the mouse with partial
functional overlap with N-ras. Genes Dev 11: 2468 2481, 1997.
JOHNSTON PA, ARCHER B, ROBINSON K, MIGNERY GA, JAHN R, AND
SUDHOF TC. rab3A attachment to the synaptic vesicle membrane
mediated by a conserved polyisoprenylated carboxy-terminal sequence. Neuron 7: 101109, 1991.
JONESON T, MCDONOUGH M, BAR-SAGI D, AND VAN AELST L. RAC
regulation of actin polymerization and proliferation by a pathway
distinct from Jun kinase. Science 274: 1374 1376, 1996.
JOU TS AND NELSON WJ. Effects of regulated expression of mutant
RhoA and Rac1 small GTPases on the development of epithelial
(MDCK) cell polarity. J Cell Biol 142: 85100, 1998.
KAHANA JA AND CLEVELAND DW. Beyond nuclear transport. Ran-GTP
as a determinant of spindle assembly. J Cell Biol 146: 12051210,
1999.
KAHN RA AND GILMAN AG. Purification of a protein cofactor required
for ADP-ribosylation of the stimulatory regulatory component of

January 2001

SMALL G PROTEINS

adenylate cyclase by cholera toxin. J Biol Chem 259: 6228 6234,

1984.
327. KAHN RA AND GILMAN AG. The protein cofactor necessary for ADPribosylation of Gs by cholera toxin is itself a GTP binding protein.
J Biol Chem 261: 7906 7911, 1986.
328. KAIBUCHI K, MIZUNO T, FUJIOKA H, YAMAMOTO T, KISHI K, FUKUMOTO Y,
HORI Y, AND TAKAI Y. Molecular cloning of the cDNA for stimulatory
GDP/GTP exchange protein for smg p21s (ras p21-like small GTPbinding proteins) and characterization of stimulatory GDP/GTP
exchange protein. Mol Cell Biol 11: 28732880, 1991.
329. KALAB P, PU RT, AND DASSO M. The ran GTPase regulates mitotic
spindle assembly. Curr Biol 9: 481 484, 1999.
330. KAMADA Y, QADOTA H, PYTHON CP, ANRAKU Y, OHYA Y, AND LEVIN DE.
Activation of yeast protein kinase C by Rho1 GTPase. J Biol Chem
271: 91939196, 1996.
331. KAMEI T, MATOZAKI T, SAKISAKA T, KODAMA A, YOKOYAMA S, PENG YF,
NAKANO K, TAKAISHI K, AND TAKAI Y. Coendocytosis of cadherin and
c-Met coupled to disruption of cell-cell adhesion in MDCK cells:
regulation by Rho, Rac and Rab small G proteins. Oncogene 18:
6776 6784, 1999.
331a.KAMEI T, TANAKA K, HIHARA T, UMIKAWA M, IMAMAURA H, KIKYO M,
OZAKI K, AND TAKAI Y. Interaction of Bnr1p with a novel Src homology 3 domain-containing Hof1p. Implication in cytokinesis in Saccharomyces cerevisiae. J Biol Chem 273: 2834128345, 1998.
332. KAPLAN DR, MORRISON DK, WONG G, MCCORMICK F, AND WILLIAMS LT.
PDGF -receptor stimulates tyrosine phosphorylation of GAP and
association of GAP with a signaling complex. Cell 61: 125133,
1990.
333. KATO M, SASAKI T, OHYA T, NAKANISHI H, NISHIOKA H, IMAMURA M, AND
TAKAI Y. Physical and functional interaction of rabphilin-3A with
-actinin. J Biol Chem 271: 3177531778, 1996.
334. KAUFFMANN-ZEH A, RODRIGUEZ-VICIANA P, ULRICH E, GILBERT C, COFFER P, DOWNWARD J, AND EVAN G. Suppression of c-Myc-induced
apoptosis by Ras signalling through PI(3)K and PKB. Nature 385:
544 548, 1997.
335. KAWASAKI H, SPRINGETT GM, MOCHIZUKI N, TOKI S, NAKAYA M, MATSUDA M, HOUSMAN DE, AND GRAYBIEL AM. A family of cAMP-binding
proteins that directly activate Rap1. Science 282: 22752279, 1998.
336. KAWATA M, FARNSWORTH CC, YOSHIDA Y, GELB MH, GLOMSET JA, AND
TAKAI Y. Posttranslationally processed structure of the human
platelet protein smg p21B: evidence for geranylgeranylation and
carboxyl methylation of the C-terminal cysteine. Proc Natl Acad
Sci USA 87: 8960 8964, 1990.
337. KAWATA M, MATSUI Y, KONDO J, HISHIDA T, TERANISHI Y, AND TAKAI Y.
A novel small molecular weight GTP-binding protein with the same
putative effector domain as the ras proteins in bovine brain membranes. Purification, determination of primary structure, and characterization. J Biol Chem 263: 1896518971, 1988.
338. KAZIRO Y. The role of guanosine 5-triphosphate in polypeptide
chain elongation. Biochim Biophys Acta 505: 95127, 1978.
339. KAZLAUSKAS A, ELLIS C, PAWSON T, AND COOPER JA. Binding of GAP to
activated PDGF receptors. Science 247: 1578 1581, 1990.
340. KHOSRAVI-FAR R, LUTZ RJ, COX AD, CONROY L, BOURNE JR, SINENSKY
M, BALCH WE, BUSS JE, AND DER CJ. Isoprenoid modification of rab
proteins terminating in CC or CXC motifs. Proc Natl Acad Sci USA
88: 6264 6268, 1991.
341. KHOSRAVI-FAR R, SOLSKI PA, CLARK GJ, KINCH MS, AND DER CJ.
Activation of Rac1, RhoA, and mitogen-activated protein kinases is
required for Ras transformation. Mol Cell Biol 15: 6443 6453, 1995.
342. KIKUCHI A, DEMO SD, YE ZH, CHEN YW, AND WILLIAMS LT. ralGDS
family members interact with the effector loop of ras p21. Mol Cell
Biol 14: 74837491, 1994.
343. KIKUCHI A, YAMAMOTO K, FUJITA T, AND TAKAI Y. ADP-ribosylation of
the bovine brain rho protein by botulinum toxin type C1. J Biol
Chem 263: 1630316308, 1988.
344. KIKUCHI A, YAMASHITA T, KAWATA M, YAMAMOTO K, IKEDA K, TANIMOTO
T, AND TAKAI Y. Purification and characterization of a novel GTPbinding protein with a molecular weight of 24,000 from bovine
brain membranes. J Biol Chem 263: 28972904, 1988.
345. KIM E, AMBROZIAK P, OTTO JC, TAYLOR B, ASHBY M, SHANNON K, CASEY
PJ, AND YOUNG SG. Disruption of the mouse Rce1 gene results in
defective Ras processing and mislocalization of Ras within cells.
J Biol Chem 274: 8383 8390, 1999.

197

346. KIM JH, LIAO D, LAU LF, AND HUGANIR RL. SynGAP: a synaptic
RasGAP that associates with the PSD-95/SAP90 protein family.
Neuron 20: 683 691, 1998.
347. KIMURA K, ITO M, AMANO M, CHIHARA K, FUKATA Y, NAKAFUKU M,
YAMAMORI B, FENG J, NAKANO T, OKAWA K, IWAMATSU A, AND KAIBUCHI
K. Regulation of myosin phosphatase by Rho and Rho-associated
kinase (Rho-kinase). Science 273: 245248, 1996.
348. KING FJ, HU E, HARRIS DF, SARRAF P, SPIEGELMAN BM, AND ROBERTS
TM. DEF-1, a novel Src SH3 binding protein that promotes adipogenesis in fibroblastic cell lines. Mol Cell Biol 19: 2330 2337, 1999.
349. KINSELLA BT AND MALTESE WA. rab GTP-binding proteins with three
different carboxyl-terminal cysteine motifs are modified in vivo by
20-carbon isoprenoids. J Biol Chem 267: 3940 3945, 1992.
350. KIRCHHAUSEN T. Adaptors for clathrin-mediated traffic. Annu Rev
Cell Dev Biol 15: 705732, 1999.
351. KIRKPATRICK D AND SOLOMON F. Overexpression of yeast homologs of
the mammalian checkpoint gene RCC1 suppresses the class of
-tubulin mutations that arrest with excess microtubules. Genetics
137: 381392, 1994.
352. KISHI K, SASAKI T, KURODA S, ITOH T, AND TAKAI Y. Regulation of
cytoplasmic division of Xenopus embryo by rho p21 and its inhibitory GDP/GTP exchange protein (rho GDI). J Cell Biol 120: 1187
1195, 1993.
353. KITAYAMA H, SUGIMOTO Y, MATSUZAKI T, IKAWA Y, AND NODA M. A
ras-related gene with transformation suppressor activity. Cell 56:
77 84, 1989.
354. KITAZAWA T, MASUO M, AND SOMLYO AP. G protein-mediated inhibition of myosin light-chain phosphatase in vascular smooth muscle.
Proc Natl Acad Sci USA 88: 93079310, 1991.
355. KIYOKAWA E, MOCHIZUKI N, KURATA T, AND MATSUDA M. Role of Crk
oncogene product in physiologic signaling. Crit Rev Oncogene 8:
329 342, 1997.
356. KLARLUND JK, CHERNIACK AD, MCMAHON M, AND CZECH MP. Role of
the Raf/mitogen-activated protein kinase pathway in p21ras desensitization. J Biol Chem 271: 16674 16677, 1996.
357. KLARLUND JK, GUILHERME A, HOLIK JJ, VIRBASIUS JV, CHAWLA A, AND
CZECH MP. Signaling by phosphoinositide-3,4,5-trisphosphate
through proteins containing pleckstrin and Sec7 homology domains. Science 275: 19271930, 1997.
358. KNAUS UG, HEYWORTH PG, EVANS T, CURNUTTE JT, AND BOKOCH GM.
Regulation of phagocyte oxygen radical production by the GTPbinding protein Rac 2. Science 254: 15121515, 1991.
359. KOBAYASHI K, KURODA S, FUKATA M, NAKAMURA T, NAGASE T, NOMURA
N, MATSUURA Y, YOSHIDA-KUBOMURA N, IWAMATSU A, AND KAIBUCHI K.
p140Sra-1 (specifically Rac1-associated protein) is a novel specific
target for Rac1 small GTPase. J Biol Chem 273: 291295, 1998.
360. KODAMA A, MATOZAKI T, FUKUHARA A, KIKYO M, ICHIHASHI M, AND TAKAI
Y. Involvement of an SHP-2-Rho small G protein pathway in hepatocyte growth factor/scatter factor-induced cell scattering. Mol Biol
Cell 11: 25652575, 2000.
361. KODAMA A, TAKAISHI K, NAKANO K, NISHIOKA H, AND TAKAI Y. Involvement of Cdc42 small G protein in cell-cell adhesion, migration and
morphology of MDCK cells. Oncogene 18: 3996 4006, 1999.
362. KOERA K, NAKAMURA K, NAKAO K, MIYOSHI J, TOYOSHIMA K, HATTA T,
OTANI H, AIBA A, AND KATSUKI M. K-ras is essential for the development of the mouse embryo. Oncogene 15: 11511159, 1997.
363. KOHNO H, TANAKA K, MINO A, UMIKAWA M, IMAMURA H, FUJIWARA T,
FUJITA Y, HOTTA K, QADOTA H, WATANABE T, OHYA Y, AND TAKAI Y.
Bni1p implicated in cytoskeletal control is a putative target of
Rho1p small GTP binding protein in Saccharomyces cerevisiae.
EMBO J 15: 6060 6068, 1996.
364. KOMURO R, SASAKI T, ORITA S, MAEDA M, AND TAKAI Y. Involvement of
rabphilin-3A in Ca2-dependent exocytosis from PC12 cells. Biochem Biophys Res Commun 219: 435 440, 1996.
365. KOMURO R, SASAKI T, TAKAISHI K, ORITA S, AND TAKAI Y. Involvement
of Rho and Rac small G proteins and Rho GDI in Ca2-dependent
exocytosis from PC12 cells. Genes Cells 1: 943951, 1996.
366. KORNFELD K, HOM DB, AND HORVITZ HR. The ksr-1 gene encodes a
novel protein kinase involved in Ras-mediated signaling in C. elegans. Cell 83: 903913, 1995.
367. KOZASA T, JIANG X, HART MJ, STERNWEIS PM, SINGER WD, GILMAN AG,
BOLLAG G, AND STERNWEIS PC. p115 RhoGEF, a GTPase activating
protein for G12 and G13. Science 280: 2109 2111, 1998.

198

TAKAI, SASAKI, AND MATOZAKI

368. KOZMA R, AHMED S, BEST A, AND LIM L. The Ras-related protein

Cdc42Hs and bradykinin promote formation of peripheral actin
microspikes and filopodia in Swiss 3T3 fibroblasts. Mol Cell Biol 15:
19421952, 1995.
369. KOZMA R, SARNER S, AHMED S, AND LIM L. Rho family GTPases and
neuronal growth cone remodelling: relationship between increased
complexity induced by Cdc42Hs, Rac1, and acetylcholine and collapse induced by RhoA and lysophosphatidic acid. Mol Cell Biol 17:
12011211, 1997.
370. KTISTAKIS NT, BROWN HA, WATERS MG, STERNWEIS PC, AND ROTH MG.
Evidence that phospholipase D mediates ADP ribosylation factordependent formation of Golgi coated vesicles. J Cell Biol 134:
295306, 1996.
371. KUEHN MJ AND SCHEKMAN R. COPII and secretory cargo capture into
transport vesicles. Curr Opin Cell Biol 9: 477 483, 1997.
372. KUGE O, DASCHER C, ORCI L, ROWE T, AMHERDT M, PLUTNER H,
RAVAZZOLA M, TANIGAWA G, ROTHMAN JE, AND BALCH WE. Sar1 promotes vesicle budding from the endoplasmic reticulum but not
Golgi compartments. J Cell Biol 125: 51 65, 1994.
373. KURIYAMA M, HARADA N, KURODA S, YAMAMOTO T, NAKAFUKU M,
IWAMATSU A, YAMAMOTO D, PRASAD R, CROCE C, CANAANI E, AND
KAIBUCHI K. Identification of AF-6 and canoe as putative targets for
Ras. J Biol Chem 271: 607 610, 1996.
374. KURODA S, FUKATA M, KOBAYASHI K, NAKAFUKU M, NOMURA N,
IWAMATSU A, AND KAIBUCHI K. Identification of IQGAP as a putative
target for the small GTPases, Cdc42 and Rac1. J Biol Chem 271:
2336323367, 1996.
375. KURODA S, FUKATA M, NAKAGAWA M, FUJII K, NAKAMURA T, OOKUBO T,
IZAWA I, NAGASE T, NOMURA N, TANI H, SHOJI I, MATSUURA Y, YONEHARA
S, AND KAIBUCHI K. Role of IQGAP1, a target of the small GTPases
Cdc42 and Rac1, in regulation of E-cadherin-mediated cell-cell
adhesion. Science 281: 832 835, 1998.
376. KURODA Y, SUZUKI N, AND KATAOKA T. The effect of posttranslational
modifications on the interaction of Ras2 with adenylyl cyclase.
Science 259: 683 686, 1993.
377. KUTAY U, BISCHOFF FR, KOSTKA S, KRAFT R, AND GORLICH D. Export of
importin from the nucleus is mediated by a specific nuclear
transport factor. Cell 90: 10611071, 1997.
378. KYRIAKIS JM, APP H, ZHANG XF, BANERJEE P, BRAUTIGAN DL, RAPP UR,
AND AVRUCH J. Raf-1 activates MAP kinase-kinase. Nature 358:
417 421, 1992.
379. LAMARCHE N, TAPON N, STOWERS L, BURBELO PD, ASPENSTROM P,
BRIDGES T, CHANT J, AND HALL A. Rac and Cdc42 induce actin
polymerization and G1 cell cycle progression independently of
p65PAK and the JNK/SAPK MAP kinase cascade. Cell 87: 519 529,
1996.
380. LAMAZE C, CHUANG TH, TERLECKY LJ, BOKOCH GM, AND SCHMID SL.
Regulation of receptor-mediated endocytosis by Rho and Rac. Nature 382: 177179, 1996.
381. LANCASTER CA, TAYLOR-HARRIS PM, SELF AJ, BRILL S, VAN ERP HE, AND
HALL A. Characterization of rhoGAP. A GTPase-activating protein
for rho-related small GTPases. J Biol Chem 269: 11371142, 1994.
382. LANGILLE SE, PATKI V, KLARLUND JK, BUXTON JM, HOLIK JJ, CHAWLA A,
CORVERA S, AND CZECH MP. ADP-ribosylation factor 6 as a target of
guanine nucleotide exchange factor GRP1. J Biol Chem 274:
27099 27104, 1999.
383. LAZAR T, GOTTE M, AND GALLWITZ D. Vesicular transport: how many
Ypt/Rab-GTPases make a eukaryotic cell? Trends Biochem Sci 22:
468 472, 1997.
384. LEBERER E, DIGNARD D, HARCUS D, THOMAS DY, AND WHITEWAY M. The
protein kinase homologue Ste20p is required to link the yeast
pheromone response G-protein subunits to downstream signalling components. EMBO J 11: 4815 4824, 1992.
385. LEE C, DELLA NG, CHEW CE, AND ZACK DJ. Rin, a neuron-specific and
calmodulin-binding small G-protein, and Rit define a novel subfamily of ras proteins. J Neurosci 16: 6784 6794, 1996.
386. LEE L, KLEE SK, EVANGELISTA M, BOONE C, AND PELLMAN D. Control of
mitotic spindle position by the Saccharomyces cerevisiae formin
Bni1p. J Cell Biol 144: 947961, 1999.
387. LEEUW T, FOUREST-LIEUVIN A, WU C, CHENEVERT J, CLARK K, WHITEWAY
M, THOMAS DY, AND LEBERER E. Pheromone response in yeast:
association of Bem1p with proteins of the MAP kinase cascade and
actin. Science 270: 1210 1213, 1995.

