3: Mechanisms Of DNA Repar II: Double Stranded Break Repair

Question 1

What is the consequence if single strand DNA breaks go unrepaired?

Answer 1

Can block DNA replication, stall transcription, turn into double strand breaks

Question 2

Bulletpoint the steps of Single Strand Break Reapir (SSBR)

Answer 2

1. Detection of SSBs
2. Removal of blocking groups from 5’ and 3’ ends, making ends compatible
3. Resynthesis
4. Resealing

Question 3

What are the two key proteins in single strand break repair?

Answer 3

PARP1 - poly(ADP-ribose) polymerase 1
XRCC1 - x-ray cross complementing protein 1

Question 4

What is the function of XRCC1

Answer 4

It’s a scaffold protein, it interacts with repair proteins to coordinate repair process ensuring correct proteins are in place at the correct time

Question 5

List some proteins that XRCC1 may interact with

Answer 5

• glycosylases
• endonucleases
• polymerases
• ligases
• PARP
• etc……

Question 6

Which one out of the 2 key proteins in SSBR has enzymatic activity

Answer 6

PARP1
(XRCC1 lacks enzymatic activity)

Question 7

Briefly outline the function of PARP1

Answer 7

• cleaves NAD+ to ADP-ribose and nicotinamide
• attaches ADP-ribose to an acceptor protein/substrate
• additional NAD+ molecules can be cleared and build linear/branched form of PAR

Question 8

What does PARP1 do

Answer 8

Recognises strand breaks and functions to post-translationally modify repair factors & chromatin surrounding the break

Question 9

What are the proteins involved at the steps in SSBR

Answer 9

1. Detection: PARP
2. Assembly of repair complex, including XRCC1
3. End processing: APE/DNA polyB
4. Gap filling: polyB
5. Ligation: Ligase 3

Question 10

What is the consequence if double stranded DNA breaks go unrepaired?

Answer 10

Loss of genetic material

Question 11

What are the 4 double strand break repair pathways

Answer 11

1. cNHEJ: classic Non-Homologous End Joining
2. Alt-EJ: Alternative(/Microhomology Mediated) End Joining
3. HRR: Homologous Recombinaion Repair
4. SSA: Single Strand Annealing

Question 12

When is cNHEJ employed?

Answer 12

If there are no available homologous sequences

Question 13

Which double strand repair pathway is employed if there are micro homologous sequences at the break ends?

Answer 13

Alternative / microhomology mediated end joining

Question 14

Which is the only error-free DSBR pathway?

Answer 14

HRR: homologous recombination repair

Question 15

What does HRR require to be occur?

Answer 15

A homologous donor sequence o copy the lost information from at the DSB site

Question 16

When does HRR occur?

Answer 16

In S and G2 phases when sister chromatid is available

Question 17

When does single strand annealing occur?

Answer 17

When a DSB is made between 2 repeated sequences

Question 18

Which DSBRs use end resection?

Answer 18

HDR, SSA, Alt-NHEJ

Question 19

Which DSBR uses end protection?

Answer 19

C-NHEJ

Question 20

What regions must be present to use the Single-Strand Annealing versus Alt-NHEJ repair pathways?

Answer 20

• SSA: substantial repeated region
• Alt-NHEJ: Micro region of homology

Question 21

What molecules are involved in the non-homologous end-joining repair pathway?

Answer 21

• Ku: mediates repair pathway, recruits catalytic subunit
• Artemis: end processing

Question 22

Is the NHEJ repair pathway prone to errors ad why?

Answer 22

It will always have errors as it cleaves, not repairs.
This is only significant in coding areas

Question 23

Briefly outline the NHEJ repair pathway

Answer 23

• Recognition of DNA damage
• Bind of NHEJ proteins, including Ku which serves as a scaffold to recruit other repair proteins to assemble at site of damage
• End processing via Artemis
• Ligation via Ligase and associated factors.

Question 24

RAD51 is exclusive to which DSBR pathway?

Answer 24

Homologous Recombination Repair