Tumour Suppressor Genes

Question 1

What is the function of tumour suppressor genes?

Answer 1

To negatively regulate growth by arresting growth or inducing apoptosis.

Question 2

Why are tumour suppressor genes said to be recessive in cancer?

What is this hypothesis known as?

Answer 2

• Because both alleles of a tumour suppressor gene must be lost to lose tumour suppressor properties.
• This is known as Knudson’s two-hit hypothesis.

Question 3

Why does familial tumour suppressor loss (e.g. with familial retinoblastoma) often result in bilateral disease, whereas somatic tumour suppressor loss often results in unilateral disease (e.g. with somatic retinoblastoma)?

Answer 3

• If the tumour suppressor loss in a particular tissue is familial, the individual is born with one mutated tumour suppressor gene, and the other must be lost by one more somatic mutation in order for tumour suppressor function to be lost.
• Since the gene is inherited in both tissues (bilaterally, e.g. in the eyes), familial cancers are more likely to develop bilaterally than unilaterally.
• In contrast, an individual that is not born with one mutated tumour suppressor gene must develop two somatic mutations in order for tumour suppressor function to be lost.

Question 4

Other than developing bilaterally, give another distinguishing characteristic of familial cancer.

Answer 4

Early onset.

Question 5

List 2 signs of retinoblastoma.

Answer 5

1 - White light reflection.

2 - Strabismus (cross-eyed).

Question 6

List 2 treatments of retinoblastoma.

Answer 6

1 - Chemotherapy.

2 - Laser therapy.

Question 7

What type of mutation is the most common cause of familial retinoblastoma?

Answer 7

Single base substitutions.

Question 8

What degree of penetrance is shown by retinoblastoma?

Answer 8

• Most mutations are associated with almost complete penetrance (the autosomal dominant genes).
• Rare alleles show incomplete penetrance.
• Although the inheritance of the mutation is dominant, remember that both genes must be lost in order for tumour suppressor function to be lost.

Question 9

Which gene is affected to cause loss of tumour suppressor function in retinoblastoma and osteosarcoma?

Answer 9

RB1.

Question 10

List 2 ways by which the second mutation can come about in familial and somatic retinoblastomas to cause loss of tumour suppressor function.

Answer 10

1 - There can be a mutation to the second gene, causing hypermethylation of the 5’ region of the nucleotides of the RB1 gene.

2 - There can be loss of heterozygosity, which can be a result of mitotic recombination, mitotic nondisjunction or large deletions.

*The second point here isn’t specific to retinoblastomas.

Question 11

What does the RB1 gene code for?

How does its protein product work as a tumour suppressor protein?

Answer 11

• The RB1 gene codes for pRB.
• pRB must be phosphorylated at the restriction point between G1 and S phase in order for the cell cycle to continue:
• pRB is a transcription factor that represses transcription by repression of E2F transcription factors.
• pRB is also bound to HDAC, which antagonises the transcription-priming function of p300.
• Early phosphorylation is mediated by the cyclin D - cdk4 / 6 complex. Phosphorylated pRB dissociates from HDAC, enabling transcriptional priming by p300.
• Late phosphorylation is mediated by the cyclin E - cdk2 complex. Hyperphosphorylated pRB dissociates from the gene, driving transcription necessary for progression into S phase.

Question 12

Give an example of a treatment for loss of tumour suppressor function.

Answer 12

Cdk inhibitors.

Question 13

What is p16ink4a?

What is its function?

Answer 13

• A tumour suppressor that is expressed in response to cellular stresses that is inactivated in human cancer.
• It prevents cdk4 and cdk6 from phosphorylating pRB, causing growth arrest.

Question 14

Which cancer is associated with a mutation in p16ink4a?

Answer 14

Familial melanoma.

Question 15

List 4 tumour suppressor functions of p53 transcription factor.

Answer 15

1 - Promotion of proapoptotic activity.

2 - Promotion of growth-arresting activity.

3 - Promotion of cellular senescence.

4 - Inhibition of angiogenesis.

Question 16

How is it possible that over 90% of cancers involve loss of p53 function?

Answer 16

Because although p53 itself is not necessarily lost by mutation in 90% of all cancers, its expression is affected by other upstream mediators.

*This reflects the multifactorial process of tumorigenesis.

Question 17

What are Li-Fraumeni and Li-Fraumeni-like syndromes?

Answer 17

• Familial predispositions to cancer that are inherited in the same way as familial retinoblastomas:
• Autosomal dominant inheritance of one mutated tumour suppressor gene, where tumour suppressor function is lost if the second gene undergoes somatic mutation.
• However in the case of Li-Fraumeni and Li-Fraumeni-like syndromes, the gene affected is p53 rather than pRB (which is the cause of familial retinoblastoma).