Volume 81

388. LEEUWEN FN, KAIN HE, KAMMEN RA, MICHIELS F, KRANENBURG OW,
AND COLLARD JG. The guanine nucleotide exchange factor Tiam1
affects neuronal morphology: opposing roles for the small GTPases
Rac and Rho. J Cell Biol 139: 797 807, 1997.
389. LEEVERS SJ, PATERSON HF, AND MARSHALL CJ. Requirement for Ras in
Raf activation is overcome by targeting Raf to the plasma membrane. Nature 369: 411 414, 1994.
390. LELIAS JM, ADRA CN, WULF GM, GUILLEMOT JC, KHAGAD M, CAPUT D,
AND LIM B. cDNA cloning of a human mRNA preferentially expressed in hematopoietic cells and with homology to a GDP-dissociation inhibitor for the rho GTP-binding proteins. Proc Natl Acad
Sci USA 90: 1479 1483, 1993.
391. LEONARD D, HART MJ, PLATKO JV, EVA A, HENZEL W, EVANS T, AND
CERIONE RA. The identification and characterization of a GDPdissociation inhibitor (GDI) for the CDC42Hs protein. J Biol Chem
267: 22860 22868, 1992.
392. LERNER EC, QIAN Y, BLASKOVICH MA, FOSSUM RD, VOGT A, SUN J, COX
AD, DER CJ, HAMILTON AD, AND SEBTI SM. Ras CAAX peptidomimetic FTI-277 selectively blocks oncogenic Ras signaling by inducing cytoplasmic accumulation of inactive Ras-Raf complexes.
J Biol Chem 270: 2680226806, 1995.
393. LEUNG T, CHEN XQ, MANSER E, AND LIM L. The p160 RhoA-binding
kinase ROK is a member of a kinase family and is involved in the
reorganization of the cytoskeleton. Mol Cell Biol 16: 53135327,
1996.
394. LEUNG T, CHEN XQ, TAN I, MANSER E, AND LIM L. Myotonic dystrophy
kinase-related Cdc42-binding kinase acts as a Cdc42 effector in
promoting cytoskeletal reorganization. Mol Cell Biol 18: 130 140,
1998.
395. LEUNG T, MANSER E, TAN L, AND LIM L. A novel serine/threonine
kinase binding the Ras-related RhoA GTPase which translocates
the kinase to peripheral membranes. J Biol Chem 270: 29051
29054, 1995.
396. LEVIN DE AND ERREDE B. The proliferation of MAP kinase signaling
pathways in yeast. Curr Opin Cell Biol 7: 197202, 1995.
397. LI C, TAKEI K, GEPPERT M, DANIELL L, STENIUS K, CHAPMAN ER, JAHN
R, DE CAMILLI P, AND SUDHOF TC. Synaptic targeting of rabphilin-3A,
a synaptic vesicle Ca2/phospholipid-binding protein, depends on
rab3A/3C. Neuron 13: 885 898, 1994.
398. LI G, DSOUZA-SCHOREY C, BARBIERI MA, COOPER JA, AND STAHL PD.
Uncoupling of membrane ruffling and pinocytosis during Ras signal
transduction. J Biol Chem 272: 1033710340, 1997.
399. LI N, BATZER A, DALY R, YAJNIK V, SKOLNIK E, CHARDIN P, BAR-SAGI D,
MARGOLIS B, AND SCHLESSINGER J. Guanine-nucleotide-releasing factor hSos1 binds to Grb2 and links receptor tyrosine kinases to Ras
signalling. Nature 363: 85 88, 1993.
400. LI S, WANG Q, CHAKLADAR A, BRONSON RT, AND BERNARDS A. Gastric
hyperplasia in mice lacking the putative Cdc42 effector IQGAP1.
Mol Cell Biol 20: 697701, 2000.
401. LIAN JP, STONE S, JIANG Y, LYONS P, AND FERRO-NOVICK S. Ypt1p
implicated in v-SNARE activation. Nature 372: 698 701, 1994.
402. LIAO Y, KARIYA K, HU CD, SHIBATOHGE M, GOSHIMA M, OKADA T,
WATARI Y, GAO X, JIN TG, YAMAWAKI-KATAOKA Y, AND KATAOKA T.
RA-GEF, a novel Rap1A guanine nucleotide exchange factor containing a Ras/Rap1A-associating domain, is conserved between
nematode and humans. J Biol Chem 274: 3781537820, 1999.
403. LIEBL EC, FORSTHOEFEL DJ, FRANCO LS, SAMPLE SH, HESS JE, COWGER
JA, CHANDLER MP, SHUPERT AM, AND SEEGER MA. Dosage-sensitive,
reciprocal genetic interactions between the Abl tyrosine kinase and
the putative GEF trio reveal trios role in axon pathfinding. Neuron
26: 107118, 2000.
404. LINNEMANN T, GEYER M, JAITNER BK, BLOCK C, KALBITZER HR, WITTINGHOFER A, AND HERRMANN C. Thermodynamic and kinetic characterization of the interaction between the Ras binding domain of
AF6 and members of the Ras subfamily. J Biol Chem 274: 13556
13562, 1999.
405. LIPPINCOTT-SCHWARTZ J, COLE NB, AND DONALDSON JG. Building a
secretory apparatus: role of ARF1/COPI in Golgi biogenesis and
maintenance. Histochem Cell Biol 109: 449 462, 1998.
406. LIU L, DUDLER T, AND GELB MH. Purification of a protein palmitoyltransferase that acts on H-Ras protein and on a C-terminal N-Ras
peptide. J Biol Chem 271: 23269 23276, 1996.
407. LLEDO PM, VERNIER P, VINCENT JD, MASON WT, AND ZOREC R. Inhibi-

January 2001

408.
409.

410.
411.
412.

413.

415.

416.

417.
418.

419.

420.
421.
422.

423.

424.

425.

426.

427.

428.
429.

430.

431.

432.

SMALL G PROTEINS

tion of Rab3B expression attenuates Ca2-dependent exocytosis in

rat anterior pituitary cells. Nature 364: 540 544, 1993.
LONART G, JANZ R, JOHNSON KM, AND SUDHOF TC. Mechanism of
action of rab3A in mossy fiber LTP. Neuron 21: 11411150, 1998.
LOUNSBURY KM AND MACARA IG. Ran-binding protein 1 (RanBP1)
forms a ternary complex with Ran and karyopherin and reduces
Ran GTPase-activating protein (RanGAP) inhibition by karyopherin . J Biol Chem 272: 551555, 1997.
LOWE M AND KREIS TE. Regulation of membrane traffic in animal
cells by COPI. Biochim Biophys Acta 1404: 53 66, 1998.
LUO L, JAN LY, AND JAN YN. Rho family GTP-binding proteins in
growth cone signalling. Curr Opin Neurobiol 7: 81 86, 1997.
LUO L, LIAO YJ, JAN LY, AND JAN YN. Distinct morphogenetic functions of similar small GTPases: Drosophila Drac1 is involved in
axonal outgrowth and myoblast fusion. Genes Dev 8: 17871802,
1994.
LUTCKE A, JANSSON S, PARTON RG, CHAVRIER P, VALENCIA A, HUBER LA,
LEHTONEN E, AND ZERIAL M. Rab17, a novel small GTPase, is specific
for epithelial cells and is induced during cell polarization. J Cell
Biol 121: 553564, 1993.
MA L, ROHATGI R, AND KIRSCHNER MW. The Arp2/3 complex mediates
actin polymerization induced by the small GTP-binding protein
Cdc42. Proc Natl Acad Sci USA 95: 1536215367, 1998.
MACDONALD SG, CREWS CM, WU L, DRILLER J, CLARK R, ERIKSON RL,
AND MCCORMICK F. Reconstitution of the Raf-1-MEK-ERK signal
transduction pathway in vitro. Mol Cell Biol 13: 6615 6620, 1993.
MACHESKY LM AND INSALL RH. Signaling to actin dynamics. J Cell
Biol 146: 267272, 1999.
MACHESKY LM, MULLINS RD, HIGGS HN, KAISER DA, BLANCHOIN L, MAY
RC, HALL ME, AND POLLARD TD. Scar, a WASp-related protein,
activates nucleation of actin filaments by the Arp2/3 complex. Proc
Natl Acad Sci USA 96: 3739 3744, 1999.
MACK D, NISHIMURA K, DENNEHEY BK, ARBOGAST T, PARKINSON J, TOH-E
A, PRINGLE JR, BENDER A, AND MATSUI Y. Identification of the bud
emergence gene BEM4 and its interactions with rho-type GTPases
in Saccharomyces cerevisiae. Mol Cell Biol 16: 4387 4395, 1996.
MACKAY DJ AND HALL A. Rho GTPases. J Biol Chem 273: 20685
20688, 1998.
MADAULE P AND AXEL R. A novel ras-related gene family. Cell 41:
31 40, 1985.
MADAULE P, AXEL R, AND MYERS AM. Characterization of two members of the rho gene family from the yeast Saccharomyces cerevisiae. Proc Natl Acad Sci USA 84: 779 783, 1987.
MADAULE P, EDA M, WATANABE N, FUJISAWA K, MATSUOKA T, BITO H,
ISHIZAKI T, AND NARUMIYA S. Role of citron kinase as a target of the
small GTPase Rho in cytokinesis. Nature 394: 491 494, 1998.
MADAULE P, FURUYASHIKI T, REID T, ISHIZAKI T, WATANABE G, MORII N,
AND NARUMIYA S. A novel partner for the GTP-bound forms of rho
and rac. FEBS Lett 377: 243248, 1995.
MAEKAWA M, ISHIZAKI T, BOKU S, WATANABE N, FUJITA A, IWAMATSU A,
OBINATA T, OHASHI K, MIZUNO K, AND NARUMIYA S. Signaling from Rho
to the actin cytoskeleton through protein kinases ROCK and LIMkinase. Science 285: 895 898, 1999.
MAEKAWA M, LI S, IWAMATSU A, MORISHITA T, YOKOTA K, IMAI Y,
KOHSAKA S, NAKAMURA S, AND HATTORI S. A novel mammalian Ras
GTPase-activating protein which has phospholipid-binding and Btk
homology regions. Mol Cell Biol 14: 6879 6885, 1994.
MAGEE AI, NEWMAN CM, GIANNAKOUROS T, HANCOCK JF, FAWELL E,
AND ARMSTRONG J. Lipid modifications and function of the ras superfamily of proteins. Biochem Soc Trans 20: 497 499, 1992.
MAGEE T AND MARSHALL C. New insights into the interaction of Ras
with the plasma membrane. Cell 98: 9 12, 1999.
MAGUIRE J, SANTORO T, JENSEN P, SIEBENLIST U, YEWDELL J, AND KELLY
K. Gem: an induced, immediate early protein belonging to the Ras
family. Science 265: 241244, 1994.
MAHAJAN R, DELPHIN C, GUAN T, GERACE L, AND MELCHIOR F. A small
ubiquitin-related polypeptide involved in targeting RanGAP1 to
nuclear pore complex protein RanBP2. Cell 88: 97107, 1997.
MALCOLM KC, ROSS AH, QIU RG, SYMONS M, AND EXTON JH. Activation
of rat liver phospholipase D by the small GTP-binding protein
RhoA. J Biol Chem 269: 2595125954, 1994.
MAMMOTO A, OHTSUKA T, HOTTA I, SASAKI T, AND TAKAI Y. Rab11BP/

433.

434.

435.

436.

437.

438.

439.

440.

441.

442.

443.

444.

445.

446.

447.
448.

449.

450.

451.

452.

199

Rabphilin-11, a downstream target of rab11 small G protein implicated in vesicle recycling. J Biol Chem 274: 2551725524, 1999.
MAMMOTO A, SASAKI T, KIM Y, AND TAKAI Y. Physical and functional
interaction of rabphilin-11 with mammalian Sec13 protein. Implication in vesicle trafficking. J Biol Chem 275: 1316713170, 2000.
MANDAI K, NAKANISHI H, SATOH A, OBAISHI H, WADA M, NISHIOKA H,
ITOH M, MIZOGUCHI A, AOKI T, FUJIMOTO T, MATSUDA Y, TSUKITA S, AND
TAKAI Y. Afadin: a novel actin filament-binding protein with one
PDZ domain localized at cadherin-based cell-to-cell adherens junction. J Cell Biol 139: 517528, 1997.
MANDIYAN V, ANDREEV J, SCHLESSINGER J, AND HUBBARD SR. Crystal
structure of the ARF-GAP domain and ankyrin repeats of PYK2associated protein . EMBO J 18: 6890 6898, 1999.
MANNING BD, PADMANABHA R, AND SNYDER M. The Rho-GEF Rom2p
localizes to sites of polarized cell growth and participates in cytoskeletal functions in Saccharomyces cerevisiae. Mol Biol Cell 8:
1829 1844, 1997.
MANSER E, HUANG HY, LOO TH, CHEN XQ, DONG JM, LEUNG T, AND LIM
L. Expression of constitutively active -PAK reveals effects of the
kinase on actin and focal complexes. Mol Cell Biol 17: 1129 1143,
1997.
MANSER E, LEUNG T, SALIHUDDIN H, TAN L, AND LIM L. A non-receptor
tyrosine kinase that inhibits the GTPase activity of p21cdc42. Nature 363: 364 367, 1993.
MANSER E, LEUNG T, SALIHUDDIN H, ZHAO ZS, AND LIM L. A brain
serine/threonine protein kinase activated by Cdc42 and Rac1. Nature 367: 40 46, 1994.
MANSOUR SJ, SKAUG J, ZHAO XH, GIORDANO J, SCHERER SW, AND
MELANCON P. p200 ARF-GEP1: a Golgi-localized guanine nucleotide
exchange protein whose Sec7 domain is targeted by the drug
brefeldin A. Proc Natl Acad Sci USA 96: 7968 7973, 1999.
MARAIS R, LIGHT Y, PATERSON HF, AND MARSHALL CJ. Ras recruits
Raf-1 to the plasma membrane for activation by tyrosine phosphorylation. EMBO J 14: 3136 3145, 1995.
MARAIS R, WYNNE J, AND TREISMAN R. The SRF accessory protein
Elk-1 contains a growth factor-regulated transcriptional activation
domain. Cell 73: 381393, 1993.
MARCUS S, POLVERINO A, CHANG E, ROBBINS D, COBB MH, AND WIGLER
MH. Shk1, a homolog of the Saccharomyces cerevisiae Ste20 and
mammalian p65PAK protein kinases, is a component of a Ras/
Cdc42 signaling module in the fission yeast Schizosaccharomyces
pombe. Proc Natl Acad Sci USA 92: 6180 6184, 1995.
MARTE BM, RODRIGUEZ-VICIANA P, WENNSTROM S, WARNE PH, AND
DOWNWARD J. R-Ras can activate the phosphoinositide 3-kinase but
not the MAP kinase arm of the Ras effector pathways. Curr Biol 7:
6370, 1997.
MARTIN GA, VISKOCHIL D, BOLLAG G, MCCABE PC, CROSIER WJ,
HAUBRUCK H, CONROY L, CLARK R, OCONNELL P, AND CAWTHON RM.
The GAP-related domain of the neurofibromatosis type 1 gene
product interacts with ras p21. Cell 63: 843 849, 1990.
MARTIN H, MENDOZA A, RODRIGUEZ-PACHON JM, MOLINA M, AND NOMBELA C. Characterization of SKM1, a Saccharomyces cerevisiae
gene encoding a novel Ste20/PAK-like protein kinase. Mol Microbiol 23: 431 444, 1997.
MARTINEZ O AND GOUD B. Rab proteins. Biochim Biophys Acta 1404:
101112, 1998.
MARTINEZ O, SCHMIDT A, SALAMERO J, HOFLACK B, ROA M, AND GOUD B.
The small GTP-binding protein rab6 functions in intra-Golgi transport. J Cell Biol 127: 15751588, 1994.
MASSOL P, MONTCOURRIER P, GUILLEMOT JC, AND CHAVRIER P. Fc
receptor-mediated phagocytosis requires CDC42 and Rac1. EMBO
J 17: 6219 6229, 1998.
MATSUDA S, KOSAKO H, TAKENAKA K, MORIYAMA K, SAKAI H, AKIYAMA T,
GOTOH Y, AND NISHIDA E. Xenopus MAP kinase activator: identification and function as a key intermediate in the phosphorylation
cascade. EMBO J 11: 973982, 1992.
MATSUI T, AMANO M, YAMAMOTO T, CHIHARA K, NAKAFUKU M, ITO M,
NAKANO T, OKAWA K, IWAMATSU A, AND KAIBUCHI K. Rho-associated
kinase, a novel serine/threonine kinase, as a putative target for
small GTP binding protein Rho. EMBO J 15: 2208 2216, 1996.
MATSUI T, YONEMURA S, TSUKITA S, AND TSUKITA S. Activation of ERM
proteins in vivo by Rho involves phosphatidyl-inositol 4-phosphate
5-kinase and not ROCK kinases. Curr Biol 9: 1259 1262, 1999.

