Bio 150 Eukaryotic Cell Structure and Function ASSEMBLY (polymerization) - addition of monomers/subunits

Miriam De Vera, PhD at the end of polymer

Lecture 3.1 Cytoskeleton o kon rate constant (M-1 sec-1)
DISASSEMBLY (depolymerization) - removal of
Cytoskeleton monomers/subunits at the end of polymer
A network of protein fibers extended throughout the o koff rate constant (sec-1)
cytoplasm Whether assembly or disassembly is promoted depends on the
Dynamic, capable of relatively rapid assembly and characteristic of the cell environment.
disassembly o So it depends on nature of free energy available to the
o Unlike the animal exoskeleton or endoskeleton, which system usually in the form of ATP or ADP
are rigid in order to support the muscles they dont
change unless they undergo osteoclastic activity Notes from ppt:
This continuous assembly & disassembly of the cytoskeleton A linear polymer of protein molecules assembles
utilizes energy (polymerizes) and disassembles (depolymerizes) by the
addition and removal of subunits at the ends of the polymer.
Figure: The rate of addition of these subunits (called monomers) is
*High resolution light microscope given by the rate constant kon, which has units of M-1 sec-1.
image of a cell The rate of loss is given by koff (units of sec-1).
*Stained with Coomassie Blue
*Network of cytoskeletal elements and Thermal stability of cytoskeletal filaments with dynamic ends:
the nucleus are visible Single Protofilament: Thermally UNSTABLE

FUNCTIONS of the cytoskeleton:

1. Structural support for maintaining cell shape
Examples:
Neurons star-like shape maintained by microtubules
Spectrin this protein, in combination with actin, Most cytoskeletal elements are filamentous.
contributes to the flexibility of the membrane of RBCs Protofilament (single protofilament)
2. Positioning of various organelles in particular locations in the o Several monomers interconnected with one another
cell interior o Functional polymer of a cytoskeletal element
Epithelial cells of digestive tract: o Can be assembled together to form sheets there is
o Both apical & basal have a particular shape a high energy bond that can connect all of the
there is a gradient of the kind of membrane proteins monomers in 1 protofilament
anchored at the apical end & basal end o Encircled figure more likely to occur than the figure on
o This anchorage is provided by cytoskeletal the left
elements o Its easier to have disassembly at the ends/terminal
(right figure) than somewhere in the middle (left figure)
3. Machinery for the movement of materials and organelles
within a cell Single protofilament thermally unstable; tendency is to
Formation of transport vesicles bind one another to become more stable
o When transport vesicles move, they are assisted by
microtubules Multiple Protofilaments: Thermally STABLE
Separation of chromosomes during anaphase
o Powered by microtubules
4. Force-generating elements allowing movement of the cells
from one place to another
Ameoboid movement

Cytoskeletal elements have dynamic property (capable of assembly

& disassembly ~unlike skeletal bones)
Figure:
*Cytoskeletal elements = polymers
(proteins)
*1 box = monomer Multiple protofilaments more thermally stable
*Down arrow assembly o Figure: Red lines = noncovalent bonds
*Process is REVERSIBLE o Bonds are at sides and downward. Everything is bound
depending on the constants together
(constant for assembly or vs. in single protofilament: bonds are only at the
constant for rxn of disassembly sides
Right: The addition or loss of a subunit at the end of one
protofilament makes or breaks one set of longitudinal bonds

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 and either one or two sets of lateral bonds. This is more
possible to occur.
Left: Breakage of the composite filament in the middle requires
breaking sets of longitudinal bonds in several protofilaments
all at the same time

Caption from Alberts: Formation of a cytoskeletal filament from more

than one protofilament allows the ends to be dynamic, while the
filaments themselves are resistant to thermal breakage. In this
hypothetical example, the stable filament is formed from 5
protofilaments. The bonds holding the subunits together in the filaments
are shown in red.

