Exam 3 Oakes Cytoskeleton II

Question 1

Microtubules:

_______ than actin

Play critical roles in ______ & ______

Answer 1

Microtubules:

More complex than actin

microtubules play a critical role in cell division and transport of proteins throughout the cells

Question 2

Microtubules:

Explain their structure

Answer 2

Microtubules:

Long, hollow cylinders made of TUBULIN

More rigid than actin

One end is attached to a MTOC or “centrosome”

MTOC serves as a nuckeation site for tubulin to originate from and spread out from inside the cell

Question 3

Explain Tubulin Structure:

Subunit is a ____

Filaments are ____

___ protofilaments per filament

Tubulin is an ______

Answer 3

Tubulin:

beta stacked ontop of alpha, giving it inherent polarity. Those are stacked ontop of each other, and those protofilaments are put together in a hollow tube

Subunit is a heterodimer (alpha and beta)

Filaments are polarized (beta always on top of alpha)

13 protofilaments per filament

Tubulin is an enzyme, it hydrolyzes GTP

Question 4

Actin uses what energy source

Tubulin uses what energy source

Answer 4

Actin: ATP

Tubulin: GTP

Question 5

The lamellapodia, aka the parts of the cell membrane that branch out, those are full of ____, specifically what type?

Answer 5

Lamellapodia: band of actin at the edges of the cell membrane, full of arp 2/3 branched actin

Question 6

Explain how microtubules undergo hydrolysis (kinda like actin)

What is microtubule catastrophe?

Answer 6

Like actin, microtubules are enzymes that utilize hydrolysis (tubulin uses GTP instead of ATP), polar filament then grows preferentially at one end.

Microtubules will spontanouesly lose their GTP cap (which is a cap at growing end for protection), which results in rapid and stunning depolymerization of the filament

LIKE STRING CHEESE (causes individual protofilaments to peel back from the plus end of the microtubule as if you were peeling string cheese)

Question 7

Explain microtubule nucleation:

____ acts as a seed pattern to grow the complex microtubule filament

There are a total of _______ in a “lock-washer spiral”

This causes an overlap, creating a “____”

Answer 7

Microtubule nucleation:
gamma-TuRC acts as a seed pattern to grow the complex microtubule filaments

Total of seven copies of gamma-TuSC in a “lock washer spiral”

This causes an overlap/ “seam” shape along the microtubule

About 2 protofilaments per TuSC

Question 8

Explain how polymerization isn’t truly restricted to just the plus end

Answer 8

Question 9

Rate limiting step of actin polymerization

Rate limiting step of tubulin polymerization

Answer 9

Actin rate limiting step: formation of the trimer

Microtubules rate limiting: seed forming in tubulin

Question 10

How many protofilaments are there in a microtubule?

Answer 10

13 protofilaments

Question 11

Explain the structure of the MTOC/ “centriole:

Answer 11

Centrosome: a collection of roughly 50 gamma tubulin ring complexes located near the nuclease and from which MOST microtubules are nucleated

Typically positioned in front of nucleus along direction of migration

Pair of centrioles inside MTOC

Question 12

Centrioles: pair of structures consiting of ____ microtubules arranged in a ____ shape

Centrioles are duplicated during ______

Microtubules will self organize with the ____ ends clustered in the center even in the absence of MTOC - this provides a general coordinate system for the cell

Answer 12

Centrioles: pair of structures consisting of 9 microtubules arranged in a barrel shape

Centrioles are duplicated during mitosis

Microtubules will self organize with the minus ends clustered in the center even in the absence of MTOC

Question 13

Top Hat Question:

Microtubules are different than actin in that…. (think energy)

Answer 13

Microtubules hydrolyze GTP instead of ATP (actin)

Question 14

Explain the role of kinesin-13 vs XMAP215 in microtubule polymerization

Answer 14

Kinesin 13 promotes destabilization by increasing the frequency of catastrophes

XMAP215 promotes growth (frequency of catastrophes surpressed and growth rate enhanced)

Question 15

Explain the microtubule crosslinkers:

MAP2 vs tau

What is more spread apart vs more clustered

Answer 15

Microtubule Crosslinkers:

MAP2 gives you more spread out structure

Tau: clustered

Question 16

Kinesin is a motor protein that moves along microtubules, typically towards ______

Kinesin is structurally similar to ______

Kinesin is a member of a large protein super-family, ____ is the only common element

Answer 16

Kinesin is a motor protein that moves along microtubules, typically towards the + end

Kinesin is structurally similar to myosin

Kinesin is a member of a large protein super family, motor domain is the only common element