200

TAKAI, SASAKI, AND MATOZAKI

453. MATSUI Y, KIKUCHI A, ARAKI S, HATA Y, KONDO J, TERANISHI Y, AND

TAKAI Y. Molecular cloning and characterization of a novel type of
regulatory protein (GDI) for smg p25A, a ras p21-like GTP-binding
protein. Mol Cell Biol 10: 4116 4122, 1990.
454. MATTAJ IW AND ENGLMEIER L. Nucleocytoplasmic transport: the
soluble phase. Annu Rev Biochem 67: 265306, 1998.
455. MATUNIS MJ, COUTAVAS E, AND BLOBEL G. A novel ubiquitin-like
modification modulates the partitioning of the Ran-GTPase-activating protein RanGAP1 between the cytosol and the nuclear pore
complex. J Cell Biol 135: 14571470, 1996.
456. MAZUR P AND BAGINSKY W. In vitro activity of 1,3--D-glucan synthase
requires the GTP-binding protein Rho1. J Biol Chem 271: 14604
14609, 1996.
457. MCBRIDE HM, RYBIN V, MURPHY C, GINER A, TEASDALE R, AND ZERIAL
M. Oligomeric complexes link Rab5 effectors with NSF and drive
membrane fusion via interactions between EEA1 and syntaxin 13.
Cell 98: 377386, 1999.
458. MCCALLUM SJ, WU WJ, AND CERIONE RA. Identification of a putative
effector for Cdc42Hs with high sequence similarity to the RasGAPrelated protein IQGAP1 and a Cdc42Hs binding partner with similarity to IQGAP2. J Biol Chem 271: 2173221737, 1996.
459. MCCORMICK F AND WITTINGHOFER A. Interactions between Ras proteins and their effectors. Curr Opin Biotechnol 7: 449 456, 1996.
460. MCKIERNAN CJ, STABILA PF, AND MACARA IG. Role of the Rab3Abinding domain in targeting of rabphilin-3A to vesicle membranes
of PC12 cells. Mol Cell Biol 16: 4985 4995, 1996.
461. MCLAUCHLAN H, NEWELL J, MORRICE N, OSBORNE A, WEST M, AND
SMYTHE E. A novel role for Rab5-GDI in ligand sequestration into
clathrin-coated pits. Curr Biol 8: 34 45, 1998.
462. MCLEOD SJ, INGHAM RJ, BOS JL, KUROSAKI T, AND GOLD MR. Activation of the Rap1 GTPase by the B cell antigen receptor. J Biol Chem
273: 29218 29223, 1998.
463. MEDEMA RH, DE LAAT WL, MARTIN GA, MCCORMICK F, AND BOS JL.
GTPase-activating protein SH2-SH3 domains induce gene expression in a Ras-dependent fashion. Mol Cell Biol 12: 34253430, 1992.
464. MELCHIOR F, PASCHAL B, EVANS J, AND GERACE L. Inhibition of nuclear
protein import by nonhydrolyzable analogues of GTP and identification of the small GTPase Ran/TC4 as an essential transport
factor. J Cell Biol 123: 1649 1659, 1993.
465. MELLMAN I AND WARREN G. The road taken: past and future foundations of membrane traffic. Cell 100: 99 112, 2000.
466. MERDES A, RAMYAR K, VECHIO JD, AND CLEVELAND DW. A complex of
NuMA and cytoplasmic dynein is essential for mitotic spindle assembly. Cell 87: 447 458, 1996.
467. MICHAUD NR, FABIAN JR, MATHES KD, AND MORRISON DK. 14 3-3 is
not essential for Raf-1 function: identification of Raf-1 proteins that
are biologically activated in a 14 3-3- and Ras-independent manner.
Mol Cell Biol 15: 3390 3397, 1995.
468. MICHIELS F, HABETS GG, STAM JC, VAN DER KAMMEN RA, AND COLLARD
JG. A role for Rac in Tiam1-induced membrane ruffling and invasion. Nature 375: 338 340, 1995.
469. MICHIELS F, STAM JC, HORDIJK PL, VAN DER KAMMEN RA, RUULS-VAN
STALLE L, FELTKAMP CA, AND COLLARD JG. Regulated membrane
localization of Tiam1, mediated by the NH2-terminal pleckstrin
homology domain, is required for Rac-dependent membrane ruffling and c-Jun NH2-terminal kinase activation. J Cell Biol 137:
387398, 1997.
470. MIKI H, SASAKI T, TAKAI Y, AND TAKENAWA T. Induction of filopodium
formation by a WASP-related actin-depolymerizing protein
N-WASP. Nature 391: 9396, 1998.
471. MILBURN MV, TONG L, DEVOS AM, BRUNGER A, YAMAIZUMI Z, NISHIMURA
S, AND KIM SH. Molecular switch for signal transduction: structural
differences between active and inactive forms of protooncogenic
ras proteins. Science 247: 939 945, 1990.
472. MILLER RK, MATHEOS D, AND ROSE MD. The cortical localization of
the microtubule orientation protein, Kar9p, is dependent upon
actin and proteins required for polarization. J Cell Biol 144: 963
975, 1999.
473. MINDEN A, LIN A, CLARET FX, ABO A, AND KARIN M. Selective activation of the JNK signaling cascade and c-Jun transcriptional activity
by the small GTPases Rac and Cdc42Hs. Cell 81: 11471157, 1995.
474. MISHIMA K, TSUCHIYA M, NIGHTINGALE MS, MOSS J, AND VAUGHAN M.
ARD 1, a 64-kDa guanine nucleotide-binding protein with a carbox-

475.

476.

477.

478.

479.

480.

481.

482.
483.
484.

485.

486.

487.

488.

489.

490.

491.
492.

493.

494.
495.

Volume 81

yl-terminal ADP-ribosylation factor domain. J Biol Chem 268:

8801 8807, 1993.
MIURA Y, KIKUCHI A, MUSHA T, KURODA S, YAKU H, SASAKI T, AND TAKAI
Y. Regulation of morphology by rho p21 and its inhibitory GDP/
GTP exchange protein (rho GDI) in Swiss 3T3 cells. J Biol Chem
268: 510 515, 1993.
MIZOGUCHI A, KIM S, UEDA T, KIKUCHI A, YORIFUJI H, HIROKAWA N, AND
TAKAI Y. Localization and subcellular distribution of smg p25A, a
ras p21-like GTP-binding protein, in rat brain. J Biol Chem 265:
1187211879, 1990.
MIZOGUCHI A, KIM S, UEDA T, AND TAKAI Y. Tissue distribution of smg
p25A, a ras p21-like GTP-binding protein, studied by use of a
specific monoclonal antibody. Biochem Biophys Res Commun 162:
1438 1445, 1989.
MIZOGUCHI A, YANO Y, HAMAGUCHI H, YANAGIDA H, IDE C, ZAHRAOUI A,
SHIRATAKI H, SASAKI T, AND TAKAI Y. Localization of Rabphilin-3A on
the synaptic vesicle. Biochem Biophys Res Commun 202: 1235
1243, 1994.
MIZUNO T, KAIBUCHI K, ANDO S, MUSHA T, HIRAOKA K, TAKAISHI K,
ASADA M, NUNOI H, MATSUDA I, AND TAKAI Y. Regulation of the
superoxide-generating NADPH oxidase by a small GTP-binding
protein and its stimulatory and inhibitory GDP/GTP exchange proteins. J Biol Chem 267: 1021510218, 1992.
MIZUNO T, KAIBUCHI K, YAMAMOTO T, KAWAMURA M, SAKODA T, FUJIOKA
H, MATSUURA Y, AND TAKAI Y. A stimulatory GDP/GTP exchange
protein for smg p21 is active on the posttranslationally processed
form of c-Ki-ras p21 and rhoA p21. Proc Natl Acad Sci USA 88:
6442 6446, 1991.
MOLLOY CJ, BOTTARO DP, FLEMING TP, MARSHALL MS, GIBBS JB, AND
AARONSON SA. PDGF induction of tyrosine phosphorylation of GTPase activating protein. Nature 342: 711714, 1989.
MOORE I, SCHELL J, AND PALME K. Subclass-specific sequence motifs
identified in Rab GTPases. Trends Biochem Sci 20: 10 12, 1995.
MOORE MS. Ran and nuclear transport. J Biol Chem 273: 22857
22860, 1998.
MOORE MS AND BLOBEL G. The GTP-binding protein Ran/TC4 is
required for protein import into the nucleus. Nature 365: 661 663,
1993.
MOORE MS AND BLOBEL G. Purification of a Ran-interacting protein
that is required for protein import into the nucleus. Proc Natl Acad
Sci USA 91: 1021210216, 1994.
MORAN MF, KOCH CA, ANDERSON D, ELLIS C, ENGLAND L, MARTIN GS,
AND PAWSON T. Src homology region 2 domains direct proteinprotein interactions in signal transduction. Proc Natl Acad Sci USA
87: 8622 8626, 1990.
MORELAND S, NISHIMURA J, VAN BREEMEN C, AHN HY, AND MORELAND
RS. Transient myosin phosphorylation at constant Ca2 during
agonist activation of permeabilized arteries. Am J Physiol Cell
Physiol 263: C540 C544, 1992.
MORII N, KAWANO K, SEKINE A, YAMADA T, AND NARUMIYA S. Purification of GTPase-activating protein specific for the rho gene products. J Biol Chem 266: 7646 7650, 1991.
MORINAGA N, ADAMIK R, MOSS J, AND VAUGHAN M. Brefeldin A inhibited activity of the sec7 domain of p200, a mammalian guanine
nucleotide-exchange protein for ADP-ribosylation factors. J Biol
Chem 274: 1741717423, 1999.
MORINAGA N, MOSS J, AND VAUGHAN M. Cloning and expression of a
cDNA encoding a bovine brain brefeldin A-sensitive guanine nucleotide-exchange protein for ADP-ribosylation factor. Proc Natl Acad
Sci USA 94: 12926 12931, 1997.
MORRISON DK AND CUTLER RE. The complexity of Raf-1 regulation.
Curr Opin Cell Biol 9: 174 179, 1997.
MORRISON DK, HEIDECKER G, RAPP UR, AND COPELAND TD. Identification of the major phosphorylation sites of the Raf-1 kinase. J Biol
Chem 268: 17309 17316, 1993.
MOSS J AND VAUGHAN M. Structure and function of ARF proteins:
activators of cholera toxin and critical components of intracellular
vesicular transport processes. J Biol Chem 270: 1232712330, 1995.
MOSS J AND VAUGHAN M. Molecules in the ARF orbit. J Biol Chem
273: 2143121434, 1998.
MOTT HR, CARPENTER JW, ZHONG S, GHOSH S, BELL RM, AND CAMPBELL
SL. The solution structure of the Raf-1 cysteine-rich domain: a

January 2001

SMALL G PROTEINS

novel ras and phospholipid binding site. Proc Natl Acad Sci USA 93:
8312 8317, 1996.
496. MOUNIER J, LAURENT V, HALL A, FORT P, CARLIER MF, SANSONETTI PJ,
AND EGILE C. Rho family GTPases control entry of Shigella flexneri
into epithelial cells but not intracellular motility. J Cell Sci 112:
2069 2080, 1999.
497. MOYA M, ROBERTS D, AND NOVICK P. DSS4 1 is a dominant suppressor of sec4 8 that encodes a nucleotide exchange protein that aids
Sec4p function. Nature 361: 460 463, 1993.
497a.MRABET L, COFFER P, ZWARTKRUIS F, FRANKE B, SEGAL AW, KOENDERMAN L, AND BOS JL. Activation of the small GTPase rap1 in human
neutrophils. Blood 92: 21332140, 1998.
498. MURPHY C, SAFFRICH R, GRUMMT M, GOURNIER H, RYBIN V, RUBINO M,
AUVINEN P, LUTCKE A, PARTON RG, AND ZERIAL M. Endosome dynamics regulated by a Rho protein. Nature 384: 427 432, 1996.
499. MURRAY MJ, CUNNINGHAM JM, PARADA LF, DAUTRY F, LEBOWITZ P, AND
WEINBERG RA. The HL-60 transforming sequence: a ras oncogene
coexisting with altered myc genes in hematopoietic tumors. Cell 33:
749 757, 1983.
500. MUSLIN AJ, TANNER JW, ALLEN PM, AND SHAW AS. Interaction of
14 3-3 with signaling proteins is mediated by the recognition of
phosphoserine. Cell 84: 889 897, 1996.
501. NAGANO F, SASAKI T, FUKUI K, ASAKURA T, IMAZUMI K, AND TAKAI Y.
Molecular cloning and characterization of the noncatalytic subunit
of the Rab3 subfamily-specific GTPase-activating protein. J Biol
Chem 273: 2478124785, 1998.
502. NAGATA K, PULS A, FUTTER C, ASPENSTROM P, SCHAEFER E, NAKATA T,
HIROKAWA N, AND HALL A. The MAP kinase kinase kinase MLK2
co-localizes with activated JNK along microtubules and associates
with kinesin superfamily motor KIF3. EMBO J 17: 149 158, 1998.
503. NAGEL W, ZEITLMANN L, SCHILCHER P, GEIGER C, KOLANUS J, AND
KOLANUS W. Phosphoinositide 3-OH kinase activates the 2 integrin
adhesion pathway and induces membrane recruitment of cytohesin-1. J Biol Chem 273: 1485314861, 1998.
504. NAKAMURA M, MASUDA H, HORII J, KUMA K, YOKOYAMA N, OHBA T,
NISHITANI H, MIYATA T, TANAKA M, AND NISHIMOTO T. When overexpressed, a novel centrosomal protein, RanBPM, causes ectopic
microtubule nucleation similar to -tubulin. J Cell Biol 143: 1041
1052, 1998.
505. NAKAMURA N, LOWE M, LEVINE TP, RABOUILLE C, AND WARREN G. The
vesicle docking protein p115 binds GM130, a cis-Golgi matrix protein, in a mitotically regulated manner. Cell 89: 445 455, 1997.
506. NAKANISHI H, KAIBUCHI K, ORITA S, UENO N, AND TAKAI Y. Different
functions of Smg GDP dissociation stimulator and mammalian
counterpart of yeast Cdc25. J Biol Chem 269: 1508515091, 1994.
507. NAKANISHI H, ORITA S, KAIBUCHI K, MIURA K, MIKI H, TAKENAWA T, AND
TAKAI Y. Kinetic properties of Ash/Grb2-interacting GDP/GTP exchange protein. Biochem Biophys Res Commun 198: 12551261,
1994.
508. NAKANO A, BRADA D, AND SCHEKMAN R. A membrane glycoprotein,
Sec12p, required for protein transport from the endoplasmic reticulum to the Golgi apparatus in yeast. J Cell Biol 107: 851 863, 1988.
509. NAKANO A AND MURAMATSU M. A novel GTP-binding protein, Sar1p,
is involved in transport from the endoplasmic reticulum to the
Golgi apparatus. J Cell Biol 109: 26772691, 1989.
510. NAKANO K, TAKAISHI K, KODAMA A, MAMMOTO A, SHIOZAKI H, MONDEN
M, AND TAKAI Y. Distinct actions and cooperative roles of ROCK and
mDia in Rho small G protein-induced reorganization of the actin
cytoskeleton in Madin-Darby canine kidney cells. Mol Biol Cell 10:
24812491, 1999.
511. NAKASHIMA S, MORINAKA K, KOYAMA S, IKEDA M, KISHIDA M, OKAWA K,
IWAMATSU A, KISHIDA S, AND KIKUCHI A. Small G protein Ral and its
downstream molecules regulate endocytosis of EGF and insulin
receptors. EMBO J 18: 3629 3642, 1999.
512. NARUMIYA S, SEKINE A, AND FUJIWARA M. Substrate for botulinum
ADP-ribosyltransferase, Gb, has an amino acid sequence homologous to a putative rho gene product. J Biol Chem 263: 1725517257,
1988.
513. NASSAR N, HORN G, HERRMANN C, SCHERER A, MCCORMICK F, AND
WITTINGHOFER A. The 2.2 A crystal structure of the Ras-binding
domain of the serine/threonine kinase c-Raf1 in complex with
Rap1A and a GTP analogue. Nature 375: 554 560, 1995.
514. NEWMAN SL, MIKUS LK, AND TUCCI MA. Differential requirements for

515.

