Ras signalling

Question 1

What are Ras proteins?

Answer 1

• Small GTPases which act as molecular switches
• 3 major isoforms: H, K and N Ras
• Regulate survival, differentiation, proliferation and apoptosis

Question 2

How are Ras proteins activated?

Answer 2

• Active in GTP bound state (vs GDP)
• regulated by catalysts GEF and GAP
• Allow fine tuning for length of time pathway is switched ‘on’

Question 3

What is the role of tyrosine kinase receptors in Ras signalling?

Answer 3

• Binding of growth factors leads to cross-phosphorylation of trk, recruits Grb2 to membrane which activates sos1 (GEF) and then activates Ras-GTP

Question 4

What two main pathways does Ras-signalling activate?

Answer 4

PI3K - cell survival, anti-apoptotic

MAPK - classical pathway, cell proliferation and differentiation

Question 5

What is the difference between Ras isoforms?

Answer 5

Show sequence homology apart from at hyper variable region which regulates binding to membrane

Question 6

What are post-translational modifications does ras undergo?

Answer 6

• farnesylation of CAAX box cleaves 3 amino acids allows for weak binding to membrane
  K ras - poly-cysteine patch
  M ras - palmitolated cycsteine
  H ras - 2 x palmitolated cycsteine

Question 7

What is the function of palmitoylation?

Answer 7

In Golgi results in Ras being trafficked by vesicles to the plasma membrane (when this is prevented H-Ras restricted to Golgi complex, also when only mono palmitoylated)

Question 8

How do membrane microdomains interacr with Ras?

Answer 8

• Ras isoforms differentially associate with cholesrerol rich and absent domains as well as GTP bound (cholesterol independent)/non GTP bound (cholesterol rich)
• Regulate effectors which may interact with Ras
• K ras exclusively occurs at cholestetol independent

Question 9

Which Ras proteins are responsible for endomembrane signalling?

Answer 9

• N-Ras is essential for mitochondril morphology

- K-Ras occurs exclusively at plasma membranes

Question 10

What is the MAPK signalling pathway?

Answer 10

Sequential series of phosphorylation events in response to a growth factor
Ras -> Raf -> MEK1/2 -> ERK1/2 -> transcription factor

Question 11

How does a single stimulus generate a specific response?

Answer 11

Use of scaffoling proteins which bring components of a signalling pathway together for a rapid response

Question 12

What role does scaffolding protein play in the Ras/MAPK pathway?

Answer 12

Scaffold: kinase suppressor of Ras (KSR)

• MEK constituitively bound to KSR, Raf and ERK recruited upon receptor binding
• G protein coupled receptors interact with scaffold beta-arrestin which can also recruit Ras, MEK and ERK

Question 13

Give an example of temporal regulation

Answer 13

Epidermal growth factor only exists for a few minutes while neuronal growth factor is present for hours, therefore length of signalling mediates efector proteins involved
EGF: proliferation
NGF: differentiation