Think: Cooking Spaghetti

Its concept applies to single and multiple protofilaments
1 kg pasta = too long so you need to break it
Kaya mong putulin yung isang kilo ng spaghetti pasta???
Kaya niyo yun? (De Vera, 2017) 2. Microfilaments
o Demonstrates that if they are greater in number (like Like 2 strings of beads (actin); smaller in diameter
mutiple protofilaments = sheets), they are more stable. o Not cylindrical since it has no lumen
Therefore, it is more difficult to break them apart Actin filaments most common type of microfilament in
o For both situations, disassembly is more likely to occur eukaryotic cells
for monomers at the terminals Polymer: F-actin (double-stranded filament)
Monomer: G-actin (globular)
Three Major Cytoskeletal Elements Some functions:
1. Microtubules o Changes in cell shape during growth
Cylindrical: w/ lumen o Cell motility in psudopodia only
Polymers: tubulin polymers
Subunits: tubulin dimers
o Wall consists of tubulin
dimers
o Dimer because of its alpha
and beta subunits
Fluorescent micrograph: Green
= microtubules in fibroblast cells
Relatively large

Ribbon model of G-actin

Theres a domain within the g-actin for the binding of a
nucleotide: ATP (ADP when in filaments)
Plus end = C-terminus; Minus end = N-terminus

Ribbon model of the dimer (next figure)

With alpha and beta subunits
Red: either a GDP or GTP molecule (so obviously a nucleotide
is needed for assembling)
Alternating dark and light green in 1 protofilament:
o Dark green = beta subunit
o Light green = alpha subunit
Plus and minus ends of tubulin protofilament
o Plus end = beta; where addition usually occurs
o Minus end = alpha; where removal usually occurs

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 3. Intermediate Filaments (IF)
Like a rope (thick and big)
Fluorescent micrograph: Green = IF in neurons
Main protein: keratin
o Function: tension-bearing
Keratin is the most stable among the 3 major cytoskeletal
elements
In order to build up 1 keratin functional structural unit, you
have to have at least a tetramer (monomer is not enough)
IF subunits do not contain a binding site for nucleotides (like
GTP, GDP, ADP, ATP)
A Model of Intermediate Filament Construction
Globular domains are of variable size and sequence
(A) Monomer of keratin
string
N-terminus and C-terminus are shown
(B) Dimer
2 monomers in parallel orientation; same ends are together
(C with C; N with N)
(C) Tetramer
2 dimers in opposite orientation (N-C; C-N); 1 functional
structural unit for IF
The tetramer is the soluble subunit of IF.
Pairs of protofilaments associate laterally to form a protofibril
Within each tetramer, the 2 dimers are offset with respect to
one another, allowing to associate with another tetramer.
Sheet several tetramers
1 sheet of keratin rolled up to form a cable
At least 4 protofibrils will laterally form a single IF
IF assembly is not accompanied by nucleotide hydrolysis, but
phosphorylation of the head domain can cause
depolymerization
In the final 10 nm rope-like filament, tetramers are packed
together in a helical array, which has 16 dimers in cross-
section. Half of these dimers are pointing in each direction
Whole Process of IF Construction
There are no plus and minus ends in IF due to the antiparallel
orientation of tetramers.
Therefore, it is more stable than others. You cant just remove
or add to the keratin tetramer assemblage in IF
Tetramer ends will be the same after arranging the dimers in
antiparallel directions assembled IF lacks structural polarity
which is critical for actin and microtubules
Recap:
All of the 3 major cytoskeletal elements have their specific
monomeric units to form the polymer
Protofilament = Initial polymer
o Ex. Microtubule protofilament initial polymer of
tubulin dimer (1 string of tubulin dimer cannot
form a complete microtubule [MT] yet bc you need
several to form 1 MT)
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NUCLEATION B) Polymerization is
Describes how initial polymerization occurs begun in the same way,
The initial process of the assembly of monomeric units into but with preformed
a larger structure (polymer) fragments of actin
filaments present to act
as nuclei for filament
growth. As indicated,
the % free subunits
reflects the critical
concentration (Cc), the
A helical polymer is stabilized by multiple contacts between point at which there is
adjacent subunits. In the case of actin, 2 actin molecules bind no net change in
relatively weakly to each other, but addition of a 3rd actin polymer.
monomer to form a trimer makes the entire group more
stable No lag phase because it started with oligomers (unlike the
one in the first figure, it started with few subunits)
When you have the nucleation phase, rate is very slow. But
Actin Polymerization (Nucleation) in this case, since you already have the oligomers
o Rate of polymerization will increase exponentially
o Equilibrium will be reached at a faster rate too
A) Polymerization is
*Nucleation - true for actin and MT. Not much for keratin (IF).
begun by raising the
salt concentration in a
solution of pure actin
MICROTUBULES (MT)
subunits
*There are certain locations or sites for the formation/assembly of
the cytoskeletal elements:
Microtubules (MT) nucleate in a microtubule-organizing center
[MTOC] (eg. Centrosome).