516.

517.
518.

519.
520.

521.

522.

523.

524.

525.

526.

527.

528.
529.

530.

532.

533.

534.

535.

201

cellular cytoskeleton in human macrophage complement receptorand Fc receptor-mediated phagocytosis. J Immunol 146: 967974,
1991.
NEWSOME TP, SCHMIDT S, DIETZL G, KELEMAN K, ASLING B, DEBANT A,
AND DICKSON BJ. Trio combines with dock to regulate Pak activity
during photoreceptor axon pathfinding in Drosophila. Cell 101:
283294, 2000.
NIELSEN E, SEVERIN F, BACKER JM, HYMAN AA, AND ZERIAL M. Rab5
regulates motility of early endosomes on microtubules. Nat Cell
Biol 1: 376 382, 1999.
NIGG EA. Nucleocytoplasmic transport: signals, mechanisms and
regulation. Nature 386: 779 787, 1997.
NISHIKAWA S AND NAKANO A. The GTP-binding Sar1 protein is localized to the early compartment of the yeast secretory pathway.
Biochim Biophys Acta 1093: 135143, 1991.
NISHIMOTO T. A new role of ran GTPase. Biochem Biophys Res
Commun 262: 571574, 1999.
NISHIMURA N, NAKAMURA H, TAKAI Y, AND SANO K. Molecular cloning
and characterization of two rab GDI species from rat brain: brainspecific and ubiquitous types. J Biol Chem 269: 1419114198, 1994.
NISHIYAMA T, SASAKI T, TAKAISHI K, KATO M, YAKU H, ARAKI K, MATSUURA Y, AND TAKAI Y. rac p21 is involved in insulin-induced membrane ruffling and rho p21 is involved in hepatocyte growth factorand 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced membrane ruffling in KB cells. Mol Cell Biol 14: 24472456, 1994.
NOBES CD AND HALL A. Rho, rac, and cdc42 GTPases regulate the
assembly of multimolecular focal complexes associated with actin
stress fibers, lamellipodia, and filopodia. Cell 81: 53 62, 1995.
NODA M, KO M, OGURA A, LIU DG, AMANO T, TAKANO T, AND IKAWA Y.
Sarcoma viruses carrying ras oncogenes induce differentiationassociated properties in a neuronal cell line. Nature 318: 7375,
1985.
NODA M, YASUDA-FUKAZAWA C, MORIISHI K, KATO T, OKUDA T, KUROKAWA K, AND TAKUWA Y. Involvement of rho in GTPS-induced
enhancement of phosphorylation of 20 kDa myosin light chain in
vascular smooth muscle cells: inhibition of phosphatase activity.
FEBS Lett 367: 246 250, 1995.
NONAKA H, TANAKA K, HIRANO H, FUJIWARA T, KOHNO H, UMIKAWA M,
MINO A, AND TAKAI Y. A downstream target of RHO1 small GTPbinding protein is PKC1, a homolog of protein kinase C, which
leads to activation of the MAP kinase cascade in Saccharomyces
cerevisiae. EMBO J 14: 59315938, 1995.
NORMAN JC, PRICE LS, RIDLEY AJ, AND KOFFER A. The small GTPbinding proteins, Rac and Rho, regulate cytoskeletal organization
and exocytosis in mast cells by parallel pathways. Mol Biol Cell 7:
1429 1442, 1996.
NOVICK P, FIELD C, AND SCHEKMAN R. Identification of 23 complementation groups required for posttranslational events in the yeast
secretory pathway. Cell 21: 205215, 1980.
NOVICK P AND ZERIAL M. The diversity of Rab proteins in vesicle
transport. Curr Opin Cell Biol 9: 496 504, 1997.
NUOFFER C AND BALCH WE. GTPases: multifunctional molecular
switches regulating vesicular traffic. Annu Rev Biochem 63: 949
990, 1994.
NUOFFER C, DAVIDSON HW, MATTESON J, MEINKOTH J, AND BALCH WE.
A GDP-bound of rab1 inhibits protein export from the endoplasmic
reticulum and transport between Golgi compartments. J Cell Biol
125: 225237, 1994.
OBAISHI H, NAKANISHI H, MANDAI K, SATOH K, SATOH A, TAKAHASHI K,
MIYAHARA M, NISHIOKA H, TAKAISHI K, AND TAKAI Y. Frabin, a novel
FGD1-related actin filament-binding protein capable of changing
cell shape and activating c-Jun N-terminal kinase. J Biol Chem 273:
1869718700, 1998.
OHBA T, NAKAMURA M, NISHITANI H, AND NISHIMOTO T. Self-organization of microtubule asters induced in Xenopus egg extracts by
GTP-bound Ran. Science 284: 1356 1358, 1999.
OHMORI T, KIKUCHI A, YAMAMOTO K, KIM S, AND TAKAI Y. Small
molecular weight GTP-binding proteins in human platelet membranes. Purification and characterization of a novel GTP-binding
protein with a molecular weight of 22,000. J Biol Chem 264: 1877
1881, 1989.
OHMORI T, TAKEYAMA Y, UEDA T, HIROYOSHI M, NAKANISHI H, OHYANAGI
H, SAITOH Y, AND TAKAI Y. Purification and characterization of a

202

TAKAI, SASAKI, AND MATOZAKI

novel GTP-binding protein with a Mr value of 24,000 from rat liver.

Biochem Biophys Res Commun 169: 816 823, 1990.
536. OHNO M, FORNEROD M, AND MATTAJ IW. Nucleocytoplasmic transport: the last 200 nanometers. Cell 92: 327336, 1998.
537. OHTA Y, SUZUKI N, NAKAMURA S, HARTWIG JH, AND STOSSEL TP. The
small GTPase RalA targets filamin to induce filopodia. Proc Natl
Acad Sci USA 96: 21222128, 1999.
538. OHTSUBO M, KAI R, FURUNO N, SEKIGUCHI T, SEKIGUCHI M, HAYASHIDA
H, KUMA K, MIYATA T, FUKUSHIGE S, AND MUROTSU T. Isolation and
characterization of the active cDNA of the human cell cycle gene
(RCC1) involved in the regulation of onset of chromosome condensation. Genes Dev 1: 585593, 1987.
539. OHTSUBO M, OKAZAKI H, AND NISHIMOTO T. The RCC1 protein, a
regulator for the onset of chromosome condensation locates in the
nucleus and binds to DNA. J Cell Biol 109: 1389 1397, 1989.
540. OHTSUKA T, HATA Y, IDE N, YASUDA T, INOUE E, INOUE T, MIZOGUCHI A,
AND TAKAI Y. nRap GEP: a novel neural GDP/GTP exchange protein
for rap1 small G protein that interacts with synaptic scaffolding
molecule (S-SCAM). Biochem Biophys Res Commun 265: 38 44,
1999.
541. OHTSUKA T, SHIMIZU K, YAMAMORI B, KURODA S, AND TAKAI Y. Activation of brain B-Raf protein kinase by Rap1B small GTP-binding
protein. J Biol Chem 271: 1258 1261, 1996.
542. OHYA T, SASAKI T, KATO M, AND TAKAI Y. Involvement of Rabphilin3
in endocytosis through interaction with Rabaptin5. J Biol Chem
273: 613 617, 1998.
543. OISHI H, SASAKI T, NAGANO F, IKEDA W, OHYA T, WADA M, IDE N,
NAKANISHI H, AND TAKAI Y. Localization of the Rab3 small G protein
regulators in nerve terminals and their involvement in Ca2-dependent exocytosis. J Biol Chem 273: 34580 34585, 1998.
544. OKA T AND NAKANO A. Inhibition of GTP hydrolysis by Sar1p causes
accumulation of vesicles that are a functional intermediate of the
ER-to-Golgi transport in yeast. J Cell Biol 124: 425 434, 1994.
545. OKA T, NISHIKAWA S, AND NAKANO A. Reconstitution of GTP-binding
Sar1 protein function in ER to Golgi transport. J Cell Biol 114:
671 679, 1991.
546. OKADA S, MATSUDA M, ANAFI M, PAWSON T, AND PESSIN JE. Insulin
regulates the dynamic balance between Ras and Rap1 signaling by
coordinating the assembly states of the Grb2-SOS and CrkII-C3G
complexes. EMBO J 17: 2554 2565, 1998.
547. OKADA T, MASUDA T, SHINKAI M, KARIYA K, AND KATAOKA T. Posttranslational modification of H-Ras is required for activation of, but not
for association with, B-Raf. J Biol Chem 271: 4671 4678, 1996.
548. OKAZAKI M, KISHIDA S, HINOI T, HASEGAWA T, TAMADA M, KATAOKA T,
AND KIKUCHI A. Synergistic activation of c-fos promoter activity by
Raf and Ral GDP dissociation stimulator. Oncogene 14: 515521,
1997.
549. OLIVIER JP, RAABE T, HENKEMEYER M, DICKSON B, MBAMALU G, MARGOLIS B, SCHLESSINGER J, HAFEN E, AND PAWSON T. A Drosophila
SH2-SH3 adaptor protein implicated in coupling the sevenless tyrosine kinase to an activator of Ras guanine nucleotide exchange,
Sos. Cell 73: 179 191, 1993.
550. OLKKONEN VM AND STENMARK H. Role of Rab GTPases in membrane
traffic. Int Rev Cytol 176: 1 85, 1997.
551. OLOFSSON B, CHARDIN P, TOUCHOT N, ZAHRAOUI A, AND TAVITIAN A.
Expression of the ras-related ralA, rho12 and rab genes in adult
mouse tissues. Oncogene 3: 231234, 1988.
552. OLSON MF, ASHWORTH A, AND HALL A. An essential role for Rho, Rac,
and Cdc42 GTPases in cell cycle progression through G1. Science
269: 1270 1272, 1995.
553. ONO Y, NAKANISHI H, NISHIMURA M, KAKIZAKI M, TAKAHASHI K, MIYAHARA M, SATOH-HORIKAWA K, MANDAI K, AND TAKAI Y. Two actions of
frabin: direct activation of Cc42 and indirect activation of Rac.
Oncogene 19: 3050 3058, 2000.
554. ORCI L, RAVAZZOLA M, MEDA P, HOLCOMB C, MOORE HP, HICKE L, AND
SCHEKMAN R. Mammalian Sec23p homologue is restricted to the
endoplasmic reticulum transitional cytoplasm. Proc Natl Acad Sci
USA 88: 8611 8615, 1991.
555. OSTERMEIER C AND BRUNGER AT. Structural basis of Rab effector
specificity: crystal structure of the small G protein Rab3A complexed with the effector domain of rabphilin-3A. Cell 96: 363374,
1999.
555a.OSULLIVAN AJ, BROWN AM, FREEMAN HN, AND GOMPERTS BD. Purifi-

556.

557.

558.

559.

560.

561.

562.
563.

564.

565.
566.

567.

568.

569.

570.

571.
572.

573.

575.

576.

577.

578.

Volume 81

cation and identification of FOAD-II, a cytosolic protein that regulates secretion in streptolysin-O permeabilized mast cells, as a
rac/rhoGDI complex. Mol Biol Cell 7: 397 408, 1996.
OTTILIE S, MILLER PJ, JOHNSON DI, CREASY CL, SELLS MA, BAGRODIA S,
FORSBURG SL, AND CHERNOFF J. Fission yeast pak1 encodes a
protein kinase that interacts with Cdc42p and is involved in the
control of cell polarity and mating. EMBO J 14: 5908 5919, 1995.
OTTO JC, KIM E, YOUNG SG, AND CASEY PJ. Cloning and characterization of a mammalian prenyl protein-specific protease. J Biol
Chem 274: 8379 8382, 1999.
PACCAUD JP, REITH W, CARPENTIER JL, RAVAZZOLA M, AMHERDT M,
SCHEKMAN R, AND ORCI L. Cloning and functional characterization of
mammalian homologues of the COPII component Sec23. Mol Biol
Cell 7: 15351546, 1996.
PAGANO A, LETOURNEUR F, GARCIA-ESTEFANIA D, CARPENTIER JL, ORCI
L, AND PACCAUD JP. Sec24 proteins and sorting at the endoplasmic
reticulum. J Biol Chem 274: 78337840, 1999.
PAI EF, KABSCH W, KRENGEL U, HOLMES KC, JOHN J, AND WITTINGHOFER A. Structure of the guanine-nucleotide-binding domain of the
Ha-ras oncogene product p21 in the triphosphate conformation.
Nature 341: 209 214, 1989.
PARADA LF, TABIN CJ, SHIH C, AND WEINBERG RA. Human EJ bladder
carcinoma oncogene is homologue of Harvey sarcoma virus ras
gene. Nature 297: 474 478, 1982.
PARK RK, LIU Y, AND DURDEN DL. A role for Shc, Grb2, and Raf-1 in
FcRI signal relay. J Biol Chem 271: 1334213348, 1996.
PASCHAL BM AND GERACE L. Identification of NTF2, a cytosolic
factor for nuclear import that interacts with nuclear pore complex
protein p62. J Cell Biol 129: 925937, 1995.
PATERSON HF, SELF AJ, GARRETT MD, JUST I, AKTORIES K, AND HALL A.
Microinjection of recombinant p21rho induces rapid changes in
cell morphology. J Cell Biol 111: 10011007, 1990.
PENNISI E. The nucleuss revolving door. Science 279: 1129 1131,
1998.
PERIN MS, FRIED VA, MIGNERY GA, JAHN R, AND SUDHOF TC. Phospholipid binding by a synaptic vesicle protein homologous to the
regulatory region of protein kinase C. Nature 345: 260 263, 1990.
PERONA R, MONTANER S, SANIGER L, SANCHEZ-PEREZ I, BRAVO R, AND
LACAL JC. Activation of the nuclear factor-kappaB by Rho, CDC42,
and Rac-1 proteins. Genes Dev 11: 463 475, 1997.
PETER M, CHAVRIER P, NIGG EA, AND ZERIAL M. Isoprenylation of rab
proteins on structurally distinct cysteine motifs. J Cell Sci 102:
857 865, 1992.
PETERSON MR, BURD CG, AND EMR SD. Vac1p coordinates Rab and
phosphatidylinositol 3-kinase signaling in Vps45p-dependent vesicle docking/fusion at the endosome. Curr Biol 9: 159 162, 1999.
PEYROCHE A, ANTONNY B, ROBINEAU S, ACKER J, CHERFILS J, AND
JACKSON CL. Brefeldin A acts to stabilize an abortive ARF-GDP-Sec7
domain protein complex: involvement of specific residues of the
Sec7 domain. Mol Cell 3: 275285, 1999.
PFEFFER SR. Transport-vesicle targeting: tethers before SNAREs.
Nat Cell Biol 1: E17E22, 1999.
PFEFFER SR, DIRAC-SVEJSTRUP AB, AND SOLDATI T. Rab GDP dissociation inhibitor: putting rab GTPases in the right place. J Biol Chem
270: 1705717059, 1995.
PIZON V, CHARDIN P, LEROSEY I, OLOFSSON B, AND TAVITIAN A. Human
cDNAs rap1 and rap2 homologous to the Drosophila gene Dras3
encode proteins closely related to ras in the effector region.
Oncogene 3: 201204, 1988.
PLUTNER H, COX AD, PIND S, KHOSRAVI-FAR R, BOURNE JR,
SCHWANINGER R, DER CJ, AND BALCH WE. Rab1b regulates vesicular
transport between the endoplasmic reticulum and successive Golgi
compartments. J Cell Biol 115: 31 43, 1991.
POPIK W AND PITHA PM. Binding of human immunodeficiency virus
type 1 to CD4 induces association of Lck and Raf-1 and activates
Raf-1 by a Ras-independent pathway. Mol Cell Biol 16: 6532 6541,
1996.
POWERS S, KATAOKA T, FASANO O, GOLDFARB M, STRATHERN J, BROACH
J, AND WIGLER M. Genes in S. cerevisiae encoding proteins with
domains homologous to the mammalian ras proteins. Cell 36: 607
612, 1984.
PREMONT RT, CLAING A, VITALE N, FREEMAN JL, PITCHER JA, PATTON
WA, MOSS J, VAUGHAN M, AND LEFKOWITZ RJ. 2-Adrenergic receptor

January 2001

579.
580.