Question 17

Explain the movement of kinesins across microtubules

Answer 17

Just like myosin and actin, kinesins use ATP hydrolysis to move along microtubules

There are two heads, the lagging head binds ATP, once ATP gets hydrolyzed to ADP, then once PI is released (just like on myosin), there is a conformational change where that foot will go in front. ADP falls off the foot in front, and so on and so on

Question 18

_________ are perfect for carrying cargo, because they kind of walk along the microtubule which acts as a higway to direct trafic

Answer 18

KINESIN is perfect for carrying cargo

one end acts as legs walking down the highway while the other end of the kinesin binds the cargo

Question 19

Dynein:

Unrelated structurally to myosin and kinesin but still couples _____ to ______

_________ cytoskeletal motor

Typically moves towards ____ end

Defects in transport are associated with

Answer 19

Dyenin:

Unrelated structurally to myosin and kinesin but still couples ATP hydrolysis to motion

MOST POWERFUL cytoskeletal motor

Typically moves towards the — (negative end)

Defects in transport related with a number of diseases

Question 20

What mades up cilia and flagella?

Function of flagella, cilia

Answer 20

DYENIN and microtubules make up cilia and flagella

Flagella: used to motility, think sperm

Cilia: whip like motion in gut

9 + 2 configuration with additional crosslinks

Question 21

What are the functions of the two drugs that affect microtubules: taxol and nocodazole

Answer 21

Taxol (chemotherapy): stabilizes microtubules

Nocodazole: depolymerizes by binding subunits

Question 22

Top Hat Question:

Microtubules play a role in what processes?

A. cargo transport

B. cell division

C. polarity

D. A and B

E. A B and C

Answer 22

Microtubules do ALL three of those things

cargo transport, cell division, and polarity

Question 23

IFs:

IFs are fromed from elongated molecules with a ________ domain…… ____ protofilaments form the intermedium filament

Lateral contacts make them __________

There are/are not binding sites for nucleotide triphosphates

Answer 23

IFs are formed from elongated molecules with an extended central alpha helical domain with eight protofilaments forming the intermedium filament

Lateral contacts make them difficult to break but easy to bend

There ARE NO binding sites for nucleotide triphosphates (so IFs are not enzymes)

Question 24

Discuss the roles of IFs

overall role?

plectin

SUN/KUSH

Answer 24

IFs play a role in physical stress. They can oprevent the nucleus from damage. They also play a role in skin cells.

They can be crosslinked to microtubules via plectin

Nuclear IFs can be ancored to microtubules and actin via SUN/KUSH proteins

Question 25

A mutation in _____ (a type of IF) is responsible for a number of different diseases present with blisters

Answer 25

A mutation in keratin cause blister diseases

Question 26

Explain the structure of septins

Are they polar?

Explain the palindrome

_____ different genes broken down in ___ different groups

Answer 26

Septins:
they are not polar

single subunit is made up of a palindromic repeat of 3-4 subunits

13 different genes for human septins, broken into 4 groups

Question 27

Septins Structure:
Front section: rich in _____ (AA)

Middle section: _____ binding domain

Back section: ____

Answer 27

Front: proline rich N terminus

Middle section: GTP binding domain

Back section: coiled coil

Question 28

Like IFs, septins seem to localize to regions that are under ___, like the _______, the _____, and the base of ______ and _____.

They can also recognize _______

Answer 28

Septins localize in regions of stress: plasma membrane, cleavage furrow, and also the base of cilia and flagella

Septins can also recognize curvature

Question 29

Explain the following steps in migration:

1. Protrusion
2. Adhesion
3. Contraction
4. Retraction

Answer 29

1. Protrusion: extension of the PM by actin arp 2/3 (actin polymerization in the lamellipodium)
2. Adhesion: coupling of actin to ECM done by “focal adhesions”…. integrin proteins span the plasma membrane and bind directly to the ECM on the outside, connecting to focal adhesion proteins which connect to the actin on the inside
3. Contraction: force generating step that is a product of myosin II pulling on actin filaments
4. Retraction: dissolutions of adhesions in rear of cell, allow it to move forward

Question 30

Rho and Rac:

Rac signaling drives ______ at the leading edge

Rho A drives ________

These two pathways are ______

Answer 30

Rac drives actin polymerization at the leading edge

(ras causes branched actin web in lamellipodia and less stress fiber formation)

Rho A drives contraction in the cell body

Rho A drives more stress fibers, integrin clustering and focal adhesion formation_

Question 31

Rac causes _____ myosin activity

Rho A causes _____ myosin activity

Answer 31

Rac causes decreased myosin activity

Rho A causes increased myosin activity