Details of cytoskeleton element-microtubule. The Microtubule associated protein-type and function, Treadmilling and dynamic instability, Structure of cilia and flagella

1. Microtubules
Dr. Sarita Nanda
Biochemistry Department
Daulat Ram College

2. Characteristics
 They are 25nm in diameter
 They are dynamic structure which continually
undergo assembly and disassembly
 They determine cell shape, cell locomotion, cell
organelle movement, separation of chromosomes
during mitosis.
 They are made up of monomers tubulin.
 The tubulins are alpha and beta. Gamma tubulin is
present in centrosome


3. Characteristics
 Tubulin dimers polymerize to form microtubule
which is made of 13 linear protofilaments.
 The dimers are arranged in head to tail manner in
parallel fashion.
 They have the fast growing plus end and slow
growing minus end.
 This gives polarity to microtubule.

4. Characteristic
 They undergo treadmilling in which fast growing end
is adding GTP bound tubulin and minus end is
continuously loosing GDP bound from the minus
end.
 In dynamic instability individual microtubules
shrink between cycles of growth and shrinkage.
 Dynamic instability is described by Tim Mitchison
and Marc Kirschner in 1984.

5.  Colchicine, colecimid, vincristine, vinblastine , taxol
affect polymerisation and block cell division.

6. Treadmilling

7. Assembly of Microtubules
 They extend from centrosome (ist describes Theodor
Boveri in 1888
 During mitosis microtubules form spindle
 Plants don not have organised centrosome
 Centrosome is microtubule organising centre
(MTOC) in which minus end is anchored. It serves
as initiation site for MT assembly.
 The key protein of centrosome is gamma tubulin.
 Gamma tubulin forms a ring complex.

8. characteristic
 The centrosome of animal cell contains a pair of
centrioles.
 Centrioles are cylindrical structures which contains nine
triplets of microtubules.
 Centrioles are necessary to form basal body, cilia,
flagella.
 The peritubular region initiates microtubular assembly.
 Centrioles have cartwheel like structure.
 Centrioles also include delta tubulin.
 Two centrioles are connected by protein called centrin.

9. Organisation of Microtubules within Cells
 They interact with microtubule associated
protein(MAPs)
 MAPS stabilise MT by capping their ends
 MAPs can also destabilise MT by severing their ends
 Several MAPs are plus end tracking proteins. The several
identified MAPs are MAP1,2, and TAU
 The neurons have dendrites and axons . The plus end of
axons are away form the body and in dendrites plus and
minus ends are oriented away from the body.
 Axons contain tau proteins and no MAP-2 whereas
dendrites contain MAP2 but no tau protein

10. Microtubule Assosciated protein

11. Microtubule

12. Microtubules motors and Movement
 The two motor protein responsible for motor
movement are kinesin and dyenin
 Kinesin moves towards plus end
 Dynein moves towards negative end.
 Dyenin was isolated by Ian Gibbons in 1965.
 Motor protien was observed by video-enhanced
microscopy.(developed by Robert Allen and shinya
Inoue in 1980. )
 These were observed in squid axons.

13. Motor Proteins
 Kinesin was identified By Brady, Ronald Vale,
thomas Reese and Michael Sheetz in squid axons and
bovine brains in 1985.
 Kinesin translocates towards plus end and dyenin
translocates towards minus end.
 Kinesin I is a molecule of 380KD consisting of two
heavy chains of 120 kD and two light chains of 64kd
each.
 It is similar to myosin which also moves towards
postive end and has molecular weight of 500kd.

14. Motor proteins
 Cytoplasmic dynein is extremely large protein of 2000kd.
It consists of two heavy chains of 500kd and variable
number of light and inermediate chains
 The molecule has head and a tail portion.
 The head binds to ATP and moves on MT
The tail binds the organellles which have to be
transported.
 There are 45 kinesinsin humans
 The plus end directed kinesin have motor domain at N
terminal end, minus end directed have motor domains
at C terminal end. Others have motor domains in the
centre.

15. Motor proteins
 Dynein may be cytoplasmic or axonemal.
 The tail region may carry cargo

16. Cargo Transport and Intracellular Organisation
 Helps to carry organelles
 Kinesin carries cargo towards the cell periphery
whereas dynein carries toward the nucleus.
 This helps in positioning ER/Golgi Apparatus.
 Mitochondria may be transported from cell body to
axon
 Neurotransmitter are carried from Golgi apparatus
to terminal branches of axons by kinsens
 Endocytic vesicles move from axon back to cell body

17. Cargo Transport and Intracellular Organisation
 Kinesin II moves mRNA towards cell cortex in
xenopus oocyte.
 Kinesin I transports actin mRNA in fibroblasts.
 ER is positioned by Kinesin I
 GA is positioned by cytoplasmic dynein.

18. Cilia and Flagella
 MT based projections
 Cilia are widespread
 Bacterial flagella quite different from eukaryotic flagella
 Bacterial flagella are protein filaments. Eukaryotic
flagella are projection of plasma membrane supported by
MT
 Cilia and flagella are quite similar structures approx 0.25
um.
 Paramecium are covered with cilia
 Flagella differ from cilia in length.

19. Cilia and Flagella
 Composed of fundamental structure axoneme composed
of MT.
 They are arranged in 9+2 pattern.
 A tubule is complete consisiting of 13 protofilament
 B tubule is incomplete consiisting of 10/11 protofilament
 The outer MT are connected to central pair by radial
spokes and to each other by nexin.
 In addition 2 arms of dyenin are attached to each A
tubule and motor activity of axonemal dynein that drive
cilia and flagella.

20. Cilia and Flagella

21. Cilia and flagella
 The basal body have 9+O arrangement
 It initiates growth of axonemal MT
 Movement of cilia and flagella result from sliding of
outer MT doublets relative to one another by motor
activity of axonemal dynein
 Dynein head binds to A tubule whereas tail binds B
tubule binds to B tubule
 Movement of dynein head groups in the minus direction
then causes a tubule of one doublet to slide towards basal
end of adjacent B tubule


22. Cilia and flagella

23. Cilia and flagella
 Since two doublets are connected by nexin, sliding
movement helps cilia and flagella to bend.
 This produces wave like oscillation.