581.

582.
583.
584.

585.

586.

587.

588.

589.

590.

591.

592.

593.

594.

595.
596.

597.

598.

599.

600.

SMALL G PROTEINS

regulation by GIT1, a G protein-coupled receptor kinase-associated

ADP ribosylation factor GTPase-activating protein. Proc Natl Acad
Sci USA 95: 1408214087, 1998.
PRYER NK, WUESTEHUBE LJ, AND SCHEKMAN R. Vesicle-mediated protein sorting. Annu Rev Biochem 61: 471516, 1992.
QADOTA H, PYTHON CP, INOUE SB, ARISAWA M, ANRAKU Y, ZHENG Y,
WATANABE T, LEVIN DE, AND OHYA Y. Identification of yeast Rho1p
GTPase as a regulatory subunit of 1,3--glucan synthase. Science
272: 279 281, 1996.
QIU RG, ABO A, MCCORMICK F, AND SYMONS M. Cdc42 regulates
anchorage-independent growth and is necessary for Ras transformation. Mol Cell Biol 17: 3449 3458, 1997.
QIU RG, CHEN J, KIRN D, MCCORMICK F, AND SYMONS M. An essential
role for Rac in Ras transformation. Nature 374: 457 459, 1995.
QIU RG, CHEN J, MCCORMICK F, AND SYMONS M. A role for Rho in Ras
transformation. Proc Natl Acad Sci USA 92: 1178111785, 1995.
RADHAKRISHNA H, AL-AWAR O, KHACHIKIAN Z, AND DONALDSON JG.
ARF6 requirement for Rac ruffling suggests a role for membrane
trafficking in cortical actin rearrangements. J Cell Sci 112: 855 866,
1999.
RADHAKRISHNA H AND DONALDSON JG. ADP-ribosylation factor 6 regulates a novel plasma membrane recycling pathway. J Cell Biol 139:
49 61, 1997.
REEDQUIST KA AND BOS JL. Costimulation through CD28 suppresses
T cell receptor-dependent activation of the Ras-like small GTPase
Rap1 in human T lymphocytes. J Biol Chem 273: 4944 4949, 1998.
REEDQUIST KA, ROSS E, KOOP EA, WOLTHUIS RM, ZWARTKRUIS FJ, VAN
KOOYK Y, SALMON M, BUCKLEY CD, AND BOS JL. The small GTPase,
Rap1, mediates CD31-induced integrin adhesion. J Cell Biol 148:
11511158, 2000.
REID T, FURUYASHIKI T, ISHIZAKI T, WATANABE G, WATANABE N, FUJISAWA K, MORII N, MADAULE P, AND NARUMIYA S. Rhotekin, a new
putative target for Rho bearing homology to a serine/threonine
kinase, PKN, and rhophilin in the rho-binding domain. J Biol Chem
271: 13556 13560, 1996.
REINHARD J, SCHEEL AA, DIEKMANN D, HALL A, RUPPERT C, AND BAHLER
M. A novel type of myosin implicated in signalling by rho family
GTPases. EMBO J 14: 697704, 1995.
REN M, XU G, ZENG J, DE LEMOS-CHIARANDINI C, ADESNIK M, AND
SABATINI DD. Hydrolysis of GTP on rab11 is required for the direct
delivery of transferrin from the pericentriolar recycling compartment to the cell surface but not from sorting endosomes. Proc Natl
Acad Sci USA 95: 6187 6192, 1998.
REN M, ZENG J, DE LEMOS-CHIARANDINI C, ROSENFELD M, ADESNIK M,
AND SABATINI DD. In its active form, the GTP-binding protein rab8
interacts with a stress-activated protein kinase. Proc Natl Acad Sci
USA 93: 51515155, 1996.
REN XD, BOKOCH GM, TRAYNOR-KAPLAN A, JENKINS GH, ANDERSON RA,
AND SCHWARTZ MA. Physical association of the small GTPase Rho
with a 68-kDa phosphatidylinositol 4-phosphate 5-kinase in Swiss
3T3 cells. Mol Biol Cell 7: 435 442, 1996.
REN XD, KIOSSES WB, AND SCHWARTZ MA. Regulation of the small
GTP-binding protein Rho by cell adhesion and the cytoskeleton.
EMBO J 18: 578 585, 1999.
REYNET C AND KAHN CR. Rad: a member of the Ras family overexpressed in muscle of type II diabetic humans. Science 262: 1441
1444, 1993.
RIBBECK K, LIPOWSKY G, KENT HM, STEWART M, AND GORLICH D. NTF2
mediates nuclear import of Ran. EMBO J 17: 6587 6598, 1998.
RICHARDS SA, LOUNSBURY KM, CAREY KL, AND MACARA IG. A nuclear
export signal is essential for the cytosolic localization of the Ran
binding protein, RanBP1. J Cell Biol 134: 11571168, 1996.
RICHARDS SA, LOUNSBURY KM, AND MACARA IG. The C terminus of the
nuclear RAN/TC4 GTPase stabilizes the GDP-bound state and mediates interactions with RCC1, RAN-GAP, and HTF9A/RANBP1.
J Biol Chem 270: 1440514411, 1995.
RICHARDSON CJ, JONES S, LITT RJ, AND SEGEV N. GTP hydrolysis is not
important for Ypt1 GTPase function in vesicular transport. Mol Cell
Biol 18: 827 838, 1998.
RIDLEY AJ, COMOGLIO PM, AND HALL A. Regulation of scatter factor/
hepatocyte growth factor responses by Ras, Rac, and Rho in MDCK
cells. Mol Cell Biol 15: 1110 1122, 1995.
RIDLEY AJ AND HALL A. The small GTP-binding protein rho regulates

601.

602.

603.

604.

605.

606.

607.

608.

609.

610.

611.

612.
613.
614.
615.
616.
617.

618.
619.

620.

621.

622.

623.
624.

203

the assembly of focal adhesions and actin stress fibers in response

to growth factors. Cell 70: 389 399, 1992.
RIDLEY AJ AND HALL A. Signal transduction pathways regulating
Rho-mediated stress fibre formation: requirement for a tyrosine
kinase. EMBO J 13: 2600 2610, 1994.
RIDLEY AJ, PATERSON HF, JOHNSTON CL, DIEKMANN D, AND HALL A. The
small GTP-binding protein rac regulates growth factor-induced
membrane ruffling. Cell 70: 401 410, 1992.
RIDLEY AJ, SELF AJ, KASMI F, PATERSON HF, HALL A, MARSHALL CJ, AND
ELLIS C. Rho family GTPase activating proteins p190, bcr and
rhoGAP show distinct specificities in vitro and in vivo. EMBO J 12:
51515160, 1993.
RIEDERER MA, SOLDATI T, SHAPIRO AD, LIN J, AND PFEFFER SR. Lysosome biogenesis requires Rab9 function and receptor recycling
from endosomes to the trans-Golgi network. J Cell Biol 125: 573
582, 1994.
ROBINSON LC, GIBBS JB, MARSHALL MS, SIGAL IS, AND TATCHELL K.
CDC25: a component of the RAS-adenylate cyclase pathway in
Saccharomyces cerevisiae. Science 235: 1218 1221, 1987.
ROBINSON NG, GUO L, IMAI J, TOH-E A, MATSUI Y, AND TAMANOI F.
Rho3 of Saccharomyces cerevisiae, which regulates the actin cytoskeleton and exocytosis, is a GTPase which interacts with Myo2
and Exo70. Mol Cell Biol 19: 3580 3587, 1999.
RODENHUIS S, SLEBOS RJ, BOOT AJ, EVERS SG, MOOI WJ, WAGENAAR
SS, VAN BODEGOM PC, AND BOS JL. Incidence and possible clinical
significance of K-ras oncogene activation in adenocarcinoma of the
human lung. Cancer Res 48: 5738 5741, 1988.
RODRIGUEZ-VICIANA P, WARNE PH, DHAND R, VANHAESEBROECK B, GOUT
I, FRY MJ, WATERFIELD MD, AND DOWNWARD J. Phosphatidylinositol3-OH kinase as a direct target of Ras. Nature 370: 527532, 1994.
ROGGE RD, KARLOVICH CA, AND BANERJEE U. Genetic dissection of a
neurodevelopmental pathway: son of sevenless functions downstream of the sevenless and EGF receptor tyrosine kinases. Cell 64:
39 48, 1991.
ROHATGI R, MA L, MIKI H, LOPEZ M, KIRCHHAUSEN T, TAKENAWA T, AND
KIRSCHNER MW. The interaction between N-WASP and the Arp2/3
complex links Cdc42-dependent signals to actin assembly. Cell 97:
221231, 1999.
ROSARIO M, PATERSON HF, AND MARSHALL CJ. Activation of the Raf/
MAP kinase cascade by the Ras-related protein TC21 is required for
the TC21-mediated transformation of NIH 3T3 cells. EMBO J 18:
1270 1279, 1999.
ROTH MG. Lipid regulators of membrane traffic through the Golgi
complex. Trends Cell Biol 9: 174 179, 1999.
ROTH MG. Snapshots of ARF1: implications for mechanisms of
activation and inactivation. Cell 97: 149 152, 1999.
ROTH MG. Arf. In: GTPases, edited by Hall A. Oxford, UK: Oxford
Univ. Press, 2000, p. 176 197.
ROTHMAN JE. Mechanisms of intracellular protein transport. Nature
372: 55 63, 1994.
ROTHMAN JE AND WIELAND FT. Protein sorting by transport vesicles.
Science 272: 227234, 1996.
ROZAKIS-ADCOCK M, FERNLEY R, WADE J, PAWSON T, AND BOWTELL D.
The SH2 and SH3 domains of mammalian Grb2 couple the EGF
receptor to the Ras activator mSos1. Nature 363: 83 85, 1993.
RUSH MG, DRIVAS G, AND DEUSTACHIO P. The small nuclear GTPase
Ran: how much does it run? Bioessays 18: 103112, 1996.
RYBIN V, ULLRICH O, RUBINO M, ALEXANDROV K, SIMON I, SEABRA MC,
GOODY R, AND ZERIAL M. GTPase activity of Rab5 acts as a timer for
endocytic membrane fusion. Nature 383: 266 269, 1996.
SACHER M, JIANG Y, BARROWMAN J, SCARPA A, BURSTON J, ZHANG L,
SCHIELTZ D, YATES J III, ABELIOVICH H, AND FERRO-NOVICK S. TRAPP,
a highly conserved novel complex on the cis-Golgi that mediates
vesicle docking and fusion. EMBO J 17: 2494 2503, 1998.
SAEZ R, CHAN AM, MIKI T, AND AARONSON SA. Oncogenic activation of
human R-ras by point mutations analogous to those of prototype
H-ras oncogenes. Oncogene 9: 29772982, 1994.
SAITOH H, SPARROW DB, SHIOMI T, PU RT, NISHIMOTO T, MOHUN TJ, AND
DASSO M. Ubc9p and the conjugation of SUMO-1 to RanGAP1 and
RanBP2. Curr Biol 8: 121124, 1998.
SALMINEN A AND NOVICK PJ. A ras-like protein is required for a
post-Golgi event in yeast secretion. Cell 49: 527538, 1987.
SANDER EE, TEN KLOOSTER JP, VAN DELFT S, VAN DER KAMMEN RA, AND

204

625.

626.

627.

628.

629.

630.

631.

632.

633.

634.
635.

636.
637.

638.

639.

640.
641.

642.

643.

644.

645.

TAKAI, SASAKI, AND MATOZAKI

COLLARD JG. Rac downregulates Rho activity: reciprocal balance
between both GTPases determines cellular morphology and migratory behavior. J Cell Biol 147: 1009 1022, 1999.
SANO K, KIKUCHI A, MATSUI Y, TERANISHI Y, AND TAKAI Y. Tissuespecific expression of a novel GTP-binding protein (smg p25A)
mRNA and its increase by nerve growth factor and cyclic AMP in
rat pheochromocytoma PC-12 cells. Biochem Biophys Res Commun 158: 377385, 1989.
SANTOS E, TRONICK SR, AARONSON SA, PULCIANI S, AND BARBACID M.
T24 human bladder carcinoma oncogene is an activated form of the
normal human homologue of BALB- and Harvey-MSV transforming
genes. Nature 298: 343347, 1982.
SASAKI T, KAIBUCHI K, KABCENELL AK, NOVICK PJ, AND TAKAI Y. A
mammalian inhibitory GDP/GTP exchange protein (GDP dissociation inhibitor) for smg p25A is active on the yeast SEC4 protein.
Mol Cell Biol 11: 2909 2912, 1991.
SASAKI T, KATO M, AND TAKAI Y. Consequences of weak interaction of
rho GDI with the GTP-bound forms of rho p21 and rac p21. J Biol
Chem 268: 23959 23963, 1993.
SASAKI T, KIKUCHI A, ARAKI S, HATA Y, ISOMURA M, KURODA S, AND
TAKAI Y. Purification and characterization from bovine brain cytosol of a protein that inhibits the dissociation of GDP from and the
subsequent binding of GTP to smg p25A, a ras p21-like GTP-binding
protein. J Biol Chem 265: 23332337, 1990.
SASAKI T AND TAKAI Y. The Rho small G protein family-Rho GDI
system as a temporal and spatial determinant for cytoskeletal
control. Biochem Biophys Res Commun 245: 641 645, 1998.
SATA M, DONALDSON JG, MOSS J, AND VAUGHAN M. Brefeldin Ainhibited guanine nucleotide-exchange activity of Sec7 domain
from yeast Sec7 with yeast and mammalian ADP ribosylation factors. Proc Natl Acad Sci USA 95: 4204 4208, 1998.
SATO K AND WICKNER W. Functional reconstitution of ypt7p GTPase
and a purified vacuole SNARE complex. Science 281: 700 702,
1998.
SCHALK I, ZENG K, WU SK, STURA EA, MATTESON J, HUANG M, TANDON
A, WILSON IA, AND BALCH WE. Structure and mutational analysis of
Rab GDP-dissociation inhibitor. Nature 381: 42 48, 1996.
SCHEKMAN R AND ORCI L. Coat proteins and vesicle budding. Science
271: 1526 1533, 1996.
SCHERLE P, BEHRENS T, AND STAUDT LM. Ly-GDI, a GDP-dissociation
inhibitor of the RhoA GTP-binding protein, is expressed preferentially in lymphocytes. Proc Natl Acad Sci USA 90: 7568 7572, 1993.
SCHIMMOLLER F, SIMON I, AND PFEFFER SR. Rab GTPases, directors of
vesicle docking. J Biol Chem 273: 2216122164, 1998.
SCHLENSTEDT G, WONG DH, KOEPP DM, AND SILVER PA. Mutants in a
yeast Ran binding protein are defective in nuclear transport. EMBO
J 14: 53675378, 1995.
SCHLUTER OM, SCHNELL E, VERHAGE M, TZONOPOULOS T, NICOLL RA,
JANZ R, MALENKA RC, GEPPERT M, AND SUDHOF TC. Rabphilin knockout mice reveal that rabphilin is not required for rab3 function in
regulating neurotransmitter release. J Neurosci 19: 5834 5846,
1999.
SCHMALZING G, RICHTER HP, HANSEN A, SCHWARZ W, JUST I, AND
AKTORIES K. Involvement of the GTP binding protein Rho in constitutive endocytosis in Xenopus laevis oocytes. J Cell Biol 130:
1319 1332, 1995.
SCHMID SL AND DAMKE H. Coated vesicles: a diversity of form and
function. FASEB J 9: 14451453, 1995.
SCHMITT HD, WAGNER P, PFAFF E, AND GALLWITZ D. The ras-related
YPT1 gene product in yeast: a GTP-binding protein that might be
involved in microtubule organization. Cell 47: 401 412, 1986.
SCHUEBEL KE, BUSTELO XR, NIELSEN DA, SONG BJ, BARBACID M,
GOLDMAN D, AND LEE IJ. Isolation and characterization of murine
vav2, a member of the vav family of proto-oncogenes. Oncogene 13:
363371, 1996.
SCHUEBEL KE, MOVILLA N, ROSA JL, AND BUSTELO XR. Phosphorylation-dependent and constitutive activation of Rho proteins by wildtype and oncogenic Vav-2. EMBO J 17: 6608 6621, 1998.
SCOLNICK EM, PAPAGEORGE AG, AND SHIH TY. Guanine nucleotidebinding activity as an assay for src protein of rat-derived murine
sarcoma viruses. Proc Natl Acad Sci USA 76: 53555359, 1979.
SEABRA MC, BROWN MS, SLAUGHTER CA, SUDHOF TC, AND GOLDSTEIN
JL. Purification of component A of Rab geranylgeranyl transferase:

646.

