LECTURE 14 - RAS AND MAPK SIGNALLING

simply describe Ras

a ubiquitous gene expressed in every animal cell

what are the 3 Ras genes found in humans

HRas

KRas

NRas

what must Ras be attached to to be functional

it must be stably associated with the membrane

describe Ras C-terminus structure

it is covalently modified with a lipid anchor and contains a CAAX box motif

what is in a CAAX box motif

C - cysteine

A - aliphatic amino acid

X - any amino acid

what is Ras also called

P21

in what state is Ras active and inactive

active - GTP bound state

inactive - GDP bound state

which molecules causes Ras activation, and how

Ras-GEF will exchange GDP for GTP, activating the Ras

what does GAP stand for

GTPase activating protein

which molecules cause Ras inactivation, and how

Ras-GAP initiates hydrolysis of GTP bound to Ras, inactivating it

what does Ras induce

cell growth, differentiation and survival

what is the connection of Ras to cancer

Ras pathway mutations are frequently involved in cancer

what sort of gene is Ras

a proto-oncogene

how many mutations can make Ras cancerous

1 mutation

what does G12V oncogenic mutation cause in Ras pathway

it will reduce the activity of Ras’GTPase, hence increasing Ras GTP, giving rise to cellular “immortality” by placing cell in hyperproliferative and survival state

which Ras pathway component is often overexpressed in tumours

Ras-GEF, increasing the presence of Ras-GTP

what sort of gene/molecule is Ras-GAP

a tumour suppressor

how many mutations must occur in Ras-GAP to have cancerous effects

2 mutations, a loss of heterozygosity

explain how Ras is activated by a RTK

• ligand binds to RTK, causing dimerization and autophosphorylation

• Grb adapter proteins bind to RTK, and Ras- GEF binds to the adapter

• Ras associates with the membrane

• Ras is activated by Ras-GEF

• Ras signals for effectors

what effectors does Ras signal for

PI3K

Rac GTPase

Raf-1(MAPKKK)

explain PI3K downstream signalling evetns

• Ras activates a specific PI3K isoform; P110γ

• Ras bind to P110γ

• PIP3 is produced by active Ras bound to the PI3 kinase, and this is integrated with other cell signalling aspects

explain Rac GTPase downstream signalling events

• Ras activates Rac GTPase

• Ras recruits Rac - GEF

• Rac - GEF exchanges GDP for for GTP on Rac, activating it

• Rac-GTPase forms/maintains lamellipodia (actin formation, microtubule growth) and this enables cell migration

simple describe the MAPKKK cascade

a 3 sequential kinase cascade

what does MAP stand for

mitogen activated protein

define a mitogen

a factor that induces cell growth and division

what is Raf a component of

Raf is a component of the MAP cascade

explain how the MAPKKK cascade works

• MAPKKK activity is induced upon binding to Ras

• MAPKKK phosphorylates and activates MAPKK (MEK)

• MAPKK phosphorylates and activates MAPK (ERK)

• MAPK must be phosphorylated for it to eb released from the protein that retains it in the cytoplasm

• MAPK phopshorylates and activates Elk1 transcription factor

• Elk1 interacts with serum response factor (SRF) to promote transcritpion of target genes

what are the target genes of Elk1

C-Fos

C-Jun

C-Myc

what are the functions of Elk1 target genes

these genes turn on the cell division cycle, transcribing for factors such as cyclins

what type of genes are the Elk1 target genes

as a group they are proto-oncogenes

explain how AP2 is involved in clathrin coated pit formation

• AP2 binds to PtdIns-4,5-P2, and the cytoplasmic domain of RTK

• the AP2 recruits clathrin to form clathrin coated vesicles

• vesicle is taken in by endocytosis