Cytoskeleton

The cytoskeleton is a network of fibers
that organizes structures and activities
in the cell
• The cytoskeleton is a network of fibers
extending throughout the cytoplasm
• It organizes the cell’s structures and activities,
anchoring many organelles
• It is composed of three types of molecular
structures:
– Microtubules
– Microfilaments
– Intermediate filaments
Copyright © 2008 Pearson Education, Inc., publishing as Pearson Benjamin Cummings

Fig. 6-20
Microtubule
0.25 μm Microfilaments

Roles of the Cytoskeleton:
Support, Motility, and Regulation
• The cytoskeleton helps to support the cell
and maintain its shape
• It interacts with motor proteins to produce
motility
• Inside the cell, vesicles can travel along
“monorails” provided by the cytoskeleton
• Recent evidence suggests that the
cytoskeleton may help regulate biochemical
activities

Fig. 6-21
Vesicle
ATP
Receptor for
motor protein
Microtubule
of cytoskeleton
Motor protein
(ATP powered)
(a)
Microtubule Vesicles
(b)
0.25 μm

Components of the Cytoskeleton
• Three main types of fibers make up the
cytoskeleton:
– Microtubules are the thickest of the three
components of the cytoskeleton
– Microfilaments, also called actin filaments, are
the thinnest components
– Intermediate filaments are fibers with diameters
in a middle range

Table 6-1
10 μm 10 μm 10 μm
Column of tubulin dimers
Tubulin dimer
Actin subunit
a b
25 nm
7 nm
Keratin proteins
Fibrous subunit (keratins
coiled together)
8–12 nm

Table 6-1a
10 μm
Column of tubulin
dimers
Tubulin
dimer
a b
25 nm

Table 6-1b
Actin subunit
10 μm
7 nm

Table 6-1c
5 μm
Keratin proteins
Fibrous subunit (keratins
coiled together)
8–12 nm

Cytoskeleton proteins revealed by Commassie staining
Cytoskeleton: filament system
Three types of filaments
and accessory proteins
(assembly of cytoskeleton,
motorproteins that move
organelles
or filaments)
Internal order
Shape and remodel surface
Move organelles
Movement
Cell division

Dynamic and adaptable
Actin filaments:
shape of the cell’s surface
and whole cell locomotion
5-9 nm diameter
Microtubules:
positions of membrane-enclosed
organelles,
intracellular transport
25 nm diameter
Intermediate filaments:
mechanical strength and
resistance to shear stress
10 nm diameter

Microtubules
• Microtubules are hollow rods about 25 nm
in diameter and about 200 nm to 25 microns
long
• Functions of microtubules:
– Shaping the cell
– Guiding movement of organelles
– Separating chromosomes during cell division

Centrosomes and Centrioles
• In many cells, microtubules grow out from a
centrosome near the nucleus
• The centrosome is a “microtubule-organizing
center”
• In animal cells, the centrosome has a pair of
centrioles, each with nine triplets of
microtubules arranged in a ring

Microtubules
• Microtubules
• I. Introduction: Long, hollow cylinders, 25 nm in diameter, made of
tubulin. The basic subunit is a heterodimer of α and β tubulin (9.8);
13 protofilaments in a typical cylinder. See below about GTP
binding, treadmilling, growth and dynamic instability (9.26). There is
a + end, fast growing, w/β tubulin at its end, and a – end, slow
growing, w/α tubulin at its end. The GTP’s are important in
assembly (9.8)
• A. They have MAPs, that influence their use- linking them together,
stabilizing them, or destabilizing them.
• B. They form a network, coming from the microtubule organizing
center, which is usually the centrosome or centriole, w/ the – end
anchored there. (9.10-13, 19)
• C. Also form cilia and flagella, and spindle fibers in mitosis.

• F. MICROTUBULE DYNAMICS: 9.25
• Key points- the cap means that subunits
are added easily- loss of GTP = harder to
add subunits, need higher subunit conc. to
add.
• Produces microtubule catastrophes!

Nucleation
• Gamma tubulin in MTOC/centriole- MT’s
grow from there

FIGURE 9.25 The structural cap model
of dynamic instability. According
to the model, the growth or shrinkage of a
microtubule depends
on the state of the tubulin dimers at the
plus end of the microtubule.
Tubulin-GTP dimers are depicted in red.
Tubulin-GDP dimers are depicted
in blue. In a growing microtubule (step 1),
the tip consists of an
open sheet containing tubulin-GTP
subunits. In step 2, the tube has begun
to close, forcing the hydrolysis of the bound
GTP. In step 3, the tube
has closed to its end, leaving only tubulin-
GDP subunits. GDP-tubulin
subunits have a curved conformation
compared to their GTP-bound
counterparts, which makes them less able
to fit into a straight protofilament.
The strain resulting from the presence of
GDP-tubulin subunits
at the plus end of the microtubule is
released as the protofilaments curl
outward from the tubule and undergo
catastrophic shrinkage (step 4).