*Cc = defined as the concentration level of free ATP-G-actin where

the rate of addition is balanced by the rate of loss and no net growth
occurs at the end
Actin microfilaments (MF): few actin subunits (monomer)
add more G monomeric units oligomeric structure actin
protofilaments
Default or natural state of typical cell:
o Low % of actin subunits slowly increasing (lag
phase) growth phase or elongation (increasing
slope for assembly/disassembly kinetics)
o In order to have growth, rate of polymerization
(addition) must be greater than depolymerization
(removal) [Kon and Koff]
o Removal at minus end of polymer; addition at plus
end
o Steady state (equilibrium phase) length of the
polymer has become relatively constant due to the
equilibration of the rate of depolymerization with Centrosomes
polymerization - Example of MTOC
Nucleation (lag phase) is a relatively slow process but will - Found in dividing animal cells; Not found in prokaryotes
eventually become exponentially faster in rate as more and plant cells
monomers are being assembled (see graph) - Composed of 2 MT:
Salt addition salt solution of pure G-actin monomers added o Mother or parent MT assembled first
to an active solution/mixture o Daughter MT formative; perpendicular to the
mother MT
Not all proteins associated with the MT can be seen
immediately
Associating proteins of MT: MAPs Microtubule Associating
(Accessory) Proteins
o There are different types of MAPs depending on the
variations of their amino acid sequences, as well as the
antibodies to which they react
MAP1, MAP2, MAP3, MAP4, etc.

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Pericentriolar Material
Newly formed MT radiate from the pericentriolar material with
the minus ends toward centrosome
One centrosome does not only contain 1 maternal and 1
daughter centriole. There are also surrounding proteinaceous
complexes associated with the centrosome called
PERICENTRIOLAR MATERIAL or CENTRIOLE MATRIX
MAPs can be found within the pericentriolar material
- Tubulin Ring Complex
First biochemically characterized proteins in the pericentriolar
material or centrosomal matrix
MT are nucleated from a complex of -tubulin and other
protein components known as the -tubulin ring complex (-
TuRC)
Complex = more than 1 protein present
& vs
o and tubulin subunits are part of the protofilament
basic structure
o -TuRC are important in the process of nucleation (MT
nucleation will not occur without -TuRC)
Figure: Terminal portion of MT
light green -tubulin
green -tubulin
blue -tubulin subunit
red = -TuRC (complex
+ accessory proteins)
Accessory proteins subunits
noncovalently attached to
gamma tubulin
Figure: Ribbon model of Tau
protein
Tau protein one of the
proteins identified as
part of -TuRC
Immunofluorescence
micrograph: shows the
distribution of tau staining
(green) and MAP2 staining
(orange) in a hippocampal
neuron in culture.
These protein complexes
associated with y-tubulin are
necessary for MT nucleation
(kaya nasa centrosome sila)
Tau protein and MAP2 main function: mechanical
stabilization of the nucleation site (for centriole elongation to
occur)
Centrosomes can also be found in basal bodies (cilia and flagella)
Basal bodies or
kinetosome serve as the
MTOC of cilia and flagella
In the MTOCs of MT, there
are MAPs which are
necessary for structural
stability (also true for the
microfilaments). However,
there are also motor
proteins for movement.
Motor proteins
- important in generating movement (unlike Tau and MAP2
which are for structural stability)
- Since cilia and flagella are for movement, you need to have
motor proteins
- include dynein, kinesin and more
o Dynein arms - associated with each pair of MT for
ciliary and flagellar movement
o Kinesin common in neuronal cells
Sliding Tubule Mechanism [Dynein]
- Made possible because there are dynein arms which walk
along the surface of the adjacent MT
- Bending of cilium and flagellum are made possible if there are
anchoring cross-link proteins
o They are steady thats why they are capable of
bending. But without this anchorage, the cilia/flagella
will only get longer
Dynein and kinesin are not the only motor proteins associated with
MT. There are 3 bases for the variation of motor proteins:
1. Motor proteins can either bind to the MT or MF
2. They also differ in the cargo associated with them (vesicles
or organelles or specific molecules)
3. Direction of movement (anterograde or retrograde)
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Recall: Basic structure of microtubule Motility of Kinesin

