 The Cytoskeleton:
 Why cells need to be motile?
• Development: cells migrate inside the embryo to their defined locations
• Host defense: motile cells constantly search for pathogens inside the
adult animal
• Wound healing: injured tissues are immediately invaded by highly
motile cells to secret extracellular matrix (ECM) proteins

Why do people need to study the basics of cell movement?
•
Uncontrolled cell migration contributes to several pathologies:
 Vascular diseases
 Chronic inflammatory diseases
 Cancer: tumor formation and metastasis
 What is the basis of cell migration?
•
A cytoskeleton composed of fibers which dynamically reorient, shrink and
grow. Based on this mechanism:
 axons of neurons can grow and connect to other neurons
 muscle cells can contract and produce force
 cells can send out small filopodia to sense their environment
 cells can divide during mitosis
 The cytoskeleton also drives internal movements
• Separation of chromosomes
• Streaming of cytosol (important for plant cells)
• Transport of membranous vesicles:
• Synaptic vesicle transport in neurons
• Endo- and exocytosis
• Membrane flow (recycling of membranes)
 The ability of eukaryotic cells:
•
to adopt a variety of shapes and
•
to carry out coordination and
•
to direct movements depend on a complex network of protein filaments
that extends through out the cytoplasm
Fig 6.1 Alberts 5th Ed
 Although unlike a skeleton made of bone, it is a highly dynamic structure that
organizes continuously as
•
cell changes shape
•
divides and
•
respond to its environment
 In fact, the cytoskeleton might equally well be called the “cytomusculature”
because
•
It is directly responsible for such movements as
 the crawling of cells on the a substratum
 muscle contraction and




 many changes in shape of a developing vertebrate embryo
• It also provides the machinery for intracellular movements such as
 the transport of organelles form one place to another in the cytoplasm
and
 the segregation of chromosomes at mitosis
The cytoskeleton is apparently absent from bacteria and
It may have been a crucial factor in the evolution of eukaryotic cells
The diverse activities of cytoskeleton depend on three types of protein
filaments:
• Actin filaments
• Microtubules and
• Intermediate filaments
Each type of filament is formed from a different protein subunit:
• Actin- actin filament
• Tubulin –microtubules and
• A family of related fibrous proteins such as:
 Vimentin or lamin – intermediate filaments
•
Actin and tubulin have been especially highly conversed through out the
evolution of eukaryotes
•
Their protein filaments bind a large variety of accessory proteins
•
Which enable the same filament to participate in distinct functions in
different regions of a cell
•
Some of these accessory proteins link
 filaments to one another or
 other cell components such as plasma membrane
•
Others control where and when actin filaments and microtubules are
assembled in the cell by regulating:
 the rate and
 extent of their polymerization
•
Yet others are motor proteins, which hydrolyze ATP to produce force and
direct movement along the filament
 The Nature of the Cytoskeleton;
•
A eukaryotic cell contains a billion or so protein molecules, which
constitute about 60% of its dry mass
•
They are thought to be about 10,000 different types of proteins in an
individual vertebrate cell and
•
Most of them are highly organized spatially
•
This organization is present at multiple levels
•
In all cells, proteins are arranged in to functional complexes:
 Most consisting of perhaps 5 to 10 proteins but
 Others as large or larger than ribosomes
•
A further level of organization involves the confinement of functionally
related proteins within the same membrane or aqueous compartment of a
membrane-bounded organelle. Such as:
 Nucleus
 Mitochondria or
 Golgi apparatus
•
An even higher level of organization is created and maintained by
cytoskeleton
 It enable the living cell, like a city, to have many specialized services
concentrated in different areas but
 Extensively interconnected by paths of communication
•
So here, we will discuss some of the basic strategies that enable the
cytoskeleton;
 To control the spatial location of protein complexes and organelles as
well as
 To provide communication paths between them
•
The Cytoplasm of a Eukaryotic Cell is Spatially Organized by Actin
Filaments, Microtubule and Intermediate Filaments;
 How can a eukaryotic cell, with a diameter of 10 µm or more, be
spatially organized by cytoskeletal protein molecules that are typically
2000 times smaller in linear dimensions?