647.
648.

649.
650.

651.

652.

653.

654.

655.

656.

657.

658.

659.

660.

661.

662.

663.

664.

665.

666.

667.

Volume 81

possible identity with the choroideremia gene product. Cell 70:

1049 1057, 1992.
SEABRA MC, GOLDSTEIN JL, SUDHOF TC, AND BROWN MS. Rab geranylgeranyl transferase. A multisubunit enzyme that prenylates
GTP-binding proteins terminating in Cys-X-Cys or Cys-Cys. J Biol
Chem 267: 1449714503, 1992.
SEABRA MC, HO YK, AND ANANT JS. Deficient geranylgeranylation of
Ram/Rab27 in choroideremia. J Biol Chem 270: 24420 24427, 1995.
SEABRA MC, REISS Y, CASEY PJ, BROWN MS, AND GOLDSTEIN JL. Protein
farnesyltransferase and geranylgeranyltransferase share a common
subunit. Cell 65: 429 434, 1991.
SEGAL AW AND ABO A. The biochemical basis of the NADPH oxidase
of phagocytes. Trends Biochem Sci 18: 43 47, 1993.
SEGEV N, MULHOLLAND J, AND BOTSTEIN D. The yeast GTP-binding
YPT1 protein and a mammalian counterpart are associated with the
secretion machinery. Cell 52: 915924, 1988.
SEKINE A, FUJIWARA M, AND NARUMIYA S. Asparagine residue in the
rho gene product is the modification site for botulinum ADPribosyltransferase. J Biol Chem 264: 8602 8605, 1989.
SELLS MA, BARRATT JT, CAVISTON J, OTTILIE S, LEBERER E, AND
CHERNOFF J. Characterization of Pak2p, a pleckstrin homology domain-containing, p21-activated protein kinase from fission yeast.
J Biol Chem 273: 18490 18498, 1998.
SELLS MA, KNAUS UG, BAGRODIA S, AMBROSE DM, BOKOCH GM, AND
CHERNOFF J. Human p21-activated kinase (Pak1) regulates actin
organization in mammalian cells. Curr Biol 7: 202210, 1997.
SENBONMATSU T, SHIRATAKI H, JIN-NO Y, YAMAMOTO T, AND TAKAI Y.
Interaction of Rabphilin3 with synaptic vesicles through multiple
regions. Biochem Biophys Res Commun 228: 567572, 1996.
SERAFINI T, ORCI L, AMHERDT M, BRUNNER M, KAHN RA, AND ROTHMAN
JE. ADP-ribosylation factor is a subunit of the coat of Golgi-derived
COP-coated vesicles: a novel role for a GTP-binding protein. Cell
67: 239 253, 1991.
SETTLEMAN J, ALBRIGHT CF, FOSTER LC, AND WEINBERG RA. Association between GTPase activators for Rho and Ras families. Nature
359: 153154, 1992.
SETTLEMAN J, NARASIMHAN V, FOSTER LC, AND WEINBERG RA. Molecular cloning of cDNAs encoding the GAP-associated protein p190:
implications for a signaling pathway from ras to the nucleus. Cell
69: 539 549, 1992.
SEWELL JL AND KAHN RA. Sequences of the bovine and yeast ADPribosylation factor and comparison to other GTP-binding proteins.
Proc Natl Acad Sci USA 85: 4620 4624, 1988.
SHIH TY, PAPAGEORGE AG, STOKES PE, WEEKS MO, AND SCOLNICK EM.
Guanine nucleotide-binding and autophosphorylating activities associated with the p21src protein of Harvey murine sarcoma virus.
Nature 287: 686 691, 1980.
SHIH TY, WILLIAMS DR, WEEKS MO, MARYAK JM, VASS WC, AND
SCOLNICK EM. Comparison of the genomic organization of Kirsten
and Harvey sarcoma viruses. J Virol 27: 4555, 1978.
SHIMIZU K, GOLDFARB M, PERUCHO M, AND WIGLER M. Isolation and
preliminary characterization of the transforming gene of a human
neuroblastoma cell line. Proc Natl Acad Sci USA 80: 383387, 1983.
SHIRATAKI H, KAIBUCHI K, SAKODA T, KISHIDA S, YAMAGUCHI T, WADA K,
MIYAZAKI M, AND TAKAI Y. Rabphilin-3A, a putative target protein for
smg p25A/rab3A p25 small GTP-binding protein related to synaptotagmin. Mol Cell Biol 13: 20612068, 1993.
SHIRATAKI H, KAIBUCHI K, YAMAGUCHI T, WADA K, HORIUCHI H, AND
TAKAI Y. A possible target protein for smg-25A/rab3A small GTPbinding protein. J Biol Chem 267: 10946 10949, 1992.
SHIRATAKI H, YAMAMOTO T, HAGI S, MIURA H, OISHI H, JIN-NO Y,
SENBONMATSU T, AND TAKAI Y. Rabphilin-3A is associated with synaptic vesicles through a vesicle protein in a manner independent of
Rab3A. J Biol Chem 269: 3271732720, 1994.
SHISHEVA A, BUXTON J, AND CZECH MP. Differential intracellular
localizations of GDP dissociation inhibitor isoforms. Insulin-dependent redistribution of GDP dissociation inhibitor-2 in 3T3L1 adipocytes. J Biol Chem 269: 2386523868, 1994.
SHISHEVA A, SUDHOF TC, AND CZECH MP. Cloning, characterization,
and expression of a novel GDP dissociation inhibitor isoform from
skeletal muscle. Mol Cell Biol 14: 3459 3468, 1994.
SHOU C, FARNSWORTH CL, NEEL BG, AND FEIG LA. Molecular cloning

January 2001

668.

669.

670.

671.

672.

673.

674.

675.

676.

677.

678.

679.
680.

681.
682.
683.

684.

685.

686.

687.

688.
689.

SMALL G PROTEINS

of cDNAs encoding a guanine-nucleotide-releasing factor for Ras

p21. Nature 358: 351354, 1992.
SIMON JP, IVANOV IE, REN J, SHOPSIN B, HERSH D, TEMPST P, ERDJUMENT-BROMAGE H, LUI M, DE LEMOS-CHIARANDINI C, ROSENFELD M,
GRAVOTTA D, MORIMOTO T, ADESNIK M, AND SABATINI DD. Regulation
of post-Golgi vesicle production in an in vitro system. Cold Spring
Harb Symp Quant Biol 60: 179 195, 1995.
SIMON MA, BOWTELL DD, DODSON GS, LAVERTY TR, AND RUBIN GM.
Ras1 and a putative guanine nucleotide exchange factor perform
crucial steps in signaling by the sevenless protein tyrosine kinase.
Cell 67: 701716, 1991.
SIMONSEN A, LIPPE R, CHRISTOFORIDIS S, GAULLIER JM, BRECH A, CALLAGHAN J, TOH BH, MURPHY C, ZERIAL M, AND STENMARK H. EEA1 links
PI(3)K function to Rab5 regulation of endosome fusion. Nature
394: 494 498, 1998.
SMALL JV. Lamellipodia architecture: actin filament turnover and
the lateral flow of actin filaments during motility. Semin Cell Biol
5: 157163, 1994.
SMELAND TE, SEABRA MC, GOLDSTEIN JL, AND BROWN MS. Geranylgeranylated Rab proteins terminating in Cys-Ala-Cys, but not
Cys-Cys, are carboxyl-methylated by bovine brain membranes in
vitro. Proc Natl Acad Sci USA 91: 1071210716, 1994.
SOLDATI T, SHAPIRO AD, SVEJSTRUP AB, AND PFEFFER SR. Membrane
targeting of the small GTPase Rab9 is accompanied by nucleotide
exchange. Nature 369: 76 78, 1994.
SONG J, KHACHIKIAN Z, RADHAKRISHNA H, AND DONALDSON JG. Localization of endogenous ARF6 to sites of cortical actin rearrangement
and involvement of ARF6 in cell spreading. J Cell Sci 111: 2257
2267, 1998.
SONNICHSEN B, LOWE M, LEVINE T, JAMSA E, DIRAC-SVEJSTRUP B, AND
WARREN G. A role for giantin in docking COPI vesicles to Golgi
membranes. J Cell Biol 140: 10131021, 1998.
SPAARGAREN M AND BISCHOFF JR. Identification of the guanine nucleotide dissociation stimulator for Ral as a putative effector molecule of R-ras, H-ras, K-ras, and Rap. Proc Natl Acad Sci USA 91:
12609 12613, 1994.
SPANG A, MATSUOKA K, HAMAMOTO S, SCHEKMAN R, AND ORCI L.
Coatomer, Arf1p, and nucleotide are required to bud coat protein
complex I-coated vesicles from large synthetic liposomes. Proc
Natl Acad Sci USA 95: 11199 11204, 1998.
SPRINGER S AND SCHEKMAN R. Nucleation of COPII vesicular coat
complex by endoplasmic reticulum to Golgi vesicle SNAREs. Science 281: 698 700, 1998.
SPRINGER S, SPANG A, AND SCHEKMAN R. A primer on vesicle budding.
Cell 97: 145148, 1999.
SRIVASTAVA SK, WHEELOCK RH, AARONSON SA, AND EVA A. Identification of the protein encoded by the human diffuse B-cell lymphoma
(dbl) oncogene. Proc Natl Acad Sci USA 83: 8868 8872, 1986.
STACEY DW AND KUNG HF. Transformation of NIH 3T3 cells by
microinjection of Ha-ras p21 protein. Nature 310: 508 511, 1984.
STADE K, FORD CS, GUTHRIE C, AND WEIS K. Exportin 1 (Crm1p) is an
essential nuclear export factor. Cell 90: 10411050, 1997.
STAHL B, CHOU JH, LI C, SUDHOF TC, AND JAHN R. Rab3 reversibly
recruits rabphilin to synaptic vesicles by a mechanism analogous to
raf recruitment by ras. EMBO J 15: 1799 1809, 1996.
STEARNS T, WILLINGHAM MC, BOTSTEIN D, AND KAHN RA. ADP-ribosylation factor is functionally and physically associated with the
Golgi complex. Proc Natl Acad Sci USA 87: 1238 1242, 1990.
STENMARK H, VITALE G, ULLRICH O, AND ZERIAL M. Rabaptin-5 is a
direct effector of the small GTPase Rab5 in endocytic membrane
fusion. Cell 83: 423 432, 1995.
STOFFLER HE, HONNERT U, BAUER CA, HOFER D, SCHWARZ H, MULLER
RT, DRENCKHAHN D, AND BAHLER M. Targeting of the myosin-I myr 3
to intercellular adherens type junctions induced by dominant active Cdc42 in HeLa cells. J Cell Sci 111: 2779 2788, 1998.
STOKOE D, MACDONALD SG, CADWALLADER K, SYMONS M, AND HANCOCK
JF. Activation of Raf as a result of recruitment to the plasma
membrane. Science 264: 14631467, 1994.
STOSSEL TP. On the crawling of animal cells. Science 260: 1086
1094, 1993.
STOWERS L, YELON D, BERG LJ, AND CHANT J. Regulation of the
polarization of T cells toward antigen-presenting cells by Ras-

690.

691.

692.

693.

694.

695.

696.

697.
698.

699.

700.

701.
702.
703.

704.
705.

706.

707.

708.

709.

710.

711.

205

related GTPase CDC42. Proc Natl Acad Sci USA 92: 50275031,
1995.
STRAUS DB AND WEISS A. Genetic evidence for the involvement of
the lck tyrosine kinase in signal transduction through the T cell
antigen receptor. Cell 70: 585593, 1992.
STROM M, VOLLMER P, TAN TJ, AND GALLWITZ D. A yeast GTPaseactivating protein that interacts specifically with a member of the
Ypt/Rab family. Nature 361: 736 739, 1993.
SULCINER DJ, IRANI K, YU ZX, FERRANS VJ, GOLDSCHMIDT-CLERMONT P,
AND FINKEL T. Rac1 regulates a cytokine-stimulated, redox-dependent pathway necessary for NF-kappaB activation. Mol Cell Biol 16:
71157121, 1996.
SUMI T, MATSUMOTO K, TAKAI Y, AND NAKAMURA T. Cofilin phosphorylation and actin cytoskeletal dynamics regulated by rho- and
Cdc42-activated LIM-kinase 2. J Cell Biol 147: 1519 1532, 1999.
SUNDARAM M AND HAN M. The C. elegans ksr-1 gene encodes a novel
Raf-related kinase involved in Ras-mediated signal transduction.
Cell 83: 889 901, 1995.
SUZUKI J, KAZIRO Y, AND KOIDE H. An activated mutant of R-Ras
inhibits cell death caused by cytokine deprivation in BaF3 cells in
the presence of IGF-I. Oncogene 15: 1689 1697, 1997.
SUZUKI T, MIKI H, TAKENAWA T, AND SASAKAWA C. Neural WiskottAldrich syndrome protein is implicated in the actin-based motility
of Shigella flexneri. EMBO J 17: 27672776, 1998.
SVITKINA TM AND BORISY GG. Progress in protrusion: the tell-tale
scar. Trends Biochem Sci 24: 432 436, 1999.
SYMONS M, DERRY JM, KARLAK B, JIANG S, LEMAHIEU V, MCCORMICK F,
FRANCKE U, AND ABO A. Wiskott-Aldrich syndrome protein, a novel
effector for the GTPase CDC42Hs, is implicated in actin polymerization. Cell 84: 723734, 1996.
TAKAHASHI K, SASAKI T, MAMMOTO A, HOTTA I, TAKAISHI K, IMAMURA H,
NAKANO K, KODAMA A, AND TAKAI Y. Interaction of radixin with Rho
small G protein GDP/GTP exchange protein Dbl. Oncogene 16:
3279 3284, 1998.
TAKAHASHI K, SASAKI T, MAMMOTO A, TAKAISHI K, KAMEYAMA T, TSUKITA
S, AND TAKAI Y. Direct interaction of the Rho GDP dissociation
inhibitor with ezrin/radixin/moesin initiates the activation of the
Rho small G protein. J Biol Chem 272: 2337123375, 1997.
TAKAI Y, KAIBUCHI K, KIKUCHI A, AND KAWATA M. Small GTP-binding
proteins. Int Rev Cytol 133: 187230, 1992.
TAKAI Y, KAIBUCHI K, KIKUCHI A, SASAKI T, AND SHIRATAKI H. Regulators of small GTPases. Ciba Found Symp 176: 128 138, 1993.
TAKAI Y, SASAKI T, SHIRATAKI H, AND NAKANISHI H. Rab3A small
GTP-binding protein in Ca2-dependent exocytosis. Genes Cells 1:
615 632, 1996.
TAKAI Y, SASAKI T, TANAKA K, AND NAKANISHI H. Rho as a regulator of
the cytoskeleton. Trends Biochem Sci 20: 227231, 1995.
TAKAISHI K, KIKUCHI A, KURODA S, KOTANI K, SASAKI T, AND TAKAI Y.
Involvement of rho p21 and its inhibitory GDP/GTP exchange
protein (rho GDI) in cell motility. Mol Cell Biol 13: 7279, 1993.
TAKAISHI K, SASAKI T, KATO M, YAMOCHI W, KURODA S, NAKAMURA T,
TAKEICHI M, AND TAKAI Y. Involvement of Rho p21 small GTP-binding
protein and its regulator in the HGF-induced cell motility. Oncogene 9: 273279, 1994.
TAKAISHI K, SASAKI T, KOTANI H, NISHIOKA H, AND TAKAI Y. Regulation
of cell-cell adhesion by rac and rho small G proteins in MDCK cells.
J Cell Biol 139: 10471059, 1997.
TAKAKURA A, MIYOSHI J, ISHIZAKI H, TANAKA M, TOGAWA A, NISHIZAWA
Y, YOSHIDA H, NISHIKAWA S, AND TAKAI Y. Involvement of a small
GTP-binding protein (G protein) regulator, small G protein GDP
dissociation stimulator, in antiapoptotic cell survival signaling. Mol
Biol Cell 11: 18751886, 2000.
TAN EC, LEUNG T, MANSER E, AND LIM L. The human active breakpoint cluster region-related gene encodes a brain protein with
homology to guanine nucleotide exchange proteins and GTPaseactivating proteins. J Biol Chem 268: 2729127298, 1993.
TANAKA K, IMAJOH-OHMI S, SAWADA T, SHIRAI R, HASHIMOTO Y, IWASAKI
S, KAIBUCHI K, KANAHO Y, SHIRAI T, TERADA Y, KIMURA K, NAGATA S,
AND FUKUI Y. A target of phosphatidylinositol 3,4,5-trisphosphate
with a zinc finger motif similar to that of the ADP-ribosylationfactor GTPase-activating protein and two pleckstrin homology domains. Eur J Biochem 245: 512519, 1997.
TANAKA K, MATSUMOTO K, AND TOH-E A. IRA1, an inhibitory regulator

206

712.