Dynamic instability:predominant in microtubules
GTP hydrolysis “catch up”
Treadmilling: predominant in actin filaments

Lateral bonds force GDP-containing
protofilaments into a linear conformation

Remodeling
• The fact that MT’s aren’t fixed means that
cells can remodel their shape- plant cells,
our cells in mitosis- round up, as MT’s
used to make spindle fibers

The time course of actin polymerization in a test tube

The structure of a microtubule and its subunits
GTP!
GTP
hollow and cylindrica and polar
13 parallel protofilaments
heterodimer

The structure of an actin monomer and actin filament
monomer
ATP
polar
two parallel protofilaments
that twist around each other
in a right-handed helix
Flexible but cross-linked and
bundled together by accessory
proteins in a living cell

The preferential growth of microtubules at the plus end
Plus end: polymerize and
depolymerize faster than
minus end
Microtubules:
Plus end- b subunit
Minus end- a subunit
Actin filaments
Plus end- barbed end
Minus end- pointed end

Fig. 6-22 Centrosome
Microtubule
Centrioles
0.25 μm
Longitudinal section
of one centriole
Microtubules Cross section
of the other centriole

GTP hydrolysis causes
filament to curve

MT drugs
• Colchicine- prevents MT formation- arrests
cells at metaphase
• Useful in determining role of MT’s in a
process

Effect of the drug taxol on microtubule organization
treatment of cancers

Actin and tubulin are highly conserved: they have to bind to
many proteins directly and indirectly
Accessory proteins and intermediate filament proteins
are not as conserved

A model of intermediate filament construction
Intermediate filaments
are only found in some
metazoans:vertebrates,
nematodes,molluscs
Not required in
every cell type
Ancesters: nuclear lamins
Parallel
Antiparrel
No polarity! “subunit”
8 parallel protofilaments
Easily bent
Hard to break

Two types of intermdiate filaments in cells of the nervous system
Neurofilaments:axons
NF-L, NF-M, NF-H proteins coassemble
axon glia
NF-M and NF-H have long C-terminal tails
That bind to neighboring filaments:uniform spacing
Regular spacing
When axons grow, subunits are added at the filament ends
and along the filament length; axon diameter increase 5 fold
In ALS (Lou Gehrig’s Disease), there is an accumulation and abnormal assembly of
Neurofilaments in motor neuron cell bodies and axon--interfere with normal axon transport

Summary
1. Three types of cytoskeletal filaments, protofilaments;
2. Subunits, polymerization, treadmilling, dynamic
instability;
3. Intermediate filaments, cell integrity, diseases caused
by mutations in the intermediate filament genes
4. Natural toxins and cytoskeleton

Cilia and Flagella
• Microtubules control the beating of cilia and
flagella, locomotor appendages of some
cells
• Cilia and flagella differ in their beating
patterns
VViiddeeoo:: CChhllaammyyddoommoonnaass VViiddeeoo:: PPaarraammeecciiuumm CCiilliiaa

Fig. 6-23
5 μm
Direction of swimming
(a) Motion of flagella
Direction of organism’s movement
Power stroke Recovery stroke
(b) Motion of cilia
15 μm

• Cilia and flagella share a common
ultrastructure:
– A core of microtubules sheathed by the plasma
membrane
– A basal body that anchors the cilium or
flagellum
– A motor protein called dynein, which drives the
bending movements of a cilium or flagellum
Animation: CCiilliiaa aanndd FFllaaggeellllaa

Fig. 6-24
0.1 μm
Triplet
(c) Cross section of basal body
0.5 μm
Microtubules
Plasma
membrane
Basal body
(a) Longitudinal
section of cilium
(b) Cross section of
cilium
Plasma
membrane
Outer microtubule
doublet
Dynein proteins
Central
microtubule
Radial
spoke
Protein cross-linking
outer
doublets
0.1 μm

• How dynein “walking” moves flagella and
cilia:
− Dynein arms alternately grab, move, and
release the outer microtubules
– Protein cross-links limit sliding
– Forces exerted by dynein arms cause doublets
to curve, bending the cilium or flagellum

Intermediate Filaments
• Intermediate filaments range in diameter
from 8–12 nanometers, larger than
microfilaments but smaller than
microtubules
• They support cell shape and fix organelles
in place
• Intermediate filaments are more permanent
cytoskeleton fixtures than the other two
classes

Cell Walls of Plants
• The cell wall is an extracellular structure that
distinguishes plant cells from animal cells
• Prokaryotes, fungi, and some protists also
have cell walls
• The cell wall protects the plant cell, maintains
its shape, and prevents excessive uptake of
water
• Plant cell walls are made of cellulose fibers
embedded in other polysaccharides and
protein

