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cytoskeleto

n
• The cytoskeleton is unique to eukaryotic cells. It
is a dynamic three-dimensional structure that
fills the cytoplasm. This structure acts as both
muscle and skeleton, for movement and
stability. The long fibers of the cytoskeleton are
polymers of subunits. The primary types of
fibers comprising the cytoskeleton are
microfilaments, microtubules, and intermediate
filaments.
The cytoskeleton is that the maintenance of
structure by the cytosol arises from a complex
network of protein filaments that traverse the
cell cytoplasm .
mitochondria

microtubule
cytoskeleton

ribosome
Intermediate
filament

microfilament ER
brief introduction of cytoskeleton
The cytoskeleton is a dynamic structure that is
responsible for whole –cell movement, changes
in cell shape, contraction of muscle cells, and
provides the machinery to move organelles
from one place to another in the cytoplasm. In
addition, cytoskeleton is the master organizer
of the cell’s cytoplasm, furnishing binding
sites for the specific localization of RNAs and
proteins that were once thought to diffuse
freely through the cytoplasm.
Φ 15nm

Φ 24-26nm

Part one 1
2
3

microtubule
13 4

12 5 5-9nm

Microtubule are hollow cylinders with 11


7
6

A diameter of 24nm. When viewed in 10


9 8

Cross section, the wall of each microtubule


Is seen to be composed of 13 tubulin
α α
dimers, which represent 13 protofilaments β β
composed of tubulin subunits.
αβ αβ
α -tubulin αβ α
heterodimer protofilaments β β
α α
β -tubulin End to end
( 13 β
) α
β
α
β
α
β
α
β
The constitutes of microtubules
he process of Microtubules assembly

α αβ αβ
β αβ α
β α β α β β
β α α
α α β α
α α α β
β α β
α β α β α
β α β β
β β α β
α β
β α
β
α
β

α - and β -tubulin subunits alternate


along the length of the protofilament,
which provides a microtubule with an
inherent polarity. 13 protofilments form
microtubule.
Microtubules undergo rapid
assembly and disassembly

α α
Plus end + β β α α
α α β α
β α βα βα β
α
β β α βα βα α αα β
α α
α α β βα βα β β β α α β β
β α β α α α β
β αβ β β α α ββ α
α α α α
α α β βα α β β αβ
α α β α β
α α β β
β α βα βββ βαα αβ α α α β

Tread-milling model
This polarity occurs β β
β
α β α β
α α β βαα α β β α β
because of the orientation β α ββ βα α α ββ αα
α β α
α
β αα β β α α β β
β α β
of the tubulin subunits in α α β βα α β β αα αα β α β
the microtubule polymer. α ββ α α ββ β
β β α αββ ββα ααβα β
β α
α β α
β βα α β β α αα β α β
α α ββ β
β ββ βα α α ββ α α β α
α
α α β β β αα β β α β
β
Minus end ββ β α α ββ α β α
- β β α αβ α
α β
β β α
β
both processes always occur more rapidly β α
at one end, called the plus end. The β
α
other, less active, end is the minus end. β
assembly
Cytoplasmic microtubules are labile structures that
have the capacity to undergo rapid assembly and
disassembly. This characteristic is important for
many microtubule functions. Such as cell mitosis,
cell migration and establishment of cellular
polarity.
Growing microtubules have an inherent structural
polarity. This polarity occurs because of the
orientation of the tubulin subunits in the
microtubule Polymer.

disassembly
tubulin MAP1A
1.Alkaline
typeI MAPs microtubule
MAP1B binding domain
Microtubule-
associated MAP2
2.Acidic
protein
typeII MAPs MAP4 protruding
(MAPs )
domain
Microtubine polymerized
protein ( Tua )

Microtubule-
associated
protein, MAP
During microtubule assembly, tubulin also
contains regions for GTP binding, for
interacting with microtubule-associated
proteins, and for several different drug-
binding sites, Such as colchicine, vinblastine
and taxol, which play an important
modulation role in assembly and
disassembly of microtubule.
The existing form of microtubule