Question 18

List 6 cancers caused by Li-Fraumeni and Li-Fraumeni-like syndromes.

Answer 18

1 - Soft tissue sarcomas.

2 - Osteosarcomas.

3 - Breast cancer.

4 - Brain tumours.

5 - Leukaemias.

6 - Adrenocortical carcinoma.

Question 19

How do p53 and pRB differ in the way they bring about tumour suppressor function?

Answer 19

To bring about their tumour suppressor functions, pRB suppresses transcription whereas p53 promotes transcription.

Question 20

How many amino acids comprise p53?

Answer 20

393.

Question 21

Describe the structure of p53.

Answer 21

1 - The sequence-specific DNA binding domain facilitates binding of p53 to target genes.

2 - The transactivation domain promotes transcription of target genes.

3 - The proline-rich domain promotes apoptosis.

4 - The oligomerisation domain enables p53 to function as a tetramer with other p53 molecules.

5 - The C-terminal regulatory domain enables post-transcriptional modification of p53.

Question 22

When is p53 activated?

Answer 22

Only in response to cellular stresses such as:

1 - DNA damage.

2 - Oncogene activation.

3 - Hypoxia.

4 - Loss of adhesion.

5 - Changes in ribosomal activity.

6 - Damage by ROS.

Question 23

What is mdm2?

What inhibits its expression?

Answer 23

• Mdm2 is a negative regulator of p53.

- Its expression is inhibited by the tumour suppressor gene p19ARF.

Question 24

List 2 tumour suppressor genes other than p19ARF that are positive regulators of p53.

Answer 24

1 - ATM.

2 - CHK2.

Question 25

List 6 tumour suppressor genes that are upregulated by p53.

Answer 25

Depending on the appropriate cellular response:

• For growth arrest:

1 - p21.

• For apoptosis:

2 - Bax.

3 - Puma.

4 - Fas.

• For anti-angiogenesis:

5 - Maspin.

• For DNA repair:

6 - p48.

Question 26

How does p21 inhibit cellular growth?

Answer 26

It is a cdk inhibitor, preventing phosphorylation of pRB.

Question 27

How is p53 activated in response to DNA damage?

Answer 27

1 - At cell cycle checkpoints, DNA damage causes ATM to autophosphorylate itself.

2 - Phosphorylated ATM phosphorylates mdm2, inactivating it. Remember mdm2 is a negative regulator of p53.

3 - Phosphorylated ATM phosphorylates chk2, activating it. Remember chk2 is a positive regulator of p53.

4 - Phosphorylated ATM phosphorylates p53, activating it directly.

Question 28

Give an example of a way in which p53 produces a tumour suppressor effect by a non-transcriptional mechanism.

Answer 28

p53 can bind to BCL-2, which triggers an apoptotic pathway by stimulating cytochrome c release from mitochondria.

Question 29

How does mdm2 inhibit p53?

Answer 29

• Mdm2 promotes poly-ubiquitylation of p53.

- This marks p53 for degradation by proteasomes.

Question 30

At which stage in tumorigenesis does loss of p53 function occur?

Why might this be?

Answer 30

• Relatively late - between the late adenoma and carcinoma stages.
• This is because p53 functions as a tetramer, which can contain a mix of mutant and non-mutant alleles, so both alleles of p53 must be inactivated for loss of p53 tumour suppression function.

Question 31

At which domain of p53 do most mutations occur?

What type of mutations are most common here?

Answer 31

• At the sequence-specific DNA binding domain.

- These are usually missense mutations.

Question 32

To which cancers do mutations in the BRCA1 and BRCA2 tumour suppressor gene predispose?

Answer 32

1 - Breast carcinoma.

2 - Ovarian tumours.

Question 33

At which domains of BRCA1 and pRB do most mutations occur?

Answer 33

Mutations to BRCA1 and pRB are evenly distributed across the molecules.

Question 34

How does BRCA1 function as a tumour suppressor gene?

Answer 34

It is involved in the DNA damage response by mediating:

1 - Chromatin remodelling.

2 - DNA recombination repair.

3 - Promoting growth arrest.

Question 35

List 3 histological differences between BRCA1-associated tumours and BRCA2-associated tumours.

Answer 35

1 - BRCA1-associated tumours have a higher mitotic rate.

2 - BRCA1-associated tumours have more lymphocyte infiltration.

3 - BRCA1-associated tumours have lower tubule formation.

Question 36

How does BRCA2 function as a tumour suppressor gene?

Answer 36

BRCA2 binds to RAD51, stimulating homologous recombination repair of double strand breaks in DNA.