BIOL2120 CELL BIOLOGY

Chapter 15

6. Cytoskeleton:
Structures for
Maintaining
Cell Shape
and Internal
Organization
Cytoskeletal Systems

• The interior of a eukaryotic cell is highly

structured

• The cytoskeleton is a network of

interconnected filaments and tubules
extending through the cytosol

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Eukaryotes Have Three Basic Types
of Cytoskeletal Elements

• Immunostaining has been used to characterize

cytoskeletal elements

o Microtubules are composed of tubulin subunits

and are about 25 nm in diameter

o Microfilaments, 7 nm wide, are composed of

actin subunits

o Intermediate filaments, 8–12 nm, are variable

in composition
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Microtubules

• Microtubules are the largest of the

cytoskeletal components of a cell

• There are two types of microtubules

• They are involved in a variety of functions

in the cell

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Two Types of Microtubules Are
Responsible for Many
Functions in the Cell
• Axonemal microtubules include the
organized and stable microtubules
found in structures such as
http://www.biologydiscussion.com/notes/useful-

o Cilia notes-on-cilia-and-flagella-1426-words-biology/517

o Flagella
o Basal bodies to which cilia and
flagella attach

• The axoneme, the central shaft of a

cilium or flagellum, is a highly ordered
bundle of MTs https://en.wikipedia.org/wiki/Cilium

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• Cytoplasmic
microtubules pervade the
cytosol and are
responsible for a variety of
functions
o Placement and
movement of vesicles
o Maintaining or altering
cell shape
o Formation of mitotic
and meiotic spindles

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Tubulin Heterodimers Are the Protein
Building Blocks of Microtubules
• MTs are straight, hollow cylinders of
varied length that consist of (usually
13) longitudinal arrays of polymers
called protofilaments

• The basic subunit of a protofilament

is a heterodimer of tubulin, one a-
tubulin and one b-tubulin

• These bind non-covalently to form

an ab-heterodimer, which does not
normally dissociate

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Subunit structure
• Although a and b subunits share only 40%
amino acid identity, they have very similar 3-D
structure

• Each has an N-terminal GTP-binding domain,

a central domain to which colchicine can bind,
and a C-terminal domain that interacts with
microtubule-binding proteins (MBPs)

• All the dimers in the MT are oriented the same

way

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MT polarity and isoforms

• Because of dimer orientation,

protofilaments have an inherent
polarity

• The two ends differ both chemically

and structurally

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 Cytoplasmic MTs Axonemal MTs
(13 protofilaments) (1 complete ring + 1 or 2 incomplete rings)

Triplet
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Microtubules Form by the Addition of
Tubulin Dimers at Their Ends

• MTs form by the reversible polymerization of tubulin

dimers in the presence of GTP and Mg2+

• Dimers aggregate into oligomers, which serve as

“nuclei” from which new MTs grow

• This process is called nucleation; the addition of

more subunits at either end is called elongation

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Critical concentration
• Microtubule assembly in vitro depends on
concentration of tubulin dimers

• The tubulin concentration at which MT assembly is

exactly balanced by disassembly is called the
critical concentration

• MTs grow when the tubulin concentration exceeds

the critical concentration and vice versa

• The rapidly growing MT end is the plus end (the

opposite end is the minus end)
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Microtubule treadmilling
• The plus and minus ends of
microtubules have different
critical concentrations

• If the [tubulin subunits] is

above the critical concentration
for the plus end but below that
of the minus end, treadmilling
will occur
o Addition of subunits at the
plus end
o Removal of subunits from
the minus end
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GTP Hydrolysis Contributes to the Dynamic
Instability of Microtubules

• Each tubulin heterodimer binds two

GTP molecules, a-tubulin binds one
and b-tubulin binds a second

• GTP is needed to promote heterodimer

interactions and addition to MTs

• The GTP bound to the b-tubulin is

hydrolyzed to GDP after the
heterodimer is added to the MT
http://www.mdpi.com/2221-3759/5/3/8/htm

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GTP-tubulin and dynamic instability

• If [GTP-tubulin] is high, it is added to

an MT, quickly creating a large GTP
cap

• If the concentration falls, the rate of

tubulin addition decreases

• At a sufficiently low concentration, the

rate of GTP hydrolysis exceeds the
rate of subunit addition and the cap
shrinks
http://www.mdpi.com/2221-3759/5/3/8/htm

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Catastrophe and rescue

• If the GTP cap completely

disappears, the MT becomes
unstable and favors loss of GDP-
bound subunits

• Individual MTs can go through

periods of growth and shrinkage
o A switch from growth to
shrinkage is called microtubule
catastrophe
o A switch back to growth phase is
called microtubule rescue

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Microtubules Originate from
Microtubule- Organizing Centers
Within the Cell
• MTs originate from a microtubule-organizing
center (MTOC)

• Many cells have an MTOC called a centrosome

near the nucleus

• In animal cells the centrosome is associated with

two centrioles, surrounded by pericentriolar
material

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g-tubulin
• Centrosomes have large ring-shaped protein complexes in
them