1
2
3 A B
13 4

12 5

11 6
7
10
9 8

doublet
singlet

A B
C

triplet
The existing form of microtubule
Singlet is one of the most familiar forms in
cytoplasm. Singlet is often affected by the
temperature, colchicum and so on.
Doublet is composed of the flagellum and
cilia. A and B microtubule built up doublet,
which is composed of 23 protofilaments.
Triplet is often viewed in centrosome, which
is composed of 33 Protofilaments .
The function of
microtubule
1. Microtubules are involved in intracellular
vesicle and organelle transport.
A good analogy for visualizing this event
would be to consider a railroad: the
microtubules would serve as the tracks and
the locomotory forces responsible for
transporting the vesicular cargo would be
generated by ATPases, such as kinesin and
dynein.
An example of the role of microtubules
in transporting organelles
The function of
microtubule
2. Microtubules are composed of cilia and
flagella that extend from the surfaces of several
different cell types.
Flagella and cilia
Cilia are prominent in the respiratory tract and on the apical
surface of the epithelial cells that line oviduct. The major
type of flagellated cell in humans is the spermatozoon. The
beating flagellum provides the force that allows the sperm to
swim.
shaft : thin protrusion extending from
the cell surface. shaft



整 Basal body : cylindrical structure on the cilia
体 basal plasma membrane.

rootlet : microfilaments coming from the Basal body


basal body, concentrating as an taper on
its tip.
rootlet
doublet Central sheath

Central microtubule

shaft
B A
A B Out arm
B A dynein
Inner arm
B
A A
B C1 C2
axoneme spoke
B
轴丝 A
A Spoke head
B
A B A B Nexin
Connect filament

9X2+2
B
A
When viewed in cross section , microtubules are
arranged in a distinctive nine-plus-two array.




体 (9X3+
0)

Cross line: ATPase ,fasten cilia and take part in contraction.


横纹
The main proteins of flagella and cilia

tubulin :分子量约 55-60KD, 构成二联体、三联体以及中央微管的蛋白


质, 二联体微管中的微管蛋白缺乏秋水仙素 (colchicine) 的结合部
位,一般为鸟苷二磷酸( GDP )所占据。

dynein :构成二联体 A 微管内、外臂的主要成分,高分子量( 300-



要 400KD )
的 ATP 酶,可使化学能转变为机械能,使纤毛或鞭毛运动
蛋 AT
白 。 P

nexin :位于二联体微管的连桥及中央微管横桥之中,分子量 150-160K


功能:稳定两个相邻二联体微管间的滑动。放射幅 、中
央鞘也是由蛋白质构成。
The motility mechanism of cilia and flagella
In the presence of ATP, dynein undergoes a
conformational change. The net effect of this change is
that the dynein arm releases from the B tubule of the
adjacent microtubule doublet pair and then reattaches
to that same doublet pair farther down the length of the
doublet. This cycle is repeated when another ATP binds
to the dynein arm.

B A
B A B A
B A B A
头部

头部
The sliding-microtubule mechanism
of ciliary and flagella motility
the structure of
sperm flagellum
Central pair

Outer doublet

Dynein

Bridge

Spoke
centrosome
and centriole

Protoplasm
centrosphere ( transpar 原生质

centrosome ency cytosol )


nucleus
centriole

Φ 0.16-
电镜: column body 0.26um

圆柱状小体 — 长度 0.16-5.6um
centriole diplosome
• located in the cytoplasm attached to the outside
of the nucleus.
• Just before mitosis, the centrosome duplicates.
• The two centrosomes move apart until they are on
opposite sides of the nucleus.
• As mitosis proceeds, microtubules grow out from
each centrosome with their plus ends growing
toward the metaphase plate. These clusters of
microtubules are called spindle fibers.