• Plant cell walls may have multiple layers:
– Primary cell wall: relatively thin and flexible
– Middle lamella: thin layer between primary
walls of adjacent cells
– Secondary cell wall (in some cells): added
between the plasma membrane and the primary
cell wall
• Plasmodesmata are channels between
adjacent plant cells

Fig. 6-28
Secondary
cell wall
Primary
cell wall
Middle
lamella
Central vacuole
Cytosol
Plasma membrane
Plant cell walls
Plasmodesmata
1 μm

Cytoskeletal filaments are all constructed from
smaller protein subunits
Intermediate filaments: smaller
elongated and fibrous subunits
Actin and microtubule filaments:
compact and globular subunits
All form as helical assemblies
of subunits
Noncovalent interactions:
rapid assembly and disassembly

MT’s are a highway-bringing
things out and
back from the center of
the cell.
/

Cilia action
• Cilia= short, many
• Flagella= long, few; NOT the same as the
bacterial flagellum!!

9+2;
nexin, radial spokes, dynein
A,B

The different sides of the
cilium may slide, depending
on the direction of sliding.
These sliding more
These sliding more

• 10 nm in diameter
• Only in animals! (??plant/fungal nucleus??)
• Variety of types- 60 genes!
• Seem to be involved in providing strength
to cells.
• Able to interact with both MT's and
microfilaments (actin filaments).

Intermediate filaments impart mechanical stability
to animal cells
Keratin filaments in epithelial cells
“desmosomes”
The most diverse family
20 in human epithelial cells
10 more in hair and nails
Diagnosis of epithelial
cancers (carcinomas)

Octamers of Tetramers make
up the structure. No polarity!
Subunits are filamentous, rather than
globular.

• Keratin- epithelial cells, hair, nails
• Neurofilaments- in, well, nerves
• Lamins- lines the nucleus

When they are mutant
• Smaller nerve fibers- a natural mutant
quail!
• Fragile skin
• Sometimes muscle weakness
• Sometimes nothing!

Microfilaments (Actin Filaments)
• Microfilaments are solid rods about 7 nm
in diameter, built as a twisted double chain
of actin subunits
• The structural role of microfilaments is to
bear tension, resisting pulling forces within
the cell
• They form a 3-D network called the cortex
just inside the plasma membrane to help
support the cell’s shape
• Bundles of microfilaments make up the core
of microvilli of intestinal cells

Fig. 6-26
Microvillus
Plasma membrane
Microfilaments (actin
filaments)
0.25 μm

• Microfilaments that function in cellular
motility contain the protein myosin in
addition to actin
• In muscle cells, thousands of actin filaments
are arranged parallel to one another
• Thicker filaments composed of myosin
interdigitate with the thinner actin fibers

• Localized contraction brought about by actin
and myosin also drives amoeboid
movement
• Pseudopodia (cellular extensions) extend
and contract through the reversible
assembly and contraction of actin subunits
into microfilaments

• Cytoplasmic streaming is a circular flow of
cytoplasm within cells
• This streaming speeds distribution of
materials within the cell
• In plant cells, actin-myosin interactions and
sol-gel transformations drive cytoplasmic
streaming
Video: CCyyttooppllaassmmiicc SSttrreeaammiinngg

Microfilaments (Actin)
• Where we’re going:
• Basic structure, polarity, treadmilling
• Muscle contraction
• Amoeboid movement

Minus end
Domains 1-4
ATP binding cleft
Subunits= G actin-bound
w/ATP; F-actin=
microfilaments
Looks like
a double
helix!

S1 is a myosin
fragment that binds to
actin- the points point
to the minus end

Treadmilling of actin filaments
actin sub units can flow through the filaments by attaching preferentially to the
(+) end and dissociating preferentially from the (-) end of the filament.
This treadmilling phenomenon occur in some moving cells.The oldest subunits
In treadmilling filament lie at the (-) end.

Treadmilling-it’s easier to
add to the + than – end at
any concentration, and at
some concentrations it’s
adding at the + end at the
rate it’s coming off the –
end= treadmilling.

The treadmilling of an actin filament
Structural difference between the two ends
D form polymer leans towards disassembly

Muscle Contraction
• Three types of muscle fibers:
• Skeletal, striated, voluntary
• Heart- more like skeletal, but not
multinucleated. Its structure allows the
propagation of an action potential (the
heart beats by itself, w/o outside signals)
• Involuntary, smooth muscle- gut, uterus,
etc.

Multinucleated
cell, arises from
fusion; great big
thing- 100mm X
100 um!
2.5 uM length
Muscle contractility

Electron micrograph of sarcomere with bands lettered

The functional anatomy of muscle fibere

Microtubules= interstate; actin= side roads

Light band http://www.youtube.com/wDaatcrhk ?bva=n0dkFmbrRJq4w Light band

Troponin binds Ca++,
moves the tropomyosin
1.5 nm- myosin binds

ARP
Filamin
Fimbrin
gelsolin
Profilin
(thymosins)

Cytoskeleton

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