Microbiology 8: Actin

Question 1

Structure of actin filament?

Answer 1

Comprised of actin protein

4 muscle specific isoforms
- alpha-actins

2 non-muscle isoforms

• beta-actin - cell cortex, leading edge of migrating cells
• gamma-actin - stress fibres

Globular (G) and filamentous (F) actin

Question 2

ATP actin and ADP actin are found at which ends of actin filaments?

Answer 2

ADP-actin at - end
ADP-Pi-actin in middle
ATP-actin at + end

Question 3

Describe function and process of ‘treadmilling’

Answer 3

New sub-units added to + end
Old sub-units lost at - end

Allows rapid reorganisation of molecules in response to stimuli

Question 4

Cytochalasin D mechanism?

Answer 4

binds to and blocks + end, preventing addition of further subunits

Question 5

Toxin Cytochalasin D mechanism?

Answer 5

binds to and blocks + end, preventing addition of further subunits

Question 6

Process of actin nucleation

• where does it occur
• process
• proteins involved
• 2 types of actin filament

Answer 6

Occurs at the cell cortex below the plasma membrane

Two types of filament can be formed (each with own role)

• branched
• unbranched

Requires activation of small GTPase in Rho family of proteins

• Rho, Rac and Cdc42
• activated at plasma membrane by a receptor

Different accessory proteins involved in each type

Question 7

Formation of unbranched actin filaments

Answer 7

• Nucleated by formins
• activated by binding to Rho-GTP
• formins recruit actin monomers to the growing filament + end

Question 8

Formation of branched actin filaments

Answer 8

Activated by Rac and Cdc42 GTPases

Requires two accessory protein complexes

• Arp2/Arp3 complex (Actin Related Proteins)
• 45% identical to actin
• Binds to and creates branches om existing filaments

Formins -> recruit monomers
WASP activated by Cdc42
or
WAVE activated by Rac

Question 9

Function of cross-linking proteins? Examples? Where are they found?

Answer 9

Mould actin filaments into bundles or networks/gels

Bundling proteins create parallel arrays

• Fimbrin (Microvilli)
• alpha-actinin (Stress fibres)

Gel-forming proteins create looser meshworks

• Spectrin (Cell cortex)
• Filamin (Leading edge, stress fibres)

Question 10

Function of integrins? What type of protein are they?

Answer 10

Integrins are membrane-spanning proteins

• > are adaptor proteins
• directly connect the actin cytoskeleton to the extracellular matrix
• important for cell adhesion and migration

Question 11

Main functions of actin based structures? Examples?

Answer 11

Major determinants of cell shape and movement

• Microvilli
• Filopodia
• Lamellipodium
• Cell cortex
• Leading edge
• Stress fibres
• Contractile ring

Question 12

Which actin structures mediate cell migration? What do they do?

Answer 12

Filopodia
- Fine membrane projections which act as sensors to co-ordinate the direction of movement

Lamellipodium
- A broad membrane extension that pushes the front of the cell forward

Stress fibres
- Conractile bundles that ‘squeeze’ the rear of the cell forward

Question 13

Which proteins act in concert to co-ordinate cell migration - mechanisms and structures involved?

Answer 13

Rho, Rac, Cdc42

Localised activation of each

-> Front of cell cdc42 activated
- Microtubule rearrangement and cell polarisation, filopodia formation via WAVE
- Rac activation, leading to lamellipodium formation via WASP
(all branched actin)

-> Rear of cell Rho activated
- formin activation and stress fibre formation
(all unbranched actin)