The Centrosome
protoplasm

nucleus

centriole
centrosphere
C
C B C
B
A A B
C B A A
C
A B
A
B A
Centriolar A B
C B AB C
satellite C C

centriolar annulet
( 9X3+0)
The structure of centrosome in electron microscope
The centrosome is composed of a centriole pair and a
surrounding cloud of amorphous substance called the
pericentriolar material (PCM). Viewed in the transverse
section, There are 9 bundles of triplet tilting just like the
wings of windmill. Between the bundles, there are many
compacting granulars.
The ultrastructure of centrosome
The function of centrosome
and centriole
As the cell enter interphase, the centrosome
is duplicated, and at the onset of mitosis,
the daughter centrosomes migrate to
opposite sides of the nucleus where they
will serve as the mitotic spindle poles.
Along with the migration, microtubule
complex is disassembled and replaced by
the mitotic apparatus.
The origin of centriole

S phase
nucleus nucleus mitosis
• Microtubules attached to opposite sides of the dyad shrink or
grow until they are of equal length.
• Microtubules motors attached to the kinetochores move them
• toward the minus end of shrinking microtubules (a dynein);
• toward the plus end of lengthening microtubules (a kinesin).
• The chromosome arms use a different kinesin to move to the
metaphase plate
Part two
microfilaments
All eukaryotic cells appear to contain filaments 8
nm in diameter, called microfilaments, that are
polymers of actins.
G-actin ( globular actin monomers ) α -actin
actin β -actin
γ -actin
F-actin ( polymerization of G-actin)
-

tread milling model


Assembly of microfilments

G-actin

+
F-actin
In physiological conditions, actin monomers (G-
actin) spontaneously self associate to form
microfilaments (F-actin). This polymerised form
of actin is in constant equilibrium with G-actin.
Monomers are able to add to the ends of
filaments, form nuclei with another two
monomers to create new filaments, and to leave
filaments. The polarity of actin subunits within
microfilaments means that the two ends are
topologically different, the narrow face of actin
subunits is exposed at the pointed end while the
more bulbous face containing the NH2 and
COOH termini is exposed at the barbed end.
Pure actin can be switched between these states
in the test-tube by altering the salt concentration.
Actin in low salt conditions is in the G-state,
while adding salts causes the actin to polymerise.
These filaments , referred to as filamentous or
F-actin, are built from polymerization of a
globular actin monomer, called G-actin, which
has a relative molecular mass of 43kDa. Each
F-actin microfilament appears as two helically
intertwined chains of G-actin monomers, for
which a complete turn of the helix occurs over
a distance of 36 nm or 13 G-actin monomers.
subunits eclipse each other at what appears to
be a crossover.

microfilments
Microfilament-associated protein

Protein function
Tropomyosin stabilizes filaments
Fimbrin bundles filaments
Myosin II slides filaments in muscle
Profilin binds actin monomers
Cap Z caps plus-ends of filaments
The function of
microfilament
Microfilaments are responsible for
contraction in nonmuscle cells. For
example, during telophase, the last
stage in mitosis, a contractile ring
forms on the cytoplasmic membrane
surface.
The cytoplasm has regions that have
the characteristics of a Pseudoplastic
gel and other regions that liquify into
the sol state.
Gel-sol conversion within the cytoplasm
is regulated by the dynamic state of
actin and its interaction with actin-
binding proteins.
Part three
intermediate
filaments
Intermediate
filaments are 10
nm in diameter .
Therefore,
Intermediate in
thickness is
between
microfilaments and
microtubules.
The structure of intermediate
filaments
Basic unit of intermediate filament——monomers
α -helix: 310 个氨基酸残基 (I-IV 型和 VI
型 IF) 或 356 个氨基酸残基 (V 型 IF)
中等纤维单体共同结构域
组成。
head ( N- 端)
Non-helix
tail ( C- 端)
head tail
( C- 端)
( N- 端) α -helix

