Cell Cycle 9

Question 1

Where does MDM2 bind on p53?

Answer 1

In the transactivation domain - blocks the recruitment of other TF.

Question 2

What happens when MDM2 binds to p53?

Answer 2

It acts as ubiquitin ligase and tags it for degradation by the proteasome.

Question 3

Describe the MDM2-p53 feedback loop.

Answer 3

At high levels of p53, it will switch on MDM2 expression.

Question 4

What does ARF do?

Answer 4

Binds to MDM2, sequestering it and preventing it from degrading p53.

Question 5

What does ARF stand for?

Answer 5

Alternative reading frame

Question 6

What protein is also transcribed from the ARF gene region?

Answer 6

p16

Question 7

What exons is p16 made up of?

Answer 7

Exon 1a, 2 and 3

Question 8

What exons is ARF made up of?

Answer 8

Exon 1B, 2 and 3

Question 9

Why do ARF and p16 have no sequence homology?

Answer 9

They use different reading frames.

Question 10

Do ARF and p16 both respond to oncogenic activation by Ras?

Answer 10

Yes

Question 11

What type of radiation causes single stranded breaks?

Answer 11

UV

Question 12

What type of radiation causes double stranded breaks?

Answer 12

Ionising

Question 13

Which proteins deal with single and double stranded breaks?

Answer 13

ATR - single

ATM - double

Question 14

What type of enzymes are ATR and ATM?

Answer 14

Serine/threonine kinase

Question 15

Briefly describe 3 ways ATM is activated by DNA damage.

Answer 15

1. Association with chromatin, causes trans-auto-phosphorylation between dimer, when dsDNA break occurs, activating monomers.
2. MRN binds to dsDNA breaks, this undergoes conformational change - causing trans-auto-phosphorylation of dimer –> activated monomers.
3. Oxidative damage causes disulphide bonds to form between monomers, resulting in transautophosphorylated –> active dimer.

Question 16

Which of the mechanisms of ATM activation is most likely correct?

Answer 16

They are all likely to be correct.

Question 17

What are the downstream effects of ATM?

Answer 17

Phosphorylates Ser15 on p53 (insufficient to activate) and phosphorylates Chk2.
Chk2 phosphorylates Ser20 on p53 - activating it.

Question 18

Name 4 things that can affect the decision for arrest/apoptosis in p53 signalling.

Answer 18

1. Strength of signal.
2. Promoter affinity.
3. PTMs
4. Other regulators.

Question 19

Give an example of a regulator that effects p53 downstream pathway.

Answer 19

ASPP2 causes p53 to bind to the Bax promoter and activate apoptosis.

Question 20

Name two things that are involved in pushing the equilibrium of p53 degradation in favour of MDM2.

Answer 20

1. Wip1 - dephosphorylates p53 in the MDM2 binding site.

2. Akt - phosphorylates MDM2, making it more efficient at p53 degradation.

Question 21

Name two other proteins in the p53 family?

Answer 21

p63 and p73

Question 22

Where is the second promoter in p63 and p73?

Answer 22

Between exon 3 and 4

Question 23

Why can the truncated forms of p63 and p73 inhibit transcription/dampen p53 response?

Answer 23

They can occupy promoters, but have no transactivation domain, as this is in exon 1-3.

Question 24

Are p63/p73 often mutated in cancer?

Answer 24

No

Question 25

Describe the function of the drug, Nutlin.

Answer 25

Nutlin binds to MDM2 and prevents degradation of p53 - WT p53 re-activation.

Question 26

Describe the function of the drug, NSC319726.

Answer 26

Binds to the structural p53 mutant R175H (DBD) and reactivate it.