Lecture 26 - Cancer

Question 1

Describe the action of PLCγ

Answer 1

PLCγ binds to EGFR, which activates it by phosphorylation. Acive PLCγ cleaves off the inositol result in a diacyl chain and an inositol (PI3).

Question 2

Describe the action of PI3 (from PLCγ)

Answer 2

It can bind to the activated EGFR, activating its kinase activity. This allows it to phosphorylate PI-4,5-BP to PI-3,4,5-TP, which acts as a second messenger in activating kinases involved in cell growth and survival (PDK1 & PKB)

Question 3

Describe the action of the diacyl chain (from PLCγ)

Answer 3

The diacyl chain can activate protein kinase C to promote cell growth.

Question 4

Is a single pathway sufficient for cell growth?

Answer 4

No, multiple pathways are needed for a significant effect.

Question 5

Describe the structure of Ras.

Answer 5

A membrane bound protein (due to a CAAX motif at C-termini), it has a post-translationally modified cysteine at the 4th amino acid from the C terminus.
A thioacyl group called a farnesyl group is attached to it allowing it to be membrane bound.

Question 6

Describe the mechanisms of Ras.

Answer 6

• Ras is a GTPase and has high affinity for GTP and GDP, very hard to dissociate from either.
• It is inactive if bound to GDP, but active if bound to GTP.
• GTPase action is extremely slow (GTP to GDP)
• A GAP (GTPase activating protein) binds to Ras, increasing its catalysis action, and inactivating it.
• Ras cannot easily dissociate, so its GDP is exchanged with a GTP by a GEF protein.
• Requires both GEF and GAP for normal function.
• Being GTP bound allows it to bind to effector proteins mediating cell growth.

Question 7

What is the consequence of any mutation to Ras?

Answer 7

Any mutation will cease Ras’ function, likely contributing to cancer development.

Question 8

How is Ras switched on?

Answer 8

Ras must be switched on by autophosphorylated EGFR, and is mediated by scaffolding proteins.
-growth factor receptor binding protein (Grb-2 & sos1)
Has one SH2 domain that binds to phosphotyrosine of autophosphorylated EGFRs.
Also has 2 SH3 domains which bind to PXXP motifs of GEF, but requires secondary structures to do so.
This mediates EGFR binding to Ras.

Question 9

Describe the structure of GEF

Answer 9

Has two PXXP motifs and is recruited to the membrane once bound to Grb-2. It then binds to Ras to facilitate GDP replacement, activating it.

Question 10

Describe the effector molecules of Ras

Answer 10

-Raf 1 (MAP kinase kinase kinase)
-MEK (MAP kinase kinase)
-MAP kinase (Erk)
Phosphorylated MAP kinase enters the nucleus and phosphorylates transcription factos.
Begins with Raf 1, the first Ras effector molecule.
Raf 1 then activates MEK.
Activated MEK then MAP kinase, which mediates gene regulation.

Question 11

What is the purpose of KSR?

Answer 11

KSR is a kinase suppressor of Ras, a membrane bound protein. It is targetted to the membrane when bound to its receptor, binding to Ras effector proteins, Raf 1, MEK and MAP kinase.

Question 12

Describe STAT3 in relation to MAP kinase

Answer 12

STAT3 is a MAP kinase substrate, existing in the cytosol as an inactive dimer with two phosphorylation sites.
One is phosphorylated by MAP kinase, and the other by EGFR.
Has an SH2 domain that mediates dimerisation following phosphorylation by EGFR.
Fully activated (phosphorylated & dimerised), induces angiogenesis and cell cycle progression gene transcription.
Enters the nucleus alongside MAP kinase to activate.