L12 L2
L1
连 连 连

N- 1A

区 1B

区 2A

区 2B C-
α - 螺旋 α - 螺旋 α - 螺旋 α - 螺旋
端 35 101 9 121

8-14 8 16-17
The structure of intermediate filaments
All IF proteins contains a subunit-spacific
NH2-terminus of variable size, a homologous
central a-helical region of approximately
310 amino acids, and a subunit-specific
COOH-terminus of variable size. Only the
homologous 310-amino acid a-helical region is
a portion of the 10-nm IF core. The variable
regions extend from the core and are
responsible for cross-linking intermediate
filaments to other cytoskeletal structures.
Assembly of intermediate filaments
COOH 单体
NH2
mononor
COOH
NH2 二聚体
COOH
NH2 超螺旋
dimer
COOH
NH2
COOH
NH2
COOH NH2
四聚体
COOH NH2
tetramer

原丝
原丝

八聚体
原纤维
八聚体

中等纤维
中等纤维
Assembly of intermediate filaments
In the formation of the intermediate filaments,
the first step is that the 310-amino acid a-helical
region of two monomers wind around each other
into a parallel coiled-coil. The IF proteins
contain the heptad repeat within the 310-amino
acid a-helical region required for coiled-coil
formation. Next two dimers link side-by-side in
an antiparallel conformation to form a tetramer.
The IF tetramers attach laterally to each other in
a staggered array, until there are eight tetramers (
32 monomers) making up the wall of IF. The
eight tetramers are wound to form the ropelike
structure of IF.
Part three intermediate filaments

The type of intermediate filaments


Type Name distribution
I keratin filament epithelium
II desmin filament muscle cells
III vimentin filament fibroblasts
IV neuroglial filament neurons
V neurofilament neurons
The character of intermediate filaments

1. The IF in various human and animal cells


are composed of a heterogeneous group of
proteins.
2. No energy in the form of ATP or GTP
hydrolysis is required for intermediate
filament polymerization.
3. Intermediate filaments have no polarity
because the tetramers have an antiparallel
conformation.
The function of intermediate filaments

1. Supporting effect, especially supporting


nucleus.
2. Transporting function, associated with
microtubule and microfilament.
3. IF affects the transporting of mRNA.
4. IF participate in the signal transducing
as a kind of information molecule.
The camparison of cytoskeleton
Microfilaments are twisted double strands consisting of a
string of proteins, from 7 nm to several cm long. The
protein is actin. Its function helps muscle contraction, cell
shape, and movement in cytoplasm.
Intermediate filamentsare made of eight subunits in
rope-strands. The proteins structure varies with different
tissue types. This component helps maintain shape,
support nerve cell extentions, and attach cells together.
Microtubules are tubes made up of spiraling, two-part
subunits. It is made of tubulin. It aids in chromosome
movement, movement of organelles, and the movement of
cilia and flagella.
   
微管 (mocrotubule)

微丝 (microfilament)

中等纤维
(intermediate filament)
The camparison of cytoskeleton
microtubule microfilament Intermediate
filament
component tubulin actin Intermediate
filament monomer

22nm 7nm 10nm


diametr
hollow tube two strands helix rope-strands helix
structure
yes yes no
polarity
yes yes no

treadmill
action
Microfilaments and Microtubules are similar in many
respects, they are polar, dynamic structures whose
assembly state is nucleotide dependent and both interact
with a host of associated proteins . IFs are apolar and
rather more static polymers which are depolymerised by
phosphorylation. The three systems differ in their
mechanical properties; MFs form visco-elastic gels; MTs
resist bending and compression; IFs are extremely tough
fibres which resist stretching. MFs are arranged as gels or
bundles in association with a large number of actin
binding proteins. MTs are usually single typically have
their minus end associated with an MTOC deep within the
cell, with the plus end toward the periphery. IFs connect
cell-cell junctions to give strength to tissues. All three
systems are interconnected to various extents.
Key point
• The structure 、 component 、
assemble process of microtubule 、
intermediate filament and microfilament.
• The structure of cilia 、 flagella and
centrosome
• The function of cytoskeleton
1. Please take the glucose as an example,
explain how the glucose is decomposed
into CO2 and H2O, and how ATP are
produced.
2. Talk about the process that proteins are
synthesized and transported.
3. Comparing the difference in
microtubule, microfilament, and
intermediate filament.
Explaining terms

1. Respiratory chain

2. Signal peptide

3. The cytoskeleton