In this figure, the kinesin heads look like feet (blue and green)
Red and orange filament: 1 protofilament of microtubule
Kinesin heads often associate with the beta subunits of the
MT
Nucelotide binding domain (or binding site) of kinesin
Microtubule dimers themselves have their domains to which
o Located in the heads
the nucleotide GDP or GTP is attached
o Have hydrolytic activity. Thus, they can convert
In this case, the red one is GTP.
triphosphate into a diphosphate
These are not the same nucleotides associated with the
While the head is attached to the beta, it is available for
motor proteins.
binding with a new ATP
At the same time that the heads rotate, diphosphate is
Nucleotide Hydrolysis
released providing the momentum for rotation/movement
o Diphosphate = released
o Triphosphate = added
ATPase domain located in the kinesin heads; converts ATP
into its diphosphate form (ADP)
o This hydrolysis provides energy for the rotation and
eventual movement/walking along the protofilament.

Direction of Movement [Kinesin]

GIF: Kinesin
Cargo: axon vesicles
When you have a free monomer (newly synthesized and Anterograde (forward) = release of
freely soluble in the cytosol of a cell), the nucleotide typically neurotransmitters
attached to it is a triphosphate (GTP or ATP) Retrograde (backward) = resorption
When it is the triphosphate which is embedded in the subunit, of neurotransmitters
thats the one which will promote addition or polymerization
If that nucleotide is a diphosphate, it promotes disassembly
So if you want to have a polymerization of cytoskeletal elements, FACTORS FAVORING ASSEMBLY AND DISASSEMBLY IN MT
the nucleotide associated with the subunit must be a Assembly Disassembly
triphosphate. But pag matatanggal siya, it should be a High temperature (37oC for Low temperature (lower than
diphosphate kasi less stable. mammalian cells) 37C)
The nucleotides mentioned are within the subunit of the
Low (osmotic) pressure w/in the High pressure
cytoskeletal elements. They are different from the
cell
nucleotides that interact with the motor proteins.
GTP cap GDP cap
High concentration of free Low concentration of free
*Motor proteins (ex.kinesin) have their own domain for binding a
tubulin (when a free tubulin is tubulin (kulang tubulin
nucleotide.
newly synthesized, it is usually disassembly of monomers)
Kinesin bound to a triphosphate)
- are the typical motor proteins associated with the MT in nerve Taxol Colchicine
cells High Ca++ concentration
Diagrammatic model of kinesin: (it interferes with the
Kinesin heads N-terminus; these noncovalent association of
are the ones which interact with the dimer subunits)
microtubule as seen in the next figure

