1. The Cytoskeleton Functions
Structural scaffold creating and supporting cell shape
Framework positioning organelles within cytoplasm
Network of molecular “roads” for intracellular transport of materials
Framework for whole cell movement
Framework for cell division

2. The Cytoskeleton Three major structural components Microtubules
Major role: support, intracellular transport
Intermediate filaments
Major role: mechanical strength to resist physical stresses
Microfilaments
Major role: muscle contraction, motility

3. The Cytoskeleton Microtubules (MTs) Major role Intracellular transport
Motor proteins drag cargo along them
Structural support
Resist compression forces
Resist shear (bending) forces
Hollow, rigid
25nm diameter, 4nm wall thickness
Radiate outward toward plasma membrane
from near nucleus (MTOC)

4. The Cytoskeleton plus-end minus-end Microtubules (MTs)
Unit = alpha / beta tubulin heterodimer
alpha subunit + beta subunit
Heterodimer is asymmetric
Beta end is called “plus” end
Alpha end is called “minus” end
Not referring to a charge difference
plus-end
minus-end

5. The Cytoskeleton Microtubules (MTs)
alpha / beta (a/b)-tubulin heterodimer
Beta subunit is a GTPase
Assembly
Polymer grows by addition of units at the “plus” end
GTP-bound tubulin can add
GTP form hydrolyzes to GDP form
GDP-bound tubulin cannot add
GDP-bound tubulin can release only from “plus” end
GDP-bound tubulin cannot release from “minus” end or from central region

6. The Cytoskeleton Dynamic instability
MTs can assemble/disassemble at different rates in different locations within a single cell
Various proteins can bind and stabilize MTs

7. The Cytoskeleton Microtubule-associated proteins (MAPs)
Form bridges crosslinking adjacent MTs for parallel alignment
Increase MT stability
Promote assembly
Regulated by phosphorylation state
Anti-tubulin antibody stain

8. The Cytoskeleton Microtubule organizing centers (MTOCs)
GTP-bound a/b-tubulin spontaneously assembles into MTs very slowly
GTP-bound a/b-tubulin add to an existing MT very rapidly
MTOCs are the nucleation points for MT assembly
Centrosome
Basal body

9. The Cytoskeleton Microtubule organizing centers (MTOCs) Centrosome
2 centrioles at right angles to each other near nucleus
Contain gamma-tubulin subunit
Nucleate “minus” end of a/b-tubulin
Plus-end is oriented outward toward plasma membrane

11. The Cytoskeleton Microtubule organizing centers (MTOCs) Basal body
Single centriole at the base of cilia and flagella

12. Eukaryotic cilia and flagella
Hair-like motile organelle projecting from cell surface
Covered by plasma membrane

13. Eukaryotic cilia and flagella
Central protein core is called an “axoneme”

14. Eukaryotic cilia and flagella
Central protein core is called an “axoneme”
Composed of 11 MTs arranged in a “9+2” array
9 outer MTs
2 central MTs
Connected by various MAPs
Locomotion caused by sliding outer tubules past each other
Action of motor proteins (dynein)

16. The Cytoskeleton Motor proteins that “walk” on MTs Kinesin gene family
Plus-end directed
Outward or “anterograde”
transport
Dynein gene family
Minus-end directed
Inward or
“retrograde”

17. The Cytoskeleton
Kinesins are composed of 2 heavy and 2 light polypeptides
Cargo-interaction domain “tail”
Different kinesins have different specificities
ATPase “head”
Binds to MT
ATP hydrolysis propels heads forward
Highly processive

18. The Cytoskeleton
Kinesins are composed of 2 heavy and 2 light polypeptides
ATPase “head”
Binds to MT
ATP hydrolysis propels heads forward
Highly processive

19. The Cytoskeleton Motor proteins that “walk” on MTs Dynein gene family
Minus-end directed
Inward or “retrograde” transport
Very large (1.5MDa)
Involved in cilia/flagella movement

20. The Cytoskeleton Three major structural components
Intermediate filaments (~65 genes)
Major role: mechanical strength to resist physical stresses
Hemidesmosomes and desmosomes

21. Intermediate filaments (IFs)
Animal specific
Strong, rope-like

22. Intermediate filaments (IFs) Animal specific Strong, rope-like
Bridged together with other cytoskeletal elements
(e.g. plectin crosslinks MTs and IFs)

23. The Cytoskeleton Intermediate filaments Composition and assembly
Monomers form dimers
Dimers form tetramers lacking polarity
Tetramers form larger fibers
Incorporation into existing filaments not limited to end regions

24. The Cytoskeleton Three major structural components
Microfilaments (MFs)
Major role: muscle contraction, motility
Solid, branched
8nm diameter
Molecular unit= actin

25. The Cytoskeleton Microfilaments (MFs) Actin molecule is asymmetric
“plus”-end versus “minus”-end
Actin is an ATPase
ATP-bound actin can be incorporated into growing MFs
plus-end of MFs grows 10x faster than minus-end
Higher dissociation rate from minus-end leads to treadmilling

26. The Cytoskeleton Microfilaments (MFs) + cytochalasin D Drugs
Cytochalasin D blocks plus-end addition leading to complete MF depolymerization
Phalloidin blocks turn-over locking MFs into polymerized state
+ cytochalasin D

27. The Cytoskeleton Microfilaments (MFs) + cytochalasin D Drugs
Cytochalasin D blocks plus-end addition leading to complete MF depolymerization
Phalloidin blocks turn-over locking MFs into polymerized state
+ cytochalasin D

28. The Cytoskeleton
Actin binding proteins
+ cytochalasin D

29. The Cytoskeleton Motors that walk on Microfilaments (MFs)
Myosin gene family
ATPase “head” domain
Cargo-interacting “tail” domain

30. The Cytoskeleton Motors that walk on Microfilaments (MFs)
Myosin gene family
Type V can walk on actin filaments carrying a bound cargo
Type II forms bipolar filaments via tail - tail interactions

31. The Cytoskeleton Myosin type II in muscle contraction Muscle fiber
Large cell, 100mm long, microns thick
Contain >100 nuclei
Derived from the fusion of many myoblast cells
Myofibrils
thin protein strands composed of repeating units called “sarcomeres” that give muscle its “striated” appearance
Sarcomere
Z, I, A, H and M regions

32. Myosin II

33. Sliding filament model of muscle contraction

34. Sliding filament model of muscle contraction