Learning objectives: Role of cytoskeleton & molecular motors Differentiate the three types of cytoskeletal elements Drugs acting on cytoskeletal elements and their uses Pathological basis of disorders associated with cytoskeletal elements Interaction between cytoskeleton and molecular motor proteins

1. Cytoskeleton & Molecular Motors

2. • Role of cytoskeleton & molecular motors
• Differentiate the three types of cytoskeletal
elements
• Drugs acting on cytoskeletal elements and
their uses
• Pathological basis of disorders associated
with cytoskeletal elements
• Interaction between cytoskeleton and
molecular motor proteins
Learning objectives

3. Introduction
• Cytoskeleton is a cellular structure
that helps cells maintain their
shape and internal organization.
• It also provides mechanical
support that enables cells to carry
out essential functions like cell-
division, anchorage and
movement.

4. 3 classes of cytoskeleton
1. Microfilaments
2. Intermediate filaments
3. Microtubules

5. 3 classes of cytoskeleton
Type Size (nm) Examples
Microfilaments 5-7 Actins
Intermediate
filaments
8-10 Lamins, Keratins, Vimentin,
Desmin, Neurofilaments
Microtubules 23-25 Tubulins

6. G-actin to F-actin

7. + and - end

8. Actin fold
• ATP binding domain of actin is known as actin fold.
• Also present in,
– hexokinase (glycolytic enzyme)
– Hsc70 (chaperone)

9. Actin is a component of thin filament

10. Role of microfilaments
• Actin-myosin complex is involved in muscle contraction
• Microfilaments help the cells to change their shape.
(Amoeboid movement of phagocytes is due to
pseudopodia (false feet) formation)
• Contractile ring that forms at the end of mitosis is made
up of actin-myosin complex. Constriction of this ring
produces two new daughter cells.

11. Actin associated proteins
Protein Location
Fimbrin Microvilli, Stereocilia
α actinin
Filopodia, Lamellipodia, Stress fibers, Adhesion
plaques
Spectrin
Cortical network of cells, i.e. specialized layer of
cytoplasmic protein on the inner face of the
plasma membrane of the cell periphery
Dystrophin Muscle cortical network
Filamin Filopodia, Pseudopodia, Stress fibers
Fascin
Filopodia, Lamellipodia, Stress fibers, Microvilli,
Acrosomal process
Villin Microvilli in intestinal and renal brush border

12. Microtubules

13. Microtubules
• Microtubules are the largest cytoskeletal
element made up of tubulin proteins.
• There are two types of tubulins - alpha and beta.
• Tubulin heterodimers organise into a hollow
shape to form microtubules.
• The hollow shape provides the mechanical
strength to bear the shear stress exerted upon
the microtubules.

14. Multiphoton fluorescence image of HeLa cells with cytoskeletal microtubules
(magenta) and DNA (cyan). Appreciate the microtubules spreading across each and
every corner of the cell.

15. Mitotic spindle

16. Microtubules in cancer and metastasis
• Cancer cells divide rapidly and are able to
metastasise to distant places.
• Microtubule inhibitors are used in the therapy of
cancer.
– Vinca alkaloids (vincristine and vinblastine) and
Taxanes (Paclitaxel) – Anti-cancer agents
– Colchicine is used to cause metaphase arrest
for karyotyping and to prevent chemotaxis in
acute gouty arthritis

17. Mechanism of action of Vinca alkaloids and taxanes
are different

18. Intermediate Filaments
Subtype Protein composition Tissue distribution
Type I Acidic Keratin Epithelia
Type II Basic Keratin
Type III
Vimentin Mesenchymal cells
Desmin Muscle cells
Glial fibrillary acidic protein
Glial cells, astrocytes,
stellate liver cells
Peripherin Diverse neuronal cells
Synemin Diverse neuronal cells
Type IV
Neurofilament – Low, Medium
and high Neurons
Type V Lamins – A, B and C Nuclear Lamina
Unclassified
Phakinin
Lens
Filensin

19. Cultured epithelial cells - Keratin is stained red and
DNA (nucleus) is stained green. Look at how the
keratin filaments are surrounding cells.