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Dynamic Instability (GTP vs GDP Cap)
Growing GTP cap = most of the added dimers have GTP
particularly in the beta subunits because beta is more subject
to hydrolysis/replacement into GDP
o Alpha subunit GTP is more embedded. It cannot be
easily removed or replaced by GDP
o In the system or in an isolated cell, accumulation of
dimers with GTP mostly associated with beta
subunit promotes growing
GDP = hydrolyzed form of GTP; promotes disassembly
Microtubules depolymerize about 100 times faster from an
end containing GDP tubulin than from one containing GTP
tubulin. A GTP cap favors growth, but if it is not lost, then
depolymerization ensues.
DynamIc Instability phenomenon wherein individual
microtubules can alternate between a period of slow growth
and period of rapid disassembly
Figure: MT in the process of assembly (plus end) and disassembly
(minus end)
Minus end (shrinking) = Light green = w/ GDP
shorter/smaller = means that
disassembly has slower rate
Plus end (growing) = longer filaments
(newly formed microtubules: each
one is a protofilament assembling
together)
Drugs or Chemicals that Promote Assembly and Disassembly
Taxol
o Family of drugs that influence microtubule elongation
o Promotes assembly or elongation
o Very useful anticancer drug. How?
Cancer cells are highly mitotic. If the MT are
intact, anaphase will not occur since the MT will
not shorten. They will only continue to elongate.
Colchicine
o Anticancer drug
o Promotes disassembly; MT destabilizer
o Kung di ka makabuo ng MT at all, di ka magkakaroon
ng sufficient redistribution of chromosomes in the
daughter cells
Dynamic instability due to the structural differences between a
growing and a shrinking MT end
A) Catastrophe and Rescue
Caption from Bruce Alberts: If the free tubulin concentration in solution
is between the critical values in Figure 16-14B (di naturo), a single
microtubule end may undergo transitions between a growing state and
a shrinking state. A growing MT has GTP-containing subunits at its end,
forming a GTP cap. If nucleotide hydrolysis proceeds more rapidly than
subunit addition, the cap is lost and the MT begins to shrink, an event
called a catastrophe. But GTP-containing subunits may still add to the
shrinking end, and if enough add to form a new cap, then MT growth
resumes, an event called rescue.
Dark green = w/ GTP, mostly in beta subunits
In the diagram, theres still a GTP cap. As long as it has a GTP
cap, it is safe because it wont disassemble, it still has the
potential to elongate
This process of addition and removal is continuous
dynamic
Catastrophe
o The rate of the accumulation of subunits (GDP) has
exceeded the addition of GTP cap
o Theres a particular phenomenon among MT and MF
that once they are part of a protofilament, their GTP
are easily replaced with GDP (the rates will vary
depending on the factors mentioned earlier) such
that kapag humabol yung formation ng subunits
upto the dulo, catastrophena siya (because wala na
yung GTP cap, madali magdisassemble or decay)
o When the rate of GTP hydrolysis has caught up to
the rate of the cap formation (theres more of the
GDP in the protofilament than the GTP cap more
prone to disassembly)
Rescue
o In cell cultures, if you can increase the concentration
of the dimers with GTP or decrease the amount of
calcium then you can easily hasten the addition of
GTP cap for it to exceed the hydrolysis of
triphosphate
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B) Model for the structural consequences of GTP hydrolysis in the In an intact microtubule, protofilaments made from GDP-
MT lattice containing subunits are forced into a linear conformation by
the many lateral bonds within the MT wall, given a stable cap
of GTP-containing subunits. Loss of the GTP cap, however,
allows the GDP-containing protofilaments to relax into their
more curved conformation. This leads to a progressive
disruption of the MT.
The electron micrographs on the right show actual MT in each
of these two states, as observed in preparations in vitreous
ice. Note particularly the curling, disintegrating GDP-
containing protofilaments at the end of the shrinking
microtubule.

