Cell cycle tutorial

Question 1

How does activation of PI-3-kinase pathway occur

Answer 1

Binding of growth factor to EDGR/insulin R stimulates the Ras/Erk pathway to lead to proliferation but also

Growth factor binding activates PIK-3. This molecule then phosphrlyates PIP2–>PIP3. PIP3 activates PDK1, which activates Akt/PKB

Akt/PKB promote cell survival

Question 2

What is the action of PKB/Akt

Answer 2

CELL SURVIVAL

1. Phosphorylates Bad, inactivating it (Bad inactivates antipoptotic regulators Bcl-2 and Bcl-XL)
2. Phosphorylates Caspase 9, inactivating it (key apoptosis protease)
3. Inactivates FOXO transcription factors (FOXO TFs transcribe CKIs e.g p27Kip, and Fas, the apoptotic death ligand)
4. Other, e.g. stimulates protein synthesis

Question 3

How do cell cycle inhibitors like Cip/Kip and INK4 control cdk activity

Answer 3

INK4.. inactivated Cdk4, displacing cyclin D from it in G1

Unphosphorylated p27Kip1 complexes with cyclin-Cdk4 are active, promoting cell cycle progression. BUT,when p27kip1 is phosphorylated, promoted by GSK3b, do not.

GSK3b is inhibited by Akt/PKB.

So p27Kip1 phosphorylated and inactivated by GSK3b, which is inhibited by Akt/PKB, so PI3 kinase (which produces the Akt) maintains active cyclin D-Cdk4

Question 4

Which molecule inhibits the PI-3-kinase pathway

Answer 4

PTEN (which is a PIP3 phosphatase)

Question 5

How do cells check for DNA damage

Answer 5

DNA damage (such as double strand breaks) lead to p53 activation.

Question 6

Which molecules can activate p53

Answer 6

MAP kinase, ATR, ATM (these are recruited and activated by DNA damage like doiuble strand breals)

Question 7

How does p53 interfere with the cell cycle

Answer 7

Phosphorylation of p53 by e.g. MAP kinase, will activate it such that it dissociates from mdm2 and forms a dimer. This will then bind to regulator region of the p21 gene.

p21 is a CKI, once bound, it inactivates G1/S-Cdk nad S-Cdk, to hold the cell in G1/s regulation point, to allow DNA repair proteins time to fix the damage.

Question 8

What are the effects of p53 on the cell

Answer 8

Activate DNA repair proteins

Induce growth arrest by holding cell at G1/s regulation point

Initiate apoptosis if DNA damage is irreparable

Question 9

Give examples of types of proteins controlling the cell cycle that are mutated in cancers

Answer 9

ONCOGENES

EGFR/HER2…. mutationally activated or overexpressed in lots of breast cancers

Ras… mutationally activated in many cancers (V12 Ras… glycine–> valine, L61 Ras… glutatmine –>leucine

Cyclin D1… overexpressed in 50% of breast cancers

B-Raf, mutationally active in menalomas

c-Myc… overexpressed in many tumours

TUMOUR SUPRESSORS:

Rb, inactivated in many cnacers

p53 (mutates in 50% of human cancers)

PTEN (activates the PKB/AKt pathway, promotes cell survival… mutated in advanced cancers)

p27KIP1, underexpression correlated with poor prognosis in many malignancies

Question 10

Outline good drug targets

Answer 10

Cell surface proteins e.g. growth factor antagonists (e.g. Hercpetin which inhibites HER2)

Enzymes e.g. kinase inhibitors

Post-translationl modification e.g. inhibitor of Ras lipid modification which attaches it to the plasma membrane