713.

714.

715.

716.

717.

718.

719.
720.

721.

722.

723.

724.

725.

726.

727.

728.

729.

730.

731.

732.

TAKAI, SASAKI, AND MATOZAKI

of the RAS-cyclic AMP pathway in Saccharomyces cerevisiae. Mol
Cell Biol 9: 757768, 1989.
TANAKA K, NAKAFUKU M, TAMANOI F, KAZIRO Y, MATSUMOTO K, AND
TOH-E A. IRA2, a second gene of Saccharomyces cerevisiae that
encodes a protein with a domain homologous to mammalian ras
GTPase-activating protein. Mol Cell Biol 10: 4303 4313, 1990.
TANAKA K AND TAKAI Y. Control of reorganization of the actin
cytoskeleton by Rho family small GTP-binding proteins in yeast.
Curr Opin Cell Biol 10: 112116, 1998.
TANG BL, PETER F, KRIJNSE-LOCKER J, LOW SH, GRIFFITHS G, AND HONG
W. The mammalian homolog of yeast Sec13p is enriched in the
intermediate compartment and is essential for protein transport
from the endoplasmic reticulum to the Golgi apparatus. Mol Cell
Biol 17: 256 266, 1997.
TANI K, OYAMA Y, HATSUZAWA K, AND TAGAYA M. Hypothetical protein
KIAA0079 is a mammalian homologue of yeast Sec24p. FEBS Lett
447: 247250, 1999.
TAPON N, NAGATA K, LAMARCHE N, AND HALL A. A new rac target
POSH is an SH3-containing scaffold protein involved in the JNK
and NF-kappaB signalling pathways. EMBO J 17: 13951404, 1998.
THERRIEN M, CHANG HC, SOLOMON NM, KARIM FD, WASSARMAN DA,
AND RUBIN GM. KSR, a novel protein kinase required for RAS signal
transduction. Cell 83: 879 888, 1995.
THERRIEN M, MICHAUD NR, RUBIN GM, AND MORRISON DK. KSR modulates signal propagation within the MAPK cascade. Genes Dev 10:
2684 2695, 1996.
THERRIEN M, WONG AM, AND RUBIN GM. CNK, a RAF-binding multidomain protein required for RAS signaling. Cell 95: 343353, 1998.
TISDALE EJ, BOURNE JR, KHOSRAVI-FAR R, DER CJ, AND BALCH WE.
GTP-binding mutants of rab1 and rab2 are potent inhibitors of
vesicular transport from the endoplasmic reticulum to the Golgi
complex. J Cell Biol 119: 749 761, 1992.
TODA T, UNO I, ISHIKAWA T, POWERS S, KATAOKA T, BROEK D, CAMERON
S, BROACH J, MATSUMOTO K, AND WIGLER M. In yeast, RAS proteins
are controlling elements of adenylate cyclase. Cell 40: 2736, 1985.
TOGAWA A, MIYOSHI J, ISHIZAKI H, TANAKA M, TAKAKURA A, NISHIOKA H,
YOSHIDA H, DOI T, MIZOGUCHI A, MATSUURA N, NIHO Y, NISHIMUNE Y,
NISHIKAWA S, AND TAKAI Y. Progressive impairment of kidneys and
reproductive organs in mice lacking Rho GDI. Oncogene 18: 5373
5380, 1999.
TOGAWA A, MORINAGA N, OGASAWARA M, MOSS J, AND VAUGHAN M.
Purification and cloning of a brefeldin A-inhibited guanine nucleotide-exchange protein for ADP-ribosylation factors. J Biol Chem
274: 12308 12315, 1999.
TOLIAS KF, CANTLEY LC, AND CARPENTER CL. Rho family GTPases
bind to phosphoinositide kinases. J Biol Chem 270: 17656 17659,
1995.
TOLIAS KF, COUVILLON AD, CANTLEY LC, AND CARPENTER CL. Characterization of a Rac1- and RhoGDI-associated lipid kinase signaling
complex. Mol Cell Biol 18: 762770, 1998.
TRAGLIA HM, OCONNOR JP, TUNG KS, DALLABRIDA S, SHEN WC, AND
HOPPER AK. Nucleus-associated pools of Rna1p, the Saccharomyces cerevisiae Ran/TC4 GTPse activating protein involved in nucleus/cytosol transit. Proc Natl Acad Sci USA 93: 76677672, 1996.
TRAHEY M AND MCCORMICK F. A cytoplasmic protein stimulates
normal N-ras p21 GTPase, but does not affect oncogenic mutants.
Science 238: 542545, 1987.
TRAHEY M, WONG G, HALENBECK R, RUBINFELD B, MARTIN GA, LADNER
M, LONG CM, CROSIER WJ, WATT K, AND KOTHS K. Molecular cloning
of two types of GAP complementary DNA from human placenta.
Science 242: 16971700, 1988.
TSUKAMOTO N, HATTORI M, YANG H, BOS JL, AND MINATO N. Rap1
GTPase-activating protein SPA-1 negatively regulates cell adhesion.
J Biol Chem 274: 1846318469, 1999.
TSUKITA S, YONEMURA S, AND TSUKITA S. ERM proteins: head-to-tail
regulation of actin-plasma membrane interaction. Trends Biochem
Sci 22: 5358, 1997.
TURNER CE, BROWN MC, PERROTTA JA, RIEDY MC, NIKOLOPOULOS SN,
MCDONALD AR, BAGRODIA S, THOMAS S, AND LEVENTHAL PS. Paxillin
LD4 motif binds PAK and PIX through a novel 95-kD ankyrin
repeat, ARF-GAP protein: a role in cytoskeletal remodeling. J Cell
Biol 145: 851 863, 1999.
UEDA T, KIKUCHI A, OHGA N, YAMAMOTO J, AND TAKAI Y. Purification

733.

734.
735.

736.

737.

738.

739.

740.
741.

742.

743.

744.
745.

746.
747.

748.

749.

750.

751.

752.

753.

754.

Volume 81

and characterization from bovine brain cytosol of a novel regulatory protein inhibiting the dissociation of GDP from and the subsequent binding of GTP to rhoB p20, a ras p21-like GTP-binding
protein. J Biol Chem 265: 93739380, 1990.
UI M AND KATADA T. Bacterial toxins as probe for receptor-Gi
coupling. Adv Second Messenger Phosphoprotein Res 24: 63 69,
1990.
ULLMAN KS, POWERS MA, AND FORBES DJ. Nuclear export receptors:
from importin to exportin. Cell 90: 967970, 1997.
ULLRICH O, HORIUCHI H, BUCCI C, AND ZERIAL M. Membrane association of Rab5 mediated by GDP-dissociation inhibitor and accompanied by GDP/GTP exchange. Nature 368: 157160, 1994.
ULLRICH O, REINSCH S, URBE S, ZERIAL M, AND PARTON RG. Rab11
regulates recycling through the pericentriolar recycling endosome.
J Cell Biol 135: 913924, 1996.
ULLRICH O, STENMARK H, ALEXANDROV K, HUBER LA, KAIBUCHI K,
SASAKI T, TAKAI Y, AND ZERIAL M. Rab GDP dissociation inhibitor as
a general regulator for the membrane association of rab proteins.
J Biol Chem 268: 1814318150, 1993.
UMIKAWA M, OBAISHI H, NAKANISHI H, SATOH-HORIKAWA K, TAKAHASHI
K, HOTTA I, MATSUURA Y, AND TAKAI Y. Association of frabin with the
actin cytoskeleton is essential for microspike formation through
activation of Cdc42 small G protein. J Biol Chem 274: 2519725200,
1999.
UMIKAWA M, TANAKA K, KAMEI T, SHIMIZU K, IMAMURA H, SASAKI T, AND
TAKAI Y. Interaction of Rho1p target Bni1p with F-actin-binding
elongation factor 1: implication in Rho1p-regulated reorganization
of the actin cytoskeleton in Saccharomyces cerevisiae. Oncogene
16: 20112016, 1998.
UNGERMANN C, SATO K, AND WICKNER W. Defining the functions of
trans-SNARE pairs. Nature 396: 543548, 1998.
URANO T, EMKEY R, AND FEIG LA. Ral-GTPases mediate a distinct
downstream signaling pathway from Ras that facilitates cellular
transformation. EMBO J 15: 810 816, 1996.
VALENCIA A, CHARDIN P, WITTINGHOFER A, AND SANDER C. The ras
protein family: evolutionary tree and role of conserved amino
acids. Biochemistry 30: 4637 4648, 1991.
VAN AELST L, BARR M, MARCUS S, POLVERINO A, AND WIGLER M.
Complex formation between RAS and RAF and other protein kinases. Proc Natl Acad Sci USA 90: 6213 6217, 1993.
VAN AELST L AND DSOUZA-SCHOREY C. Rho GTPases and signaling
networks. Genes Dev 11: 22952322, 1997.
VAN AELST L, JONESON T, AND BAR-SAGI D. Identification of a novel
Rac1-interacting protein involved in membrane ruffling. EMBO J
15: 3778 3786, 1996.
VAN AELST L, WHITE MA, AND WIGLER MH. Ras partners. Cold Spring
Harbor Symp Quant Biol 59: 181186, 1994.
VAN DER SLUIJS P, HULL M, WEBSTER P, MALE P, GOUD B, AND MELLMAN
I. The small GTP-binding protein rab4 controls an early sorting
event on the endocytic pathway. Cell 70: 729 740, 1992.
VAN DER SLUIJS P, HULL M, ZAHRAOUI A, TAVITIAN A, GOUD B, AND
MELLMAN I. The small GTP-binding protein rab4 is associated with
early endosomes. Proc Natl Acad Sci USA 88: 6313 6317, 1991.
VANRHEENEN SM, CAO X, LUPASHIN VV, BARLOWE C, AND WATERS MG.
Sec35p, a novel peripheral membrane protein, is required for ER to
Golgi vesicle docking. J Cell Biol 141: 11071119, 1998.
VENKATESWARLU K AND CULLEN PJ. Signalling via ADP-ribosylation
factor 6 lies downstream of phosphatidylinositide 3-kinase. Biochem J 345: 719 724, 2000.
VENKATESWARLU K, OATEY PB, TAVARE JM, AND CULLEN PJ. Insulindependent translocation of ARNO to the plasma membrane of
adipocytes requires phosphatidylinositol 3-kinase. Curr Biol 8:
463 466, 1998.
VERNOS I, RAATS J, HIRANO T, HEASMAN J, KARSENTI E, AND WYLIE C.
Xklp1, a chromosomal Xenopus kinesin-like protein essential for
spindle organization and chromosome positioning. Cell 81: 117
127, 1995.
VITALE G, RYBIN V, CHRISTOFORIDIS S, THORNQVIST P, MCCAFFREY M,
STENMARK H, AND ZERIAL M. Distinct Rab-binding domains mediate
the interaction of Rabaptin-5 with GTP-bound Rab4 and Rab5.
EMBO J 17: 19411951, 1998.
VITALE N, MOSS J, AND VAUGHAN M. ARD1, a 64-kDa bifunctional
protein containing an 18-kDa GTP-binding ADP-ribosylation factor

January 2001

755.

756.

757.

758.
759.

760.

761.

762.

763.

764.

765.

766.

767.

768.

769.

770.

771.
772.

773.

774.
775.

776.

SMALL G PROTEINS

domain and a 46-kDa GTPase-activating domain. Proc Natl Acad

Sci USA 93: 19411944, 1996.
VOGEL US, DIXON RA, SCHABER MD, DIEHL RE, MARSHALL MS,
SCOLNICK EM, SIGAL IS, AND GIBBS JB. Cloning of bovine GAP and its
interaction with oncogenic ras p21. Nature 335: 90 93, 1988.
VOGELSTEIN B, FEARON ER, HAMILTON SR, KERN SE, PREISINGER AC,
LEPPERT M, NAKAMURA Y, WHITE R, SMITS AM, AND BOS JL. Genetic
alterations during colorectal-tumor development. N Engl J Med
319: 525532, 1988.
VOGLER O, KRUMMENERL P, SCHMIDT M, JAKOBS KH, AND VAN KOPPEN
CJ. RhoA-sensitive trafficking of muscarinic acetylcholine receptors. J Pharmacol Exp Ther 288: 36 42, 1999.
VOJTEK AB AND DER CJ. Increasing complexity of the Ras signaling
pathway. J Biol Chem 273: 1992519928, 1998.
VOJTEK AB, HOLLENBERG SM, AND COOPER JA. Mammalian Ras interacts directly with the serine/threonine kinase Raf. Cell 74: 205214,
1993.
VOLLMER P, WILL E, SCHEGLMANN D, STROM M, AND GALLWITZ D.
Primary structure and biochemical characterization of yeast GTPase-activating proteins with substrate preference for the transport
GTPase Ypt7p. Eur J Biochem 260: 284 290, 1999.
VOSSLER MR, YAO H, YORK RD, PAN MG, RIM CS, AND STORK PJ. cAMP
activates MAP kinase and Elk-1 through a B-Raf- and Rap1-dependent pathway. Cell 89: 73 82, 1997.
WADA M, NAKANISHI H, SATOH A, HIRANO H, OBAISHI H, MATSUURA Y,
AND TAKAI Y. Isolation and characterization of a GDP/GTP exchange
protein specific for the Rab3 subfamily small G proteins. J Biol
Chem 272: 38753878, 1997.
WALCH-SOLIMENA C, COLLINS RN, AND NOVICK PJ. Sec2p mediates
nucleotide exchange on Sec4p and is involved in polarized delivery
of post-Golgi vesicles. J Cell Biol 137: 14951509, 1997.
WALCZAK CE, VERNOS I, MITCHISON TJ, KARSENTI E, AND HEALD R. A
model for the proposed roles of different microtubule-based motor
proteins in establishing spindle bipolarity. Curr Biol 8: 903913,
1998.
WALWORTH NC, GOUD B, KABCENELL AK, AND NOVICK PJ. Mutational
analysis of SEC4 suggests a cyclical mechanism for the regulation
of vesicular traffic. EMBO J 8: 16851693, 1989.
WANG Y, OKAMOTO M, SCHMITZ F, HOFMANN K, AND SUDHOF TC. Rim is
a putative Rab3 effector in regulating synaptic-vesicle fusion. Nature 388: 593598, 1997.
WARNE PH, VICIANA PR, AND DOWNWARD J. Direct interaction of Ras
and the amino-terminal region of Raf-1 in vitro. Nature 364: 352
355, 1993.
WATANABE G, SAITO Y, MADAULE P, ISHIZAKI T, FUJISAWA K, MORII N,
MUKAI H, ONO Y, KAKIZUKA A, AND NARUMIYA S. Protein kinase N
(PKN) and PKN-related protein rhophilin as targets of small
GTPase Rho. Science 271: 645 648, 1996.
WATANABE N, KATO T, FUJITA A, ISHIZAKI T, AND NARUMIYA S. Cooperation between mDia1 and ROCK in Rho-induced actin reorganization. Nat Cell Biol 1: 136 143, 1999.
WATANABE N, MADAULE P, REID T, ISHIZAKI T, WATANABE G, KAKIZUKA
A, SAITO Y, NAKAO K, JOCKUSCH BM, AND NARUMIYA S. p140mDia, a
mammalian homolog of Drosophila diaphanous, is a target protein
for Rho small GTPase and is a ligand for profilin. EMBO J 16:
3044 3056, 1997.
WATERS MG AND PFEFFER SR. Membrane tethering in intracellular
transport. Curr Opin Cell Biol 11: 453 459, 1999.
WEBER E, JILLING T, AND KIRK KL. Distinct functional properties of
Rab3A and Rab3B in PC12 neuroendocrine cells. J Biol Chem 271:
6963 6971, 1996.
WEIGERT R, SILLETTA MG, SPANO S, TURACCHIO G, CERICOLA C, COLANZI
A, SENATORE S, MANCINI R, POLISHCHUK EV, SALMONA M, FACCHIANO F,
BURGER KN, MIRONOV A, LUINI A, AND CORDA D. CtBP/BARS induces
fission of Golgi membranes by acylating lysophosphatidic acid.
Nature 402: 429 433, 1999.
WEISS A AND LITTMAN DR. Signal transduction by lymphocyte antigen receptors. Cell 76: 263274, 1994.
WELCH MD, IWAMATSU A, AND MITCHISON TJ. Actin polymerization is
induced by Arp2/3 protein complex at the surface of Listeria
monocytogenes. Nature 385: 265269, 1997.
WESTWICK JK, LAMBERT QT, CLARK GJ, SYMONS M, VAN AELST L,
PESTELL RG, AND DER CJ. Rac regulation of transformation, gene