*Since this demonstrates that there is continual assembly and

disassembly for MT, a phenomenon known as treadmilling can be
demonstrated

D) Treadmilling behavior of a microtubule (observed in a living cell)

From Bruce Alberts: The addition of GTP-containing tubulin subunits to

the end of a protofilament causes the end to grow in a linear From Bruce Alberts: The microtubule shown
conformation that can readily pack into the cylindrical wall of the appears to be sliding from left to right, but, in
microtubule. Hydrolysis of GTP after assembly changes the fact, the microtubule lattice remains
conformation of the subunits and tends to force the protofilament into a stationary (as shown by landmark within the
curved shape that is less able to pack into the MT wall. microtubule lattice indicated by the red
Figure: Visual demonstration of what happens when GTP is arrowhead, while the plus end (on the right)
being replaced by GDP grows and the minus end (On the left)
If you have a string of GTP-bearing beta subunits, it is rigid or shrinks. The plus end also displays dynamic
straight instability.
But if the GTP is hydrolyzed para siyang rod na maraming
segments na biglang humina as seen in the curved
protofilament (linear arrangement before) which suggests na
very fragile na yung noncovalent bonds between subunits
more prone to breakage
*Beta lang pinapakita not alpha since sa beta yung madaling palitan
Figure: Frame by frame from a video
o Green = fluorescent-labeled MT
C) Growing and Shrinking o Triangles = markers of position
o White line = another marker
As seen in the markers, it looks like its moving which is
actually the basis for the MT being able to move a cargo (like
vesicles) within the cell
How is this movement happening? At first, you can see that
it is short it then elongates which means that the rate of
assembly is becoming greater than the rate of disassembly
You do not only have addition, but you also have removal as
seen in its movement away from the white marker
treadmilling (whatever is being added in the plus ends is
compensated by the removal of subunits at the minus ends)
It results generally in: elongation and apparent movement
Explains whats going on when MT moves vesicles during
sorting and trafficking
True for both actin filaments and microtubules

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MICROFILAMENTS (MF)
Actin filaments
- Also known as microfilaments
- Two-stranded helical polymers of the protein actin
- Appear as flexible structures, with a diameter of 5-9 nm, and Myosin head in the
they are organized into a variety of linear bundles, 2D sarcomere
Secondary structure:
networks, and 3D gels. Chains in a helical
curled w/o tail
- Although actin filaments are dispersed throughout the cell, arrangement
No tail because of the
they are most highly concentrated in the cortex, just beneath Head region and tail
tendency to curl
the plasma membrane
One chain only curling
Function in transport of Function in muscle contraction
cytoplasmic vesicles (tension) and cytokinesis

Organization of Accessory Proteins in a Sarcomere

Shown in the figure:

Actin
Myosin
ABPs
o Titin - anchoring protein to the Z line for the myosin
o Nebulin - supporting protein associated with the actin
filaments
o Caps:
Figure: Diagrammatic representation of actin [Actin at the crossroads]
CapZ - capping protein associated with Z line
Actin binds to a very large variety of accessory proteins in all eukaryotic
Tropomodulin - cap towards the M line
cells. This diagram shows the most of the interactions that have been
demonstrated, using either genetic or biochemical techniques, in the
Not shown: regulatory proteins in the sarcomere (associated with
yeast Saccharomyces cerevisiae. Accessory proteins that operate in the
actin, troponin, and tropomyosin)
same intracellular process are shown in the same color, as indicated in
the key.
From Bruce Alberts:
Actin most common MF in cells
Each giant titin molecule extends from the Z disc to the M line---a
All of the different colored dots are the processes (involving
distance of over 1m.
actin) and associated proteins which are involved in specific
Part of each titin molecule is closely associated with a myosin
functions
thick filament (which switches polarity at the M line)
Actin at the center: portion of F actin
the rest of the titin molecule is elastic and changes length as the
The diagrammatic representation shows that there are
sarcomere contracts and relaxes
specific domains on the F actin that has a particular ABP
Each nebulin molecule is exactly the same as the length of a thin
(actin binding proteins)
filament.
Accessory proteins for actin: ABP equivalent of MAP
The actin filaments are also coated with tropomyosin and troponin
o Microtubule = MAPs
(not shown) and are capped at both ends.
o Actin MF = ABP
Tropomodulin caps the minus end of the actin filaments
CapZ anchors the plus end at the Z disc, which also contains -
Myosin (another MF or myofilament)
actinin
Myosin I and II first myosins discovered
Myosin I Myosin II
In nonmuscle cells In muscle and nonmuscle cells
(thats why they are more
common)
With 1 heavy chain but no tail With 2 heavy chains and a tail
region region