 The answer lies in polymerization
 For each of the three major types of cytoskeletal protein, thousands of
identical protein molecules assemble in to linear filaments
o
that can be long enough, if necessary, to stretch from one side of
the cell to the other
 Such filaments connect protein complexes and organelles in different
regions of the cell and serve as tracks for transport between them
 In addition, they provide mechanical support, which is especially
important for animal cells since they do not have rigid external walls
 The cytoskeleton forms an internal framework for the large volume of
cytoplasm, supporting it like a framework of girders supporting a
building
•
It is easy to see how filaments arose in evolution:
 any protein with an appropriately oriented pair of complementary selfbinding sites on its surface can form a long helical filament
Fig 3.44 Alberts 3rd Ed
Fig 3.45 Alberts 3rd Ed
Fig 3.46 Alberts 3rd Ed
•
Each of the three principal types of protein filaments that make up
cytoskeleton is helical polymer that has a different arrangement in the cell
and a distinct function
Fig 16.2 Alberts 3rd Ed
•
Their functions depend on a large followers of accessory proteins that link
the filaments
 to one another and
 to other cell components
•
Accessory proteins are also essential
o
for the controlled assembly of the protein filaments in particular
location and
o
they provide the motors that either:
o
move organelles along the filament or
o
move the filaments themselves
 Microtubules;
•
Dynamic Microtubules Originate from the Centrosome;
 Microtubules are polar structures:
o
one end i.e. the plus end is capable of rapid growth while
o
other end i.e. minus end tends to lose subunits if not stabilized
 In most cells, the minus ends of microtubules are stabilized by
embedding them in a structure called the centrosome and
 Rapidly growing ends are then free to add tubulin molecules
 The centrosome generally lies next to the nucleus, near the center of
the cell
Fig 16.3 Alberts 3rd Ed
Fig 16.4 Alberts 3rd Ed
•
The Microtubule Network Can Find the Center of the Cell;
 What determines how the cytoplasmic array of microtubules is
normally positioned in a cell?
Fig 16.5 Alberts 3rd Ed
Fig 16.6 Alberts 3rd Ed
 This simple experiment suggest that the cytoplasmic array of
microtubules originating from the centrosome can act as a surveying
device that is able to find the center of the cell
•
Motor Proteins Use the Microtubule Network as a Scaffold to
Position Membrane-Bounded Organelles;
 As we have just seen in the case of fish pigment cells, cytoskeletal
filaments serve not only as structural supports but also as lines of
transport
 If a living vertebrate cells is observed in a light microscope, its
cytoplasm is seen to be continual motion
 Over the course of minutes:
o
mitochondria and smaller membrane-bounded organelles change
their positions by periodic saltatory movements
o
which are much more sustained and directional than the continual
small Brownian movements caused by random thermal motions
 These and other intracellular movements in eukaryotic cells are
generated by motor proteins
o
Which bind to either an actin filament or a microtubule and
use the energy derived from repeated cycles of ATP hydrolysis to
move steadily along it
Fig 5.21 Alberts 3rd Ed
Fig 5.22 Alberts 3rd Ed
Dozens motor proteins have now been identified
They differ
o in the type of filament they bind to
o the direction in which they move along the filament and
o the cargo they carry
The first motor protein to be discovered was myosin – a protein that
moves along actin filament and
is especially abundant in skeletal muscle where it forms a major part
of the contractile apparatus
Other types of myosins were subsequently found in non-muscular
cells
All myosins have similar motor domains – the part of the protein that
generates movement
But they differ markedly in domains that are responsible for attaching
the myosin molecules to other component of the cell
o







 The motor proteins that move along microtubules are distinct from
the myosins and belong to one of two families:
o
Kinesins – generally move towards the plus end of microtubules
i.e. away from centrosome and
o
Dyneins – move towards the minus end i.e. toward centrosome
Fig 16.7 Alberts 3rd Ed
 Microtubule-dependent motor proteins play an important part in
positioning membrane-bound organelles within a eukaryotic cell. For
example:
o
The membrane tubules of the endoplasmic reticulum (ER) align
with microtubules and extend almost to the edge of the cell
o
Whereas the Golgi apparatus is located near the centrosome
 When cells are treated with a drug that depolymerizes microtubules,
both of these organelles change their location:
o
ER collapses to the center of the cell and
o
Golgi apparatus fragments in to small vesicles that disperse
though out the cytoplasm
 When the drug is removed, the organelles return to their original
positions, dragged by motor proteins moving along the reformed
microtubules
Fig 16.8 Alberts 3rd Ed
•
Microtubules are Hollow Tubes Formed from Tubulin;
 Microtubules are formed from molecules of tubulin -protein
 Each of which is heterodimer consisting of closely related and tightly
linked globular polypeptides called
o
α - tubulin and
o
β - tubulin
Fig 16.8 Alberts 3rd Ed
 Dynamic instability of microtubule requires an input of energy to shift
the chemical balance between polymerization and depolymerization
 That energy comes from the hydrolysis of GTP
Fig 16.33 Alberts 3rd Ed
 Many of the microtubule arrays are labile and
 Depend on this liability got their function
 One of the most striking examples is
o
mitotic spindle which forms after the cytoplasmic microtubules
disassemble at the onset of mitosis
(A)
the mitotic spindle is the target of a variety of specific antimitotic
drugs that
o act by interfering with the exchange of tubulin subunits between
the microtubules and free tubulin pool i.e.