777.

778.

779.
780.
781.

782.

783.
784.

785.

786.

787.
788.

789.

790.

791.

792.

793.

794.

795.

796.

797.

207

expression, and actin organization by multiple, PAK-independent

pathways. Mol Cell Biol 17: 1324 1335, 1997.
WHITE J, JOHANNES L, MALLARD F, GIROD A, GRILL S, REINSCH S, KELLER
P, TZSCHASCHEL B, ECHARD A, GOUD B, AND STELZER EH. Rab6 coordinates a novel Golgi to ER retrograde transport pathway in live
cells. J Cell Biol 147: 743760, 1999.
WHITEHEAD I, KIRK H, AND KAY R. Retroviral transduction and oncogenic selection of a cDNA encoding Dbs, a homolog of the Dbl
guanine nucleotide exchange factor. Oncogene 10: 713721, 1995.
WHITMAN M AND MELTON DA. Involvement of p21ras in Xenopus
mesoderm induction. Nature 357: 252254, 1992.
WIELAND F AND HARTER C. Mechanisms of vesicle formation: insights
from the COP system. Curr Opin Cell Biol 11: 440 446, 1999.
WILDE A AND ZHENG Y. Stimulation of microtubule aster formation
and spindle assembly by the small GTPase Ran. Science 284: 1359
1362, 1999.
WINTER D, PODTELEJNIKOV AV, MANN M, AND LI R. The complex
containing actin-related proteins Arp2 and Arp3 is required for the
motility and integrity of yeast actin patches. Curr Biol 7: 519 529,
1997.
WOLTHUIS RM, ZWARTKRUIS F, MOEN TC, AND BOS JL. Ras-dependent
activation of the small GTPase Ral. Curr Biol 8: 471 474, 1998.
WU J, MATUNIS MJ, KRAEMER D, BLOBEL G, AND COUTAVAS E. Nup358,
a cytoplasmically exposed nucleoporin with peptide repeats, RanGTP binding sites, zinc fingers, a cyclophilin A homologous domain, and a leucine-rich region. J Biol Chem 270: 14209 14213,
1995.
XIONG ZL, KITAMURA K, AND KURIYAMA H. ATP activates cationic
currents and modulates the calcium current through GTP-binding
protein in rabbit portal vein. J Physiol (Lond) 440: 143165, 1991.
XU GF, LIN B, TANAKA K, DUNN D, WOOD D, GESTELAND R, WHITE R,
WEISS R, AND TAMANOI F. The catalytic domain of the neurofibromatosis type 1 gene product stimulates ras GTPase and complements ira mutants of S. cerevisiae. Cell 63: 835 841, 1990.
XUE F AND COOLEY L. Kelch encodes a component of intercellular
bridges in Drosophila egg chambers. Cell 72: 681 693, 1993.
YAKU H, SASAKI T, AND TAKAI Y. The Dbl oncogene product as a
GDP/GTP exchange protein for the Rho family: its properties compared with those of Smg GDS. Biochem Biophys Res Commun 198:
811 817, 1994.
YAMADA M, TACHIBANA T, IMAMOTO N, AND YONEDA Y. Nuclear transport factor p10/NTF2 functions as a Ran-GDP dissociation inhibitor
(Ran-GDI). Curr Biol 8: 1339 1342, 1998.
YAMAGATA K, SANDERS LK, KAUFMANN WE, YEE W, BARNES CA,
NATHANS D, AND WORLEY PF. Rheb, a growth factor- and synaptic
activity-regulated gene, encodes a novel Ras-related protein. J Biol
Chem 269: 1633316339, 1994.
YAMAGUCHI A, URANO T, GOI T, AND FEIG LA. An Eps homology (EH)
domain protein that binds to the Ral-GTPase target, RalBP1. J Biol
Chem 272: 31230 31234, 1997.
YAMAGUCHI T, SHIRATAKI H, KISHIDA S, MIYAZAKI M, NISHIKAWA J, WADA
K, NUMATA S, KAIBUCHI K, AND TAKAI Y. Two functionally different
domains of rabphilin-3A, Rab3A p25/smg p25A-binding and phospholipid- and Ca2-binding domains. J Biol Chem 268: 27164
27170, 1993.
YAMAMOTO K, KONDO J, HISHIDA T, TERANISHI Y, AND TAKAI Y. Purification and characterization of a GTP-binding protein with a molecular weight of 20,000 in bovine brain membranes. Identification as
the rho gene product. J Biol Chem 263: 9926 9932, 1988.
YAMAMOTO M, MARUI N, SAKAI T, MORII N, KOZAKI S, IKAI K, IMAMURA
S, AND NARUMIYA S. ADP-ribosylation of the rhoA gene product by
botulinum C3 exoenzyme causes Swiss 3T3 cells to accumulate in
the G1 phase of the cell cycle. Oncogene 8: 1449 1455, 1993.
YAMAMOTO T, KAIBUCHI K, MIZUNO T, HIROYOSHI M, SHIRATAKI H, AND
TAKAI Y. Purification and characterization from bovine brain cytosol of proteins that regulate the GDP/GTP exchange reaction of
smg p21s, ras p21-like GTP-binding proteins. J Biol Chem 265:
16626 16634, 1990.
YAMANASHI Y AND BALTIMORE D. Identification of the Abl- and rasGAP-associated 62 kDa protein as a docking protein, Dok. Cell 88:
205211, 1997.
YAMASHITA T, YAMAMOTO K, KIKUCHI A, KAWATA M, KONDO J, HISHIDA T,
TERANISHI Y, SHIKU H, AND TAKAI Y. Purification and characterization

208

TAKAI, SASAKI, AND MATOZAKI

of c-Ki-ras p21 from bovine brain crude membranes. J Biol Chem

263: 1718117188, 1988.
798. YAMOCHI W, TANAKA K, NONAKA H, MAEDA A, MUSHA T, AND TAKAI Y.
Growth site localization of Rho1 small GTP-binding protein and its
involvement in bud formation in Saccharomyces cerevisiae. J Cell
Biol 125: 10771093, 1994.
799. YANG B, GONZALEZ L JR, PREKERIS R, STEEGMAIER M, ADVANI RJ, AND
SCHELLER RH. SNARE interactions are not selective. Implications
for membrane fusion specificity. J Biol Chem 274: 5649 5653, 1999.
800. YANG C, SLEPNEV VI, AND GOUD B. Rab proteins form in vivo complexes with two isoforms of the GDP-dissociation inhibitor protein
(GDI). J. Biol Chem 269: 3189131899, 1994.
801. YANG CH AND SNYDER M. The nuclear-mitotic apparatus protein is
important in the establishment and maintenance of the bipolar
mitotic spindle apparatus. Mol Biol Cell 3: 1259 1267, 1992.
802. YAO I, HATA Y, IDE N, HIRAO K, DEGUCHI M, NISHIOKA H, MIZOGUCHI A,
AND TAKAI Y. MAGUIN, a novel neuronal membrane-associated
guanylate kinase-interacting protein. J Biol Chem 274:
11889 11896, 1999.
803. YAO I, OHTSUKA T, KAWABE H, MATSUURA Y, TAKAI Y, AND HATA Y.
Association of membrane-associated guanylate kinase-interacting
protein-1 with Raf-1. Biochem Biophys Res Commun 270: 538 542,
2000.
804. YATANI A, OKABE K, POLAKIS P, HALENBECK R, MCCORMICK F, AND
BROWN AM. ras p21 and GAP inhibit coupling of muscarinic receptors to atrial K channels. Cell 61: 769 776, 1990.
805. YOKOYAMA N, HAYASHI N, SEKI T, PANTE N, OHBA T, NISHII K, KUMA K,
HAYASHIDA T, MIYATA T, AND AEBI U. A giant nucleopore protein that
binds Ran/TC4. Nature 376: 184 188, 1995.
806. YORK RD, YAO H, DILLON T, ELLIG CL, ECKERT SP, MCCLESKEY EW,
AND STORK PJ. Rap1 mediates sustained MAP kinase activation
induced by nerve growth factor. Nature 392: 622 626, 1998.
807. YOSHIDA Y, KAWATA M, MIURA Y, MUSHA T, SASAKI T, KIKUCHI A, AND
TAKAI Y. Microinjection of smg/rap1/Krev-1 p21 into Swiss 3T3 cells
induces DNA synthesis and morphological changes. Mol Cell Biol
12: 34073414, 1992.
808. YOSHIHISA T, BARLOWE C, AND SCHEKMAN R. Requirement for a GTPase-activating protein in vesicle budding from the endoplasmic
reticulum. Science 259: 1466 1468, 1993.
809. ZALCMAN G, CLOSSON V, CAMONIS J, HONORE N, ROUSSEAU-MERCK MF,
TAVITIAN A, AND OLOFSSON B. RhoGDI-3 is a new GDP dissociation
inhibitor (GDI). Identification of a non-cytosolic GDI protein interacting with the small GTP-binding proteins RhoB and RhoG. J Biol
Chem 271: 30366 30374, 1996.
810. ZENG J, REN M, GRAVOTTA D, DE LEMOS-CHIARANDINI C, LUI M, ERDJUMENT-BROMAGE H, TEMPST P, XU G, SHEN TH, MORIMOTO T, ADESNIK
M, AND SABATINI DD. Identification of a putative effector protein for
rab11 that participates in transferrin recycling. Proc Natl Acad Sci
USA 96: 2840 2845, 1999.
810a.ZERIAL M AND HUBER LA. Guidebook to the Small GTPases. Oxford,
UK: Oxford Univ. Press, 1995.
811. ZHANG FL AND CASEY PJ. Protein prenylation: molecular mechanisms and functional consequences. Annu Rev Biochem 65: 241
269, 1996.
812. ZHANG J, KING WG, DILLON S, HALL A, FEIG L, AND RITTENHOUSE SE.
Activation of platelet phosphatidylinositide 3-kinase requires the

813.

814.

815.

816.
817.

818.

819.

820.

821.

822.

823.

824.

825.

826.
827.

828.

Volume 81

small GTP-binding protein Rho. J Biol Chem 268: 2225122254,

1993.
ZHANG Q, CALAFAT J, JANSSEN H, AND GREENBERG S. ARF6 is required
for growth factor- and rac-mediated membrane ruffling in macrophages at a stage distal to rac membrane targeting. Mol Cell Biol 19:
8158 8168, 1999.
ZHANG Q, COX D, TSENG CC, DONALDSON JG, AND GREENBERG S. A
requirement for ARF6 in Fc receptor-mediated phagocytosis in
macrophages. J Biol Chem 273: 1997719981, 1998.
ZHANG XF, SETTLEMAN J, KYRIAKIS JM, TAKEUCHI-SUZUKI E, ELLEDGE
SJ, MARSHALL MS, BRUDER JT, RAPP UR, AND AVRUCH J. Normal and
oncogenic p21ras proteins bind to the amino-terminal regulatory
domain of c-Raf-1. Nature 364: 308 313, 1993.
ZHANG Z, VUORI K, WANG H, REED JC, AND RUOSLAHTI E. Integrin
activation by R-ras. Cell 85: 61 69, 1996.
ZHAO L, HELMS JB, BRUGGER B, HARTER C, MARTOGLIO B, GRAF R,
BRUNNER J, AND WIELAND FT. Direct and GTP-dependent interaction
of ADP ribosylation factor 1 with coatomer subunit . Proc Natl
Acad Sci USA 94: 4418 4423, 1997.
ZHAO L, HELMS JB, BRUNNER J, AND WIELAND FT. GTP-dependent
binding of ADP-ribosylation factor to coatomer in close proximity
to the binding site for dilysine retrieval motifs and p23. J Biol Chem
274: 14198 14203, 1999.
ZHAO ZS, LEUNG T, MANSER E, AND LIM L. Pheromone signalling in
Saccharomyces cerevisiae requires the small GTP-binding protein
Cdc42p and its activator CDC24. Mol Cell Biol 15: 5246 5257, 1995.
ZHENG Y, BAGRODIA S, AND CERIONE RA. Activation of phosphoinositide 3-kinase activity by Cdc42Hs binding to p85. J Biol Chem 269:
1872718730, 1994.
ZHENG Y, BENDER A, AND CERIONE RA. Interactions among proteins
involved in bud-site selection and bud-site assembly in Saccharomyces cerevisiae. J Biol Chem 270: 626 630, 1995.
ZHENG Y, FISCHER DJ, SANTOS MF, TIGYI G, PASTERIS NG, GORSKI JL,
AND XU Y. The faciogenital dysplasia gene product FGD1 functions
as a Cdc42Hs-specific guanine-nucleotide exchange factor. J Biol
Chem 271: 33169 33172, 1996.
ZHENG Y, OLSON MF, HALL A, CERIONE RA, AND TOKSOZ D. Direct
involvement of the small GTP-binding protein Rho in lbc oncogene
function. J Biol Chem 270: 90319034, 1995.
ZHENG Y, ZANGRILLI D, CERIONE RA, AND EVA A. The pleckstrin
homology domain mediates transformation by oncogenic dbl
through specific intracellular targeting. J Biol Chem 271: 19017
19020, 1996.
ZHU Y, DRAKE MT, AND KORNFELD S. ADP-ribosylation factor 1 dependent clathrin-coat assembly on synthetic liposomes. Proc Natl
Acad Sci USA 96: 50135018, 1999.
ZIGMOND SH. Signal transduction and actin filament organization.
Curr Opin Cell Biol 8: 66 73, 1996.
ZIMAN M, OBRIEN JM, OUELLETTE LA, CHURCH WR, AND JOHNSON DI.
Mutational analysis of CDC42Sc, a Saccharomyces cerevisiae gene
that encodes a putative GTP-binding protein involved in the control
of cell polarity. Mol Cell Biol 11: 35373544, 1991.
ZWARTKRUIS FJ, WOLTHUIS RM, NABBEN NM, FRANKE B, AND BOS JL.
Extracellular signal-regulated activation of Rap1 fails to interfere in
Ras effector signalling. EMBO J 17: 59055912, 1998.