Chapter 6

Question 1

What is Knudson’s Two Hit hypothesis?

Answer 1

A gene with a germline mutation predisposes n individual to cancer

Question 2

are oncogene mutations dominant or recessive?

Answer 2

dominant - one active is enough

Question 3

Are Tumor-suppressor genes dominant or recessive?

Answer 3

recessive - two must be activated

Question 4

Name a few well-known tumor-suppressor genes and the syndromes they predispose to

Answer 4

1. RB1 - Retinoblastoma
2. p53 - Li-Fraumeni
3. APC - Colorectal cancer
4. BRCA - breast and ovarian cancer

Question 5

What happens if RB protein is lost?

Answer 5

There will be no inhibition of E2F and cells will progress through the cell cycle

Question 6

What happens if the RB protein suffers a missense mutation?

Answer 6

In a missense mutation, the tertiary structure of the protein will not have the same function. This means that it will not be able to bind to E2F and inhibit the cell cycle progression

Question 7

what is a consequence of increased activity for Cyclin D/cdk4 complex?

Answer 7

Increased activity of this comples will hyperphosphorylate the RB protein and hereby activate binding from E2F

Question 8

Which virus protein inhibits the function of RB protein?

Answer 8

The E7 protein from the HPV virus. It binds to RB and inhibits its function

Question 9

Why is p53 called the guardian of the genome?

Answer 9

This protein is the core in many pathways that regulate cell survival and reaction to DNA damage

Question 10

What is the function of MDM2?

Answer 10

MDM2 binds to p53 and adds an ubiquitine group to it, marking it for degradation

Question 11

Name the 4 domains of the p53 protein and briefly explain their function

Answer 11

1. Transactivation domain and MDM2 binding - MDM2 binds to this domain and ubiquitinates p53
2. DNA binding domain - p53 is a transcription factor for many proteins involved in DNA damage repair and cell cycle arrest. It binds to DNA and regulates their transcription
3. Tetramerization domain - a fully functional p53 is a tetramer. When activates it creates one
4. Regulatory domain - this domain is meant for cofactors to bind to p53. Cofactors, for example, regulate the binding strength to DNA molecules and the rate of gene transcription

Question 12

Briefly explain the p53-MDM2 feedback loop.

Answer 12

When p53 concentration is high in the cell, it binds to DNA and induces the formation of MDM2. The MDM2 then binds to p53 and marks it for degradation. Low concentrations of p53 stop the formation of MDM2, so more p53 can be created, then the cycle starts again

Question 13

At what level is p53 regulated?

Answer 13

At a post-transcriptional level

Question 14

Describe 2 upstream pathways of p53 activation

Answer 14

1. DNA damage > ATM > Chk2 > p53
2. Oncogene active > p14Arf > MDM2 inhibition > inhibition of p53 degradation > more p53
3. Cell stress > ATR > Casein kinase II > p53

Question 15

How does p14 help p53?

Answer 15

p14 binds to MDM2 which inhibits it from binding to p53

Question 16

Describe a few downstream pathways of p53 in DNA damage repair and apoptosis

Answer 16

1. p53 > p21 > p21 binds to cyclin/cdk > Cell cycle arrest
2. p53 > activation of pro-apoptotic proteins like NOXA, PUMA, Death receptor, Bax etc + inhibition of anti-apoptotic factors like Bcl2 > apoptosis via the caspase pathway

Question 17

What does Bcl2 inhibit?

Answer 17

The release of Cytochrome C in the plasma membrane from the mitochondria

Question 18

What is the effect of p53 on miRNA?

Answer 18

p53 activates the transcription of miRNA-34A which is an inhibitor for proteins like Myc, Cyclins/cdk - thus preventing cell cycle progression

Question 19

What determines the decision-making activity of p53?

Answer 19

1. Amount of p53 - high concentrations of p53 will induce its effects
2. Modification (phosphorylation and acetylation)
3. Co-factors at the promoter region
4. Co-activators binding to p53 - co-activators can decide the binding strength of p53 to the DNA or other proteins, hereby regulating its activity

Question 20

What could a difference in DNA binding affinity for p53 mean?

Answer 20

The difference between apoptosis and cell cycle arrest

Question 21

High p53 and low binding strength can induce…

Answer 21

… apoptosis (activates Bax)

Question 22

Low p53 and high binding strength can induce…

Answer 22

… cell cycle arrest (activation of p21)

Question 23

What is the function of co-factors and p53?

Answer 23

Co-factors bind to p53 and can determine the binding strength to a certain promoter

Question 24

What is the function of ASPP?

Answer 24

ASPP is a co-factor of p53 which increases binding strength to pro-apoptotic promoter regions

Question 25

In which domain can you find the most mutations of p53?

Answer 25

In the DNA binding domain. 90% of the mutations happen in that region

Question 26

How can gain-of-function of p53 actually inhibit it?

Answer 26

p53 activates MDM2 in a negative feedback loop. A gain-of-function means higher activity, so less p53 is needed to activate more MDM2. MDM2 degrades p53 and it cannot function

Question 27

Which viral protein inhibits p53?

Answer 27

E6 of the HPV virus. HPV E7 inhibits RB

Question 28

Under what condition can introducing a functional p53 gene cure cancer?

Answer 28

The wild type p53 gene in an individual must be malfunctioning

Question 29

what therapeutic strategy can be used for MDM2?

Answer 29

An MDM2 inhibior can decrease the p53 inhibition in a cancer cell

Question 30

During therapy, why would endogenous, wildtype p53 be inhibited?

Answer 30

If there is a treatment that inhibits the feedback loop of p53, p53 will also be produced in normal body cells. This can lead to apoptosis of healthy cells. A new p53 gene can be induced in the cancer cells that has a different, lower affinity for the drug used. Like that, the p53 in the cancer is not inhibited while normal cells will suffer less.