DNA repair

Question 1

How do bacteria repair thymine dimers produced by UV light.

Answer 1

A complex forms between DNA and a photoreactivating enzyme. This is capable of absorbing UV light at a higher wavelength than DNA. The energy absorbed is transfered in into the DNA which reverses the cyclobutyl ring.

Question 2

How do mammals repair the O6-methylguanine adduct.

Answer 2

Through the action of O6-alkylguanine methyltransferase.

Question 3

How does O6-alkylguanine methyltransferase repair the O6-methylguanine adduct?

Answer 3

It transfers the methyl group from the guanine onto the SH group found in the active site of the enzyme.

Question 4

Why is O6-alkylguanine methyltransferase referred to as a suicide enzyme?

Answer 4

Once the methyl group has been transferred onto the enzyme, the active site is occluded so can no longer catalyze another reaction.

Question 5

Which Mut is responsible for recognising different replication errors during MMR

Answer 5

MutS family members

Question 6

What does MutS dimerise with and what does this complex do?

Answer 6

MutL - the complex scans DNA and detects nicks in the new DNA strand. These are present because DNA ligase is yet to fill these gaps

Question 7

Why is it important that the MutL/MutS complex recognises nicks in the new strand of DNA

Answer 7

So it removes the correct, inappropriate base following replication, not the wild type base that would be found on the complimentary strand.

Question 8

What denotes the MutL/MutS dimer type that forms in response to DNA damage?

Answer 8

The type of DNA damage results in different dimer combinations

Question 9

90% of MMR mutations occur in which MutL and MutS family members?

Answer 9

Msh2 and Mlh1 which are found in a majority of DNA repairing complexes.

Question 10

Which hereditary cancer is associated with MMR mutations?

Answer 10

Hereditary nonpolyposis colon cancer.

Question 11

What mediates the specificity of base excision repair?

Answer 11

DNA glycosylases

Question 12

What is the process of BER?

Answer 12

1) excision of inappropriate base
2) AP-site cleavage
3) Gap filling
4) If gap filling is incorrect, proofreading ensures the correct gap is put in.
5) Ligation of new base

Question 13

When would cells use BER Vs NER?

Answer 13

BER when there is a single, inappropriate base Vs NER when there is a bulky adduct on the DNA e.g a pyrimidine dimer.

Question 14

What is the process of NER

Answer 14

1. Distorted DNA is recognised and a partial opening in the strand is made. (XPC)
2. Formation of the open structure and recognition of the damaged strand (XPB and XPD are the helicases; TFIIH, XPA and RPA also involved)
3. Dual excision by structure specific nucleases (ERCC1-XPF and XPG)\
4. Excision and DNA repair synthesis (DNA pol e and DNA ligase)

Question 15

What is the role of XPB/XPD

Answer 15

Both are helicases which unwind 28-30 bases in length of damaged DNA Vs undamaged DNA. (during NER)

Question 16

Why does Xeroderma pigmentosum cause a high incidence of cancer?

Answer 16

Mutations lie within the global genomic repair pathway. This is involved in repair of all regions of the genome, repair of all types of bulky adducts.

Question 17

What does gloabl genomic repair require?

Answer 17

XPC and all other NER factors except for CSA and CSB.

Question 18

Why does Cockayne syndrome not cause a large increase in cancer incidence?

Answer 18

Because mutations are within the transcription-coupled repair pathway, a protein complex that dislocates stalled transcription machinery. They therefore have normal overall DNA damage repair; but different preferential gene specific repair after UV light damage. This doesn’t require XPC

Question 19

Is homologous recombination error free?

Answer 19

Mostly - it uses the WT sister chromatid as a template for the exact replacement of lost DNA

Question 20

Is NHEJ error free?

Answer 20

No

Question 21

What proteins are required for homologous recombination>

Answer 21

R51, RPA, BRCA1/2

Question 22

What proteins are required for NHEJ

Answer 22

KU70, KU80, DNA-PK, LIG4, XLF, XRCC4

Question 23

How are interstrand crosslinks repaired?

Answer 23

A double strand break is formed at the stalled replication fork (Mus81, Eme1)
Unhooking of the crosslink is performed by XPF and ERCC1
Gap resection and recombination (Rad51)
Resolution of recombination followed by excision of the crosslink and re-synthesis

Question 24

What is the role of TLS polymerase and how does it achieve this>

Answer 24

Damage tolerance, it is capable of misincorporation at the lesion, extending over and past it. Following this DNA polymerase takes back over as normal.

Question 25

Which cell type is capable of repairing most types of DNA lesion>?

Answer 25

Human embryonic stem cells (more effectively than more differentiated cell types)

Question 26

Which cell types are defective in BER?

Answer 26

Monocytes and muscle cells

Question 27

What DNA damaging event induces the activation of ATR?

Answer 27

Stalled or damaged replication forks

Question 28

What DNA damaging event induces the activation of ATM

Answer 28

DNA DSBs