Lecture 17 & 18 - Microtubules

Question 1

What is the main purpose of the cytoskeleton?

Answer 1

it maintains cell shape, organization, and provides support for internal and external movement

Question 2

What are the 3 classes of cytoskeletal filaments? What proteins make them?

Answer 2

1. microfilaments: actin
2. microtubules: tubulin
3. intermediate filaments: various proteins

Question 3

What is a tubulin subunit?

Answer 3

a heterodimer made of two closely related globular proteins called alpha-tubulin and beta-tubulin

Question 4

A microtubule is built from 13 ________ _________

Answer 4

parallel protofilaments; each composed of alpha/beta-tubulin heterodimers stacked head to tail and then folded into a tube

Question 5

Why is the helical shape advantageous?

Answer 5

the helical microtubule lattice makes them stiff and hard to bend

Question 6

What gives microtubules polarity?

Answer 6

the orientation of the subunits: the microtubule plus end grows and shrinks more rapidly than its minus end

Question 7

What are the 3 phases of microtubule growth, starting with individual dimers?

Answer 7

1. Lag phase - nucleation
2. Elongation phase
3. Plateau phase

Question 8

What is microtubule nucleation?

Answer 8

the process in which several tubulin molecules interact to form a microtubule seed
- tubulin dimers assemble into protofilaments
- laterally associated linear protofilaments
- slow process: must form out of nothing

Question 9

Dynamic instability

Answer 9

a process in which individual microtubules alternate between cycles of growth and shrinkage

Question 10

What does the addition of GTP-tubulin to the plus end of a protofilament do?

Answer 10

GTP-tubulin = alpha tubulin

it causes the end to grow in a linear conformation that assembles into the cylindrical wall of the microtubule

Question 11

What happens to GTP in the microtubule?

Answer 11

at the (+) end it stabilizes the microtubule by creating a GTP cap, this allows the microtubule to grow
GTP gets hydrolyzed once its in the microtubule - this region is unstable and will not stand by itself without the GTP cap

if all GTP gets hydrolyzed, the microtubule begins to shrink

Question 12

Catastrophe vs rescue

Answer 12

Catastrophe is the change from growth to shrinkage

Rescue is the change from shrinkage to growth

Question 13

How do scientists think catastrophe works?

Answer 13

hydrolysis of GTP after assembly changes the conformation of the subunits and tends to force the protofilament into a curved shape that is less able to pack into the microtubule wall

Question 14

What complex does nucleation depend on?

Answer 14

gamma-tubulin ring complex
- anchored to accessory proteins
- prevents nucleation from being very slow by providing support for tubulins to grow
- forms helical pitch so tubulin subunits can join to that instead of forming out of nowhere

Question 15

Where do microtubules generally nucleate from?

Answer 15

the microtubule-organizing center (MTOC) where gamma-tubulin is most enriched
- many animals possess a single, well-defined MTOC called the centrosome

Question 16

Centrosomes

Answer 16

composed of 2 centrioles and are surrounded by a dense mass of protein termed the pericentriolar material
- gamma-tubulin is in the pericentriolar material

Question 17

What structure is gamma-tubulin located in? When is it not there?

Answer 17

the centromere; when the cell is dividing

Question 18

Microtubule associated proteins (MAPs)

Answer 18

proteins that bind and stabilize microtubules
- help microtubules interact with other things

Question 19

Map2 and Tau

Answer 19

MAPs
- set the spacing of the microtubule bundles
- Map2 is a long protein => larger spacing of bundles
- Tau is a shorter protein => smaller spacing of bundles

Question 20

What can cause neurodegenerative diseases?

Answer 20

Tau mutations
- affects how neurons communicate => remove Tau => spacing is too tight (or microtubules are not bundled) => vesicles cannot travel through

Question 21

What are classic kinesins?

Answer 21

motors that move towards the (+) end of microtubules
- moves from center (nucleus) to plasma membrane

Question 22

Kinesin

Answer 22

• a tetramer protein
• two heavy chains & two light chains
• heavy chain has many domains
• head splits ATP and converts the energy into motion
• tail is cargo-binding

Question 23

Movement of kinesins

Answer 23

• at least one kinesin head is always attached to the microtubule
• progressive steps (all the same size)
• for long-range

Question 24

What is the common element in the kinesin family?

Answer 24

the motor domain of the heavy chain

Question 25

Kinesin-13 proteins

Answer 25

(microtubule depolymerases)
- induce depolymerization uniquely from both ends of the microtubule
- incapable of movement
- regulate microtubule dynamics to control spindle assembly

Question 26

Kinesin-14

Answer 26

unusual: it moves from (+) end to (-) end in motility assays

• its tail can bind microtubules and allows it to organize microtubule bundles
  - parallel microtubules
  - antiparallel microtubules
  - clustering microtubules
  - overlap microtubules

Question 27

What is another microtubule motor protein, besides kinesin?

Answer 27

cytoplasmic dynein

Question 28

Dynein

Answer 28

• much larger and more complex than kinesin
• head is force generating motor (AAA = ATPase domain)
• stalk contains the microtubule-binding site at its tip
• tail binds cargo
• ATP changes the conformational structure to disassociate microtubule binding

Question 29

Dynein steps and direction

Answer 29

big but irregular; moves towards minus ends

Question 30

Compare kinesin and dynein

Answer 30

Kinesin:
- small
- moves towards plus end
- ATP driven
- regular steps

Dynein:
- big
- moves towards minus end
- ATP driven
- irregular movements

Question 31

Microtubule motors move _______ in the secretory pathway

Answer 31

vesicles; kinesin-mediated and dynein-mediated transport of vesicles, vesicular-tubular structures, and organelles

Question 32

If both motor proteins are attached to a vesicle, how does it get transported in either direction?

Answer 32

kinesin must be inhibited for minus-end transport
- kinesin-binding proteins prevent kinesin-microtubule binding

Question 33

Cilia and flagella

Answer 33

hairlike cell appendages that have a bundle of microtubules at their core
- the core is called the axoneme, which is composed of microtubules and their associated proteins
- axonemal dynein bends the axoneme which moves the cilium and flagellum

Question 34

Melanosomes

Answer 34

rapid movement is mediated by dynein (or dynamin??) and microtubules
- melanosome is an organelle that synthesizes and stores melanin
- coordinated movement because microtubules are uniformly polarized