o blocking the polymerization of tubulin into microtubules
o one of these is colchicine – an alkaloid extracted from the
meadow saffron that has been used medicinally in treatment of
gout since ancient Egyptian times
o The drug taxol, extracted from the bark of yew trees has opposite
effect
Cilia and Flagella;
 Cilia and flagella are highly specialized and efficient motility
structures built from microtubules and dynein
Both are hair like appendages about 0.25 µm in diameter with a
bundle of microtubules at their core
Fig 16.39 Alberts 3rd Ed
 Flagella are found on sperm and many protozoa
o By their undulating motion, they enable the cells to which they are
attached to swim through liquid media
 Cilia tend to be shorter than flagella and are organized in a similar
fashion but they beat with a whip-like motion
o
•
 Ciliary beating can either propel single cells through fluid as
o in swimming of the protozoan Paramecium or
o can move fluid over the surface of a group of cells as in a tissue
 In the human body
o
huge numbers of cilia (109/cm2 or more) line our respiratory tract,
sweeping layers of mucus, trapped particles of dust and bacteria
up to the mouth where they are swallowed and ultimately
eliminated
 Likewise, cilia along the oviduct help to sweep eggs towards the
uterus and the flagellum propels sperm
 The movement of a cilium or a flagellum is produced by the bending
of its core called axoneme
 It is composed of microtubules and their associated proteins, arranged
in a distinctive and regular pattern
 Nine special doublet microtubules (comprising one complete and one
partial microtubule fused together so that they share a common tubule
wall) are arranged in a ring around a pair of single microtubule
Fig 16.41 Alberts 3rd Ed
 Actin Filaments;
•
They consists of a tight helix of uniformly oriented actin molecules also
known as globular actin or G actin
Fig 16.49 Alberts 3rd Ed
•
Like microtubule, actin filament is a polar structure with two structurally
different ends:
 a relatively inert and slow growing minus end ( pointed end) and
 a faster growing plus end (barbed end)
•
Three dimensional structure of actin has been solved by x-ray diffraction /
crystallography
Fig 16.50 Alberts 3rd Ed
Fig 16.51 Alberts 3rd Ed
•
The role of ATP hydrolysis in actin polymerization is similar to the role of
GTP hydrolysis in tubulin polymerization
•
Actin is involved in a remarkably wide range of structures i.e.
 from stiff and relatively permanent extensions of the cell surface to the
dynamic three dimensional networks at the leading edge of a
migrating cell
•
So the varied forms and functions of actin in eukaryotic cells depend on a
versatile repertoire of actin-binding proteins that
 cross link actin filaments into loose gels
 binding them into stiff bundles
 attach them to plasma membrane or
 forcibly move them relative to one another
Fig 16.65 Alberts 3rd Ed
•
For example; solate
 Tropomyosin binds along the length of actin filaments, making them
more rigid and alternating their affinity for other proteins
 Filamin cross links actin filaments in to a loose gel
 Fimbrin and α-actinin form bundles of parallel actin filaments
 Gelsolin mediates Ca2+- dependent fragmentation of actin filaments,
thereby causing a rapid solation of actin gels
 Various forms of myosin use energy of ATP hydrolysis to move along
actin filaments either
i. Carrying membrane-bounded organelles from one location in the
cell to another or
ii. Moving adjacent actin filaments against each other
 Sets of actin-binding proteins are thought to act cooperatively in
generating the movements of the cell surface including:
•
o
Cytokinesis
o
Phagocytosis and
o
Cell locomotion
Muscle;
 Many of the proteins that associate with actin filaments in eukaryotic
cell were first discovered in muscle
 Muscle contraction is the most familiar and the best understood of all
kinds of movement of which animals are capable
 For example, in vertebrates:
o
Running
o
Walking
o
Swimming and
o
Flying all depend on the ability of skeletal muscle to contract
rapidly on its scaffolding of bone
o
while involuntary movements such as heart pumping and
o
gut peristalsis depend upon on the contraction of cardiac and
smooth muscle respectively
 The long thin muscle fibers of skeletal muscle are huge single cells
formed during development by the fusion of many separate cells
 The nuclei of the contributing cells are retained in this large cell and
lie just beneath the plasma membrane
 But bulk of the cytoplasm is made up of mycofibrils
 Which are contractile elements of the muscle cell
 They are cylindrical structures 1 to 2 µm in diameter and are often as
long as the muscle cell itself
Fig 16.82Alberts 3rd Ed
Fig 16.83 Alberts 3rd Ed
Fig 16.69 Alberts 3rd Ed / Fig 16.54 Alberts 5th Ed
Fig 16.88 Alberts 3rd Ed
Fig 16.89 Alberts 3rd Ed
 How contraction occurs? What is actual mechanism?