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GENERAL CLASSIFICATION OF ABPs b) Crosslinking proteins
Action of ABPs
Protein Action Examples
Monomer Sequesters G-actin and Thymosin
binding prevent polymerization Profilin
protein Promotes actin assembly by
adding monomers to plus end
and recharging ADP-G-actin
to ATP-G-actin
End-blocking Regulate length of MF by Cap-Z
(capping) binding to one end of the B-actinin ABPs which promote crosslinking
filament, preventing further How are crosslinking proteins associated with gelation and
disassembly or assembly solation?
Actin cross- Crosslink actin to form Filamin o Gelation crosslinkages; MF crosslink for the gel
linking networks Fimbrin state of the cytoplasm
Crosslink actin in parallel to Villin o Solation crosslinkages degraded/removed for
form bundles cytoplasm to become soluble (sol state)
Filament- Bind to interior of existing Gelsolin Importance of transition of gel state to sol state: amoeboid
severing filament, causing breaks and Severin movement
making the cytoplasm more Fragmin
fluid c) Dystrophin
Membrane Link actin to plasma Vinculin
MF-binding membrane through Dystrophin
proteins membrane proteins Spectrin

F-actin

Dystrophin ABP associated with sarcolemma (muscle cell

membrane)
Necessary to keep cell membrane intact
Figure: Example of F-actin undergoing depolymerization and
o Muscle cells contract and relax. They are capable of
polymerization (addition)
being stretched. During relaxation, when the muscle
Gray: associated nucleotide is ADP = depolymerization or
is more elongated, dystrophin prevents it from
removal of actin monomers
snapping
Yellow: associated nucleotide is ATP = polymerization or
o It stabilizes the structure of the sarcolemma
addition of actin monomers
There are many dystrophins. There are particular dystrophin
Profilin directs actin monomers to barbed ends
family of genes which code for the protein.
But if theres a mutation, one of the disorders resulting from
Visual examples of the different proteins:
the mutation of dystrophin gene is Duchenne Muscular
a) Capping Proteins Dystrophy (or atrophy) no muscle tone and the muscle
tissues are not strong enough
CapZ - stabilizing protein
which supports the d) Actin Related Proteins (ARP) Complexes
attachment of the actin to the
Z line Analogous to Y-TuRC
Cofilin - opposite function of Important for nucleation
CapZ; promotes disassembly of MF
of polymer

10
Other functions of actin and myosin
1. Maintenance of cell shape and structure
- Interaction of myosin with MF helps stiffen the plasma
membrane, reducing likelihood of surface deformation
- Figure: Artificial Human Stem Cells
o Typical shape of stem cells is round. But in this case,
the shapes are star-shaped and flower-shaped
o The researchers who made this culture of stem cells
added certain molecules for the actin filaments to
attach to them and form these specific shapes (kaya
artificially structured)
o These human stem cells have different shapes
because of the modifications done
2. Component of stress fibers
- MF and associated proteins beneath the plasma membrane
of focal contacts form stress fibers
o Recall: Focal adhesion (or contact) points = part of
intercellular junctions internally associated with MF
- These help maintain tension between the cell and
substratum of the ECM
3. Amoeboid movement
- Related to #2 (Stress fibers). You cannot generate stress
fibers without the actin or myosin filaments and these are
necessary for amoeboid movement
- Actin network in trailing edge contract to squeeze the sol
portion toward the leading edge
- Trailing edge of amoeboid cell = crosslinkages in the
ectoplasm; gelatin-like
- Leaving edge of pseudopodia = not much crosslinks
movable state of cytoplasm allows amoeboid movement
4. Intracellular translocation
- Formation of endocytotic vesicles: one way by which these
vesicles can be internalized into the cell is through the
involvement of actin filaments
o Cargo of actin filaments: vesicles
5. Structural support of microvilli
- Actin filaments: main cytoskeletal structures within the
microvilli
- These are responsible for the movement and elongation of
villi during digestive absorption
- Elongation occurs in some animal cells when a lot of food
materials are digested and absorbed through the lumen in
the digestive tract
- Elongation is made possible by the sliding of actin filaments
6. Cytokinesis in non-muscle cells
- During cell division, a contractile ring, comprised of non-
muscle myosin II and actin filaments is formed and placed
at the center of the cell
- This ring contracts using the
energy released from ATP
hydrolysis and physically cleaves
the cell into two
- Contractile ring becomes smaller
and smaller for the daughter cells
to separate [cytokinesis]
7. Cytoplasmic streaming
- Stationary bundles of actin lie just under the nonmoving
cortical cytoplasm [easily observed in plants Ex. Hydrilla]
- Cross-bridges are formed between myosin bound to the ER
and the stationary MF
- The binding and sliding of the ER along the filaments propel
the entire cytoplasm
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 Cytoplasmic Streaming

