L26 - Cell Shape and Cytoskeleton

Question 1

Name some factors which may define the shape of a cell

Answer 1

Adjoining cells
Cell adhesions
ECM
cell functions

Question 2

Which sub cellular behaviours can change cell shape

Answer 2

Migration
Phagocytosis
Transport
Cytoskeletal dynamics

Question 3

Give examples of some intermeidate filaments

Answer 3

Keratin, vimentin, lamin

Question 4

F actin is

Answer 4

Filamentous actin

Question 5

Describe F actin under cytoplasmic conditions

Answer 5

Very stable and not a large pool of unpolymersied filament proteins since lots of actin is incorportated into cytoplasmic strucutres

Question 6

Describe some of the strucutres that actin makes within the cell

Answer 6

Cortical actin - around outside 
Microvilli
Flipodia 
Lamellipodia 
Podosome 
Internal stress fibres

Question 7

What are the three stages in the process to form actin

Answer 7

Nucleation - elongation - steady state

Question 8

What is the step in actin filament formation which is unfavourable

Answer 8

Initial oligomerisation

Bottleneck

Question 9

Describe when a saturation point is reached in actin filament formation

Answer 9

When the rate of polymerisation = the rate of depolymerisation

Question 10

Actin monomer addtion occurs at the

Answer 10

+ end (barbed)

Question 11

Monomers are only allowed to bind if they have what bound

Answer 11

ATP

Question 12

Descrieb the role of profillin

Answer 12

Binds monomers and allows phosphorylation increasing the rate of monomer addition at the + end

Question 13

Summary of actin filaments

Answer 13

Polar filaments
Regulated by direct ATP bidning
Treadmilling
Many accessory proteins involved filling

Question 14

Describe the role of profillin

Answer 14

Binds monomers promoting nucelotide ech.

Aids with the defliver of monomers to the barbed end

Question 15

Describe how the Arp2/3 complex acts as a nucleator

Answer 15

Resembles actin so is able to form a startin point for the addition of new monomers

Question 16

As well as being a nucelator how else may Arp2/3 act

Answer 16

Also interacts with existing filaments to cause branching

Question 17

Describe the role of gelosin

Answer 17

Acts to cap and sever actin - by binding to the plus end of an elongating fibre and preventing any further growth

Question 18

Describe the action of a-actinin filamin

Answer 18

Causes bundling and crosslinking which reinforces the strength of the fibre

Question 19

What are small GTPases

Answer 19

Small monomeric (21 kDa) proteins with intrinsic GTPase activity thus they are capable of hydrolysing GTP

Question 20

Many GTPase have a __________________ which targets them to ______________

Answer 20

Post-translational lipid modifications - targets them to specific sites of the membrane

Question 21

All of the small GTPases belong to a large family where the archetypal member is

Answer 21

Ras

Question 22

Functions of Ran

Answer 22

Mitotic spindle organisation

Question 23

Functions of Rab

Answer 23

Endosomal trafficking

Question 24

Functions of Ras

Answer 24

Proliferation - Oncogene

Question 25

What are members of the Rho family

Answer 25

Rac Cdc42 Rho

Question 26

What is the function of the rho family

Answer 26

Involved in the cytoskeleton and migration

Question 27

GAP

Answer 27

GTPase activating proteins GTP --> GDP Reduce activation

Question 28

GEF

Answer 28

Guanine exchange factors GDP --> GTP Activate

Question 29

GDI

Answer 29

Guanine dissociation inhibitors | Prevents dissociation of GDP meaning Rac1 remains in the inactive state for longer

Question 30

GTP hydrolysis acts as a

Answer 30

Timer

Question 31

What about GTPases causes activation of downstream signalling

Answer 31

NOT THE GTP | But the conformational change that it goes through when GTP is bound

Question 32

Describe the switch regions and how they change

Answer 32

Binding of nucleotides causes a structural change in the switch regions Bound nucleotides dictate the signalling activity GTP hydrolysis is an intrinsic property of the protein

Question 33

How many switch regions does Rac1 have

Answer 33

2

Question 34

What is the function of RhoA family members

Answer 34

Stabilises and consolidates actin filaments into a more rigid skeletal framework known as stress fibres

Question 35

What is the function of Rac1

Answer 35

Controls organisation of actin into dynamic ruffling strucutres or lamellipodia

Question 36

What is the function of Cdc42

Answer 36

Controls polymerisation of actin filaments and formation of actin spikes/filopodia

Question 37

Wihat two tools are used for investigating GTPase function

Answer 37

Dominant negative GTPases | Consitutively active GTPases

Question 38

Describe how a constituibely active GTPase may b made

Answer 38

Sub of the catalytic glutamine in switch 2 stops GTP hydrolysis Always GTP bound Signalling always active

Question 39

Describe how a dominant negative GTPase may be made

Answer 39

Subsituition of the P-loop stops the nucelotide binding Nucelotdie is free This acts to mop up active GEFs from acting on functional dominant negative GTPases

Question 40

Effect of a constituitvely active rho mutant

Answer 40

Stress fibre formation

Question 41

Effect of a constitutively active cdc24 or Rac

Answer 41

Membrane ruffling (cdc42) of filopodia formation (Rac)

Question 42

What do activated Rho proteins bind

Answer 42

16 AA sequence in effector proteins - CRIB motif

Question 43

What is a CRIB motif

Answer 43

Cdc42/Rac1 interactive/binding

Question 44

Describe the activtion of Rac or cdc42

Answer 44

Stimulates the formation of new actin filaments

Question 45

Describe how Rac and Cdc42 lead to activation of ARP2/3 what is the overall effect of this

Answer 45

Rac activates WAVE Cdc42 activates WASP Activates ARP2/3 Leads to formation of actin filaments

Question 46

Describe what happens upon activation of Rho

Answer 46

Act of Rho kinase Phosphorylation of myosin Increases myosin contractility and the formation of stress fibres