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Cell and Molecular > L25 - Cytoskeleton and Cell Shape > Flashcards

L25 - Cytoskeleton and Cell Shape Flashcards

1
Q

What 3 things define cell shape?

A

Adjoining cells – physically boxes cells in
Cell adhesions
Extra-cellular matrix

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2
Q

What cells show the important of cell shape for function?

A

Nerves
Muscles
Red blood cells

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3
Q

Why is cell shape vital for cellular activity?

A

Migration – cell shape changes drive movement
Phagocytosis – cytoskeleton changes shape when engulfing
Transport – cytoskeleton movements used to move vesicles
Cytoskeletal dynamics

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4
Q

F-actin monomers role?

A

No structural role

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5
Q

F-actin polymers role?

A

Cortical actin - around the edge of the cell to maintain its shape
Filopodium - to sense surrounding environment
Lamellipodium – important for migration
Podosome – used to anchor the cell
Internally stress fibres – many stripes together to provide strength

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6
Q

The actin cytoskeleton in the inactive state is?

A

Very stable

Very little unpolymerized actin in cytoplasm

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7
Q

What regulated disassembly of F-actin?

A

Phosphorylation

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8
Q

Why do actin filament self-polymerise?

A

Actin in solution will polymerise by itself – because of their shape
Initial step is energetically unfavourable

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9
Q

Actin polymerisation steps

A
  1. Add salt to a group of monomers to allow polymerisation
  2. Olgiomers polymerise to form a growing actin filament
  3. A point where the rate of monomers added and falling off is equal – steady state of fibres
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10
Q

What is actin treadmilling?

A

This is the process of how we regulate actin polymerisation in cells

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11
Q

How are monomers added to actin filaments?

A

Added to the + end

Removed at the – end

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12
Q

What regulates the addition of monomers to actin filaments?

A

Phosphorylation regulates this - ATP  ADP and Pi resulting in monomer release

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13
Q

What accessory protein is associated with actin filaments?

A

Profilin – binds to monomers enabling them to be phosphorylated so they can be added to the + end
They increase the rate of monomer addition

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14
Q

What are the 4 mechanism of actin binding proteins?

A

Branching
- Arp2/3 interacts with existing filaments to induce branching
Capping/severing
- Gelsolin binds to + end to block further growth
Nucleators
- Arp2/3 with profilin initiates fibre growth
Bundling/crosslinking
- Α-actin filamin reinforce the structure to give it strength

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15
Q

What are small GTPases?

A

Small monomeric 21kDa proteins
- Don’t confuse with heterodimeric G proteins
Have intrinsic GTPase activity that can bind to and hydrolyse GTP

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16
Q

What modifications do small GTPases have?

A

Many have post-translational lipid modifications to target them to specific membrane sites

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17
Q

What family do small GTPases belong to?

A

All small GTPases belong to a large family

Archetypcal member being Ras

18
Q

What are some examples of the Ras superfamily of GTPases?

A 
1 Ran - mitotic spindle organisation 
27 Arf - membrane budding 
61 Rab - endosomal trafficking 
36 Ras - cell proliferation and a oncogene 
20 Rho - cytoskeleton and migration
19
Q

If RAC is bound to GTP it is?

A

Active
This is a timed process due to its enzymatic activity
GTP is broken down to GDP fairly quickly

20
Q

If RAC is bound to GDP it is?

A

Inactive

21
Q

What does active RAC activate?

A

Downstream targets - PAK and WAVE2

22
Q

What helps catalyse the exchange of GDP for GTP?

A

Guanine nucleotide exchange factor (GEF)

23
Q

What helps catalyse the breakdown of GTP to GDP?

A

GTPase activating protein (GAP)

24
Q

What slows down the dissociation of GDP?

A

Guanine nucleotide dissociation inhibitor (GDI)

25
Q

What causes a change in the structure of GTPase switch regions?

A

Binding of nucleotide causes a structural change in the switch regions
Bound nucleotide dictates signalling activity
GTP hydrolysis is an intrinsic property of the protein that turns signalling off

26
Q

What do RhoA, Rac1 and Cdc42 coordinate?

A

Coordinate actin cytoskeletal organisation

In turn controls cell morphology, cell movement and cell polarity

27
Q

What does Cdc42 control?

A

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

28
Q

What does Rac1 control?

A

Controls organisation of new actin filaments into dynamic ruffling structures or lamellipodia

29
Q

What does RhoA control?

A

Stabilises actin filaments into a more rigid skeletal framework - stress fibres

30
Q

What have dominant negative and constitutively active mutant GTPases shown?

A

Point mutations in the nucleotide-binding sites of small GTPases can make the protein

  • Constitutively active - always on
  • Dominant negative - always off and inhibitory
31
Q

What does a constitutively active GTPase mutant cause?

A

Substitution of the catalytic glutamine in switch 2 stops GTP hydrolysis
Always GTP bound
Always signalling active

32
Q

What does a dominant negative GTPase mutant cause?

A

Substitution of the P-loop stops nucleotide binding
Nucleotide-free
Mops up active GEFs

33
Q

What part of the cell do Rho mutants affects?

A

The actin cytoskeleton

34
Q

Constitutively active Rho activation leads to?

A

Stress fibre formation

35
Q

Constitutively active Rac activation leads to?

A

Membrane ruffles

36
Q

Constitutively active Cdc42 activation leads to?

A

Filopodia formation

37
Q

What do activated Rho proteins bind to?

A

A specific 16 amino acid sequence in effector proteins

CRIB motif - Cdc42 Rac1 Interactive Binding

38
Q

How does Rac stimulate actin filaments?

A

Activates WAVE proteins
These activate Arp2/3
These create actin filaments

39
Q

How does Cdc42 stimulate actin filaments?

A

Activates WASP proteins
These activate Arp2/3
These create actin filaments

40
Q

How does Rho activate increases in myosin contractility and stress fibre formation?

A

Activates Rho kinase

Activates myosin phosphorylation

Cell and Molecular (24 decks)

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