20. Unique features of intermediate filaments
• Intermediate filaments are less dynamic
• They do not contain polarity.
• They are tissue-specific, e.g. Neurofilament is present in
neurons only.

21. Keratin is a marker of epithelial cells
H&E staining of carcinoma. Look at the whorled pattern of
keratin. This is known as Keratin pearls.

22. Diseases due to defective intermediate
filaments
• Epidermolysis bullosa simplex
(bullous formation after a trivial
trauma) – due to keratin 5 mutation
• Hutchinson-Gilford Progeria –
mutation in Lamin A protein
• Amyotrophic lateral sclerosis (ALS)–
defect in neurofilaments (NF-L, NF-
H)

23. Mutations in the gene encoding lamin A and
C (they are formed by alternative splicing)
Defective scaffolding of nucleus -
Accumulation of farnesylated lamin A
Signs and symptoms of premature aging
(progeria)
Pathogenesis of progeria

24. Comparison of cytoskeletal elements
Microfilaments Intermediate
filaments
Microtubules
Size (nm) 5-7 8-10 23-25
Subunit
proteins
Actins (alpha, beta
and gamma)
Lamins, Keratins,
Vimentin, Desmin,
Neurofilaments
Tubulins (alpha
and beta)
Structure Beaded structure α helical rods that
assemble into rope
like filaments.
Hollow tubules.
Requirement of
nucleotide
hydrolysis
ATP hydrolysis for
polymerisation of G-
actin to F-actin
Not needed Needed

25. Role of cytoskeleton
• Giving shape & strength to the cell
• Anchorage
• Intracellular transport
• Muscle contraction
• Beating of cilia
• Amoeboid movement of
phagocytic cells

26. Molecular motors
• Molecular motors are protein that convert
chemical energy to mechanical work.
• Most of the molecular motors utilise the free
energy of hydrolysis of ATP.

27. Classes of molecular motors
Class Example
Cytoskeletal
motors
Myosin, Dynamin, Kinesin, Dynein
Rotary motors FoF1-ATP synthase
Nucleic acid
motors
DNA polymerase, RNA polymerase,
Helicase, Topoisomerases

28. Cytoskeletal Motors
Molecular motor
Cytoskeletal
railroad
Fuel
used
Function
Myosin Actin ATP Muscle contraction
Dynamin Actin GTP
Receptor mediated
endocytosis
Kinesin Tubulin ATP
Anterograde transport (e.g.
neuronal transport)
Dynein Tubulin ATP
Retrograde
transport (e.g. cilia)

29. Kinesin, a molecular motor protein moving
along the microtubules.

30. Eukaryotic flagellum showing the 9+2 arrangement
of microtubules. Dynein arms are shown in blue.

31. Dynein arm defect leads to Primary ciliary
dyskinesia/Kartagener syndrome
A – Normal cilia B – Cilia of Kartagener’s
syndrome

32. Summary
• The cytoskeleton is a cellular structure that helps cells maintain their shape and
internal organization.
• Cytoskeleton consists of three major class of protein elements namely
microfilaments, intermediate filaments and microtubules.
• Microfilament is formed by actin family of proteins
• Microtubules are the largest cytoskeletal element made up of tubulin proteins.
• Microtubule inhibitors are used in the therapy of cancer.

33. Summary
• Intermediate filaments are less dynamic and are made of at least 40 different subunit
proteins.
• Cytoskeletal Molecular motors are the vehicles carrying the cargo. They run upon the
cytoskeletal framework.
• Myosin is the motor protein that move along actin filaments using the energy of ATP
hydrolysis while Kinesin is the motor protein that move along tubulin using the energy of ATP
hydrolysis
• Dynamin requires free energy of hydrolysis of GTP.
• Primary ciliary dyskinesia (PCD), also called immotile ciliary syndrome or Kartagener
syndrome is an autosomal recessive disorder due to dynein arm defect.