• g-tubulin ring complexes (g-TuRCs) nucleate the assembly

of new MTs away from the centrosome

• Loss of g-TuRCs prevents a cell from nucleating MTs

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MTOCs Organize and Polarize the
Microtubules Within Cells
• MTOCs nucleate and anchor MTs

• MTs grow outward from the MTOC with a fixed polarity

• Because of this, dynamic growth and shrinkage of MTs

occurs at the plus ends, near the cell periphery

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Microtubule Stability Is Tightly Regulated in
Cells by a Variety of Microtubule-Binding
Proteins (MBPs)

• Cells regulate MTs with great precision

• Some MBPs use ATP to drive vesicle or organelle transport

or to generate sliding forces between MTs

o Motor proteins, e.g. kinesins, are the driving force

behind most active transport of proteins and vesicles in
the cytoplasm and the separation of the chromosomes
during mitosis and meiosis

Videos for demonstration

https://www.youtube.com/watch?v=gbycQf1TbM0 26
Microtubule-Stabilizing Proteins
• MTs can be stabilized by proteins that “capture” and protect the
growing plus ends

• These are +-end tubulin interacting proteins (+–TIPs)

• These decrease the likelihood that MTs will undergo catastrophic

subunit loss

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Microtubule-Destabilizing Proteins
• Some proteins promote depolymerization of MTs
o Stathmin binds to tubulin heterodimers and prevents
their polymerization
o Catastrophins act at the ends of MTs and promote the
peeling of subunits from the ends

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Microfilaments

• Microfilaments are the smallest of the

cytoskeletal filaments

• They are best known for their role in muscle

contraction

• They play a role in cell migration, amoeboid

movement, and cytoplasmic streaming

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• Development and
maintenance of cell
shape (via
microfilaments just
beneath the plasma
membrane at the cell
cortex)

• Structural core of
microvilli

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Actin Is the Protein Building Block of
Microfilaments

• Actin is a very abundant protein in all eukaryotic

cells

• Once synthesized, it folds into a globular-shaped

molecule that can bind ATP or ADP
(G-actin; globular actin)

• G-actin molecules polymerize to form

microfilaments, F-actin

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G-Actin Monomers Polymerize into
F-Actin Microfilaments

• G-actin monomers can polymerize reversibly into

filaments with a lag phase and elongation
phase, similar to tubulin assembly

• F-actin filaments are composed of two linear

strands of polymerized G-actin, wound into a
helix

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Actin-Binding Proteins Regulate the
Polymerization, Length, and
Organization of Actin

• Cells can precisely control where actin

assembles and the structure of the
resulting network

• They use a variety of actin-binding

proteins (ABPs) to do so:

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Proteins That Link Actin to Membranes

• MFs are connected to the plasma membrane and

exert force on it during cell movement or cytokinesis

• This (indirect) connection to the membrane requires

one or more linking proteins, e.g. band 4.1, spectrin,
ankyrin, etc.

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Proteins That Promote Actin Branching
and Growth

• Besides bundles and loose networks, actin can form

a dendritic network (treelike)

• A complex of actin-related proteins, the Arp2/3

complex, nucleates new branches on the sides of
filaments

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Cell Signaling Regulates Where and
When Actin-Based Structures
Assemble

• Both plasma membrane lipids, e.g.

phosphatidylinositol, and several
monomeric G proteins regulate the
formation, stability, and breakdown of MFs

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Rho Family GTPases
• The cytoskeleton of cells exposed to certain growth factors
can undergo a dramatic change

• Many signals that result in these changes act via a family of

monomeric G proteins called Rho GTPases

• Activation of the Rho pathway results in formation of stress

fibers (contractile actin bundles) for cell adhesion and
migration
o Myosins are motor proteins responsible for actin-based
motility

Video for demonstration

https://www.youtube.com/watch?v=UmxKxpKua2M 41
 Video for demonstration
https://www.youtube.com/watch?v=t3u2_pAEB94 42
Intermediate Filaments
• Intermediate filaments are the most stable and least
soluble cytoskeletal components and are not
polarized

• An abundant intermediate filament (IF) is keratin,

an important component of structures that grow
from skin in animals

• The nuclear lamina, on the inner surface of the

nuclear envelope, disassemble and then
reassemble during nuclear division

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Intermediate Filament Proteins Are
Tissue Specific

• IFs differ greatly in amino acid composition from

tissue to tissue

• Animal cells can be distinguished based on the types

of IF proteins they contain—intermediate filament
typing

• They are grouped into six classes:

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Intermediate Filaments Assemble from
Fibrous Subunits

• IF proteins are fibrous rather than globular

• All have a homologous rod-like central domain

conserved in size, secondary structure, and to some
extent, in sequence

• Flanking the central helical domain are N- and C-

terminal domains that differ greatly among IF
proteins

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The Cytoskeleton Is a
Mechanically Integrated Structure
• Cellular architecture depends on the unique
properties of the cytoskeletal elements working
together
o MTs resist bending when a cell is compressed
o MFs serve as contractile elements that
generate tension
o IFs are elastic and can withstand tensile forces

• Linker proteins, e.g. plectin, connecting them

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