 The sliding filament model was first proposed in 1954 which provided
the crucial understanding of contractile mechanism
Fig 16.85 Alberts 3rd Ed
Fig 16.50 Alberts 3rd Ed
Fig 16.91 Alberts 3rd Ed
 The force generating molecular interaction takes place only when a
signal passes to the skeletal muscle from its motor nerve
 The signal from the nerve triggers an action potential in the muscle
cell plasma membrane and
 this electrical excitation spreads rapidly in to a series of membranous
folds, the transverse tubules or T tubules
 The signal is then relayed across a small gap to the sarcoplasmic
reticulum
Fig 16.92 Alberts 3rd Ed
 Intermediate Filaments;
•
These are tough and durable protein fibers found in cytoplasm of most but
not all animal cells
•
They are called intermediate because in electron micrographs their
apparent diameter (8-10 nm) is between
 that of the thin actin filaments and
 the thick myosin filament of muscle cells where they were first
described
•
Further they are also intermediate in diameter between actin filaments and
microtubules
•
In most animals cells, an extensive network of intermediate filaments
surrounds the nucleus and extend out to the cell periphery, where they
interact with the plasma membrane
Fig 16.12 Alberts 3rd Ed
•
In addition, a tightly woven basketwork of intermediate filaments i.e. the
nuclear lamina underlies the nuclear envelope
•
Intermediate filaments are particularly prominent in the cytoplasm of cells
that are subject to mechanical stress. For example;
 They are present in large numbers in epithelia
 Where they linked from cell to cell at specialized junctions
o
along the length of nerve cell axons and
o
in all kinds of muscle cells
•
Intermediate Filaments are Polymers of Fibrous Proteins;
 Unlike actin and tubulin, which are globular proteins, many types of
intermediate filament protein monomers are all highly elongated
fibrous molecules that have
•
o
an amino –terminal head
o
a carboxyl-terminal tail and
o
central rod domain
Fig 16.13 Alberts 3rd Ed
Fig 16.14 Alberts 3rd Ed
Epithelial Cells Contain a Highly Diverse Family of Keratin
Filaments;
 The cytoplasmic intermediate filaments in vertebrate cells can be
grouped in to three classes:
i. Keratin filaments
ii. Vimentin and vimentin-related filaments and
iii. Neurofliaments
 Each formed by polymerization of their corresponding subunit
proteins
The anti-parallel arrangement of dimers implies that the tetramer and hence the intermediate filament
that is formed id a non-polarized structure i.e. it is same at both ends and symmetrical along its length
Table 16.1 Alberts 3rd Ed
 The nuclear lamina is a mesh work of intermediate filaments that lines
the inside surface of the nuclear membrane in eukaryotic cells
Fig 16.18Alberts 3rd Ed
 It is typically 10-20 nm thick and is interrupted in the region of
nuclear pores to provide a passageway for macromolecules entering
and leaving the nucleus
 In mammalian cells, the nuclear lamina is composed of lamins which
are homologous to other intermediate filament proteins but
 Differ from them at least four ways:
i.
Their central rod domain is somewhat longer
ii.
They contain a nuclear transport signal that directs them from the
cytosol (where they are made) in to nucleus
iii.
They assemble in to a two-dimensional, sheet like lattice which
is thought to require their association with other proteins
iv.
The mesh work they form is usually dynamic and
o
rapidly disassembles at the start of mitosis and
o
reassembles at the end of mitosis
o
•
The disassembly and reassembly is mediated by the
phosphorylation and dephosphorylation of several serine
residues on the lamins
Intermediate Filaments Provide Mechanical Stability to Animal Cells;
 There is increasing evidence that a major function of cytoplasmic
intermediate filaments is to resist mechanical stress
 In human genetic disease – Epidermolysis bullosa simplex;
 mutation in keratin genes that are normally expressed in the basal cell
layer of epidermis disrupt the keratin filament network in these cells
 Making them very sensitive to mechanical injury. Therefore,
o
a gentle squeeze can cause the mutant basal cells to rupture and
o
the skin is blistered in affected individuals
o
A similar condition can be produced in transgenic mice that
express mutant keratins of this type
Fig 16.19 Alberts 3rd Ed
x -------------------------- x --------------------------x------------------------ x