Steve Ward

Question 1

Cell migration

Answer 1

A fundamental and critical function that requires the coordination of several cellular processes that operate in a cycle
A highly integrated, multi-step process

Question 2

Roles of cell migration

Answer 2

Wound healing (innate and adaptive immune response)
Regeneration
Development
Tumour metastases and angiogenesis

Question 3

Chemokinesis

Answer 3

Migration driven by soluble chemokines, without a cue gradient to provide a directional bias

Question 4

Haptokinesis

Answer 4

Migration along a surface, using immobilised ligands e.g. chemokines/integrins, without a cue gradient to provide a directional bias

Question 5

Chemotaxis

Answer 5

Migration driven by a gradient of soluble chemokines, occurring when there is an asymmetry in chemoattractant (i.e. in its rate/density) and local cue gradients can be followed

Question 6

Haptotaxis

Answer 6

Migration along a surface, guided by a gradient of immobilised chemoattractants/adhesion receptor ligands, providing a directional bias

Question 7

Roles of chemotaxis

Answer 7

Immune cell development, immunosurveillance, activation
Sourcing of nutrients (by prokaryotes)
Metastatic cancer migration to growth factors
Embryonic cell development
Formation of multi-cellular structures in protozoa

Question 8

Amoeboid chemotaxis

Answer 8

Consists of 3 key components, each with overlapping but distinct properties

1. Pseudopodia formation
2. Polarisation
3. Directional sensing

Question 9

Pseudopodia formation in amoeboid chemotaxis

Answer 9

Regulated by the chemoattractant
G-protein independent
Enhanced by PIP3

Question 10

Polarisation in amoeboid chemotaxis

Answer 10

Elongation of the cell shape
Cells ‘turn’ towards the gradient because the anterior is more sensitive than the posterior
G-protein dependent
Enhanced by PIP3

Question 11

Directional sensing in amoeboid chemotaxis

Answer 11

Occurs even in immobilised cells
Proteins containing PH domains accumulate at the leading edge of the membrane
G-protein dependent
PIP3 independent

Question 12

Mediators involved in establishing and maintaining cell polarity in response to extracellular stimuli

Answer 12

RhoGTPases
PI3Ks
Integrins
Microtubules
Vesicular transport

Question 13

Cdc42

Answer 13

Rho G protein

“Master regulator” of cell polarity

Question 14

How does Cdc42 influence polarity?

Answer 14

Restricts where lamellipodia form

Localises Golgi and MTOC in front of the nucleus, oriented towards the leading edge of the cell

Question 15

How does Cdc42 exert its effects on MTOC?

Answer 15

Through PAR6 and aPKC, leading to activation of dyne motors that “pull” the MTOC into its new position

Question 16

What does the polarisation of PIP3/PIP2 in response to cellular exposure chemoattractants involve?

Answer 16

The localised accumulation of PI3Ks and PTEN, which generate/remove PIP3/PIP2
PI3Ks rapidly accumulate at the leading edge of cells in response to chemoattractant, whereas PTEN is restricted to the sides/rear, restricting protrusions to the front of the cell

Question 17

Initial experiments

Answer 17

Cells with altered PI3K/PTEN activity can usually still migrate but exhibit significantly reduced directionality along a chemoattractant gradient

Question 18

More recent experiments

Answer 18

PI3K/PTEN activity are dispensable for chemotaxis

Question 19

Hoeller and Kay, 2007

Answer 19

Multiple KO strain lacking all 5 Dictyostelium class I pi3k genes as well as PTEN was still able to undergo chemotaxis in strong chemoattractant gradients, but showed reduced speed
Therefore showing that the polarisation of membrane PIP3 is not essential for directed chemotaxis but ensures rapid movement

Question 20

How is activation of the actin polymerisation machinery achieved?

Answer 20

Through activation of Rac GEFs

Actin polymerisation is required for protrusions

Question 21

Rac positive feedback loops for maintaining directional protrusions

Answer 21

Stimulates recruitment/activation of PI3K

Microtubule polymerisation activates Rac and Rac, in turn, stabilises microtubules

Question 22

WASP/WAVE proteins

Answer 22

Targets of Rac and Cdc42 - associate with WASP/WAVE proteins, that can then bind to Arp2/3 complexes via their WCA domains, promoting Arp2/3 activation

Question 23

Arp2/3 complex

Answer 23

Binds to pre-existing actin filaments at 70 degrees, leading to the nucleation of new actin filaments that branch off in all directions
This results in the formation of a dense web/branched network of actin filaments

Question 24

Actin polymerisation is regulated by…

Answer 24

…accessory proteins that control filament length and stability

Question 25

Profilin

Answer 25

Facilitates growth of the actin filament through (+) end elongation
Profilin binds to the (+) end of actin monomers, leading the (-) end free to bind to the (+) end of the growing actin polymer
Controlling the local concentration of profiling can lead to explosive polymer growth in the required areas of the cell

Question 26

Cofilin

Answer 26

Destabilises the actin filament helix through binding and inducing torsional strain, leading to depolymerisation
Selectively binds to ADP-actin (most new actin in growing polymers is ATP-actin), therefore selectively destabilising older polymers

Question 27

CapZ

Answer 27

Capping protein
Binds to ("caps") the (+) end of actin filaments, favouring their disassembly

Question 28

Tropomodulin

Answer 28

Capping protein
Binds to ("caps") the (-) end of actin filaments, favouring growth

Question 29

How are protrusions stabilised?

Answer 29

Through the formation of adhesions (Rac- and Cdc42-dependent)
Adhesions stabilise lamellipodia through mediating attachment to the ECM