8. Changes in cell shape during development

- Explains the differentiation in the shape of epithelial cells in
the digestive tract (left figure)

o Cells in the apical region contract to Types of IFAPs

become columnar (found in the more
mature digestive epithelia)
o Imagine it as an apron wherein the
apical actin-filament bundles are
drawstrings na hinila para maging skirt-
like

- Also applicable in nerve cells:

IFAPs
o crosslink the IF with one another, forming a bundle or a
network, and with other cell structures, including the
- Growth cone functions to explore the environment and to plasma membrane
elongate the axon along an appropriate pathway Lamins
- Nerve cells elongate because of the formation of the growth o A, B, C differ based on aa sequence and antibody
cone wherein you have the extending filopodia reactions
- Axon = microtubules (dark orange) o associated with nuclear lamina, keeping it intact
- Lamellipodia (and filopodia) of growth cone = actin Vimentin-like
filaments (light orange) o first IFAPs found in mesenchyme cells
o For it to extend for neuronal growth Keratins
o for epithelial cells
INTERMEDIATE FILAMENTS (IF) o classified based on the amino acids that make them
- Smaller than MT but more compact since it has no lumen up:
- Has higher tensile strength acidic aa type 1
- Greater stability (not easily disassembled unlike MT & MF) basic aa type 2
- It is not involved in movement (depolymerization and o whether alpha or beta keratin:
polymerization not frequent) alpha = soft (hair, fur, epithelial cells)
- More on structural and anchorage beta = rigid (horns, hooves, carapace)
- Experiment: MT or MF of a cell culture denatured or Neurofilaments (NF)
degraded thru a chemical procedure but IF not denatured o found in the axons of neurons
cells still maintain their normal shape Different IFAPs have their tissue or cell-specificity such that
- Associating proteins: IFAP (IF Associating/Accessory one can distinguish cancerous from normal cells based on
Proteins) what kind of chemistry their IF have
- IF networks are not only easily deformed, but they withstand Remember: IF are not subject to high rates of disassembly
large stresses and strains without rupture; they are thereby stable (not involved in movement) bc theres no + or end in
well suited to maintain cell integrity its basic unit which is the tetramer

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Extensive cross-linking in the axon neurofilament provide great
tensile strength, while in glial cells, the IF have fewer cross bridges.

SEM of IF in an axon of a nerve cell:

a) Typical neuron

transmits nerve impulses

with crosslinkages (IFAPs) in the axon

b) Glial cell

does not transmit nerve impulses

less cross-linking IFAPs in the axon

Cross section of an axon

MT larger & fewer

NF IF; smaller but more numerous
since they are the ones which
provide mechanical stability to the
structure of the axon (more so than
the MT)

*Read the caption!

*blue = plasma membrane
*Desmosomes - Intercellular junctions associated with the IF for
these type of cells (squamous epithelial cells)

Come to me, all you who are weary and burdened,

and I will give you rest. Matthew 11:28

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