DNA Damage Repar

Question 1

What are 3 cellular responses to DNA damage?

Answer 1

Repair
Apoptosis
Cell cycle arrest

Question 2

Whats p53, which two things is it particularly important for? What happens when it mutates?

Answer 2

Effector in DNA damage repair, particularly important for senescence (cell cycle arrest) and apoptosis) - mutated in many cancers

Question 3

What are ATM ATR and DNA-PK?

Answer 3

Transducers - all kinases involved in DNA damage response that can phosphorylate etc downstream for either senescence, apoptosis, DNA repair.

Question 4

What happens when DNA damage is found? Starting with sensor…

Answer 4

Proteins sense DNA damage, signal to transducers to phosphorylate downstream to signal effectors to elicit a cellular response e.g. apoptosis, senescence or repair

Question 5

Why do you have cell cycle check points? What would happen if you didnt and a cell underwent mitosis?

Answer 5

Temporary arrest provides time for DNA damage to be repaired. If you didn’t and mitosis occurred, at metaphase you could either have a situation where two chromosomes can’t be pulled apart e.g. if crosslinked, or DNA may break during anaphase.

Question 6

How many check points are there? What is G1 G2 check point for? What is M for?

Answer 6

Check OK to progress to replication all DNA present Check DNA has been replicated before mitosis. Are chromosomes attached to spindle?

Question 7

When would you be most likely to get Senescence and apoptosis following DNA damage?

Answer 7

If there is a lot of DNA damage or damage persists for a long time

Question 8

What is senescence

Answer 8

Cell there but can never reenter cell cycle.

Question 9

Why don’t you want a cell with DNA damage to keep dividing?

Answer 9

Prone to cancer

Question 10

3 stages of BER? Give an example of when BER can be used?

Answer 10

Enzyme removes base
Enzyme repairs where base was
Enzyme fixes break that was formed

e.g. of a base that has been deaminated

Question 11

Which is the most common form of DNA damage that is implicated in cancers?

Answer 11

DSBs

Question 12

What is the difference between BER, mismatch repair and NER?

Answer 12

BER - replaces base - any point in cell cycle
Mismatch - switches base soon after DNA replication error (e.g. error in exonuclease function of DNA polymerase).
NER - replaces small section of DNA and bases around it, polymerases replace

Question 13

What 2 types of DSB repair are there? One type has 3 subtypes, what are they? Which of the two types is best and why?

Answer 13

Non homology directed repair

Homology directed repair

i) Simple annealing
ii) Synthesis dependant
iii) DSB repair - synthesis dependant and doesn’t lose DNA

Homology directed repair better as if done properly less prone to error.

Question 14

What happens in non-homology directed repair(3)?

note: there are 5 major factors/enzymes to remember

Answer 14

• Protein complex Ku70/80 recognises broken ends and brings in other factors
• Factors brought in that bring the ends together and DNA-Pkcs process them to remove lesions (can lose DNA)
• Ligate ends - DNA ligase IV, XLF, XRCC4

Question 15

Which is the easiest way to DSB repair? What is the disadvantage?

Answer 15

Non homology directed

Error prone

Question 16

What happens during simple annealing homology directed repair (1) . What is the advantage? disadvantage?

Answer 16

1) Uses complementary sequences on 2 single strand DNAs to bind together

Error free

Causes deletions (the overhang on each single strand).

Question 17

What happens during synthesis dependant Homology directed repair (2)? When is it best done and why? Why is this better than single strand annealing?

Answer 17

1) Uses other undamaged chromosome as a template
2) Requires DNA polymerase to copy other chromosome

Best at DNA replication as can use identical copy of DNA strand rather than opposite maternal/paternal copy which could have different alleles.

Better than single strand because no loss of genetic information

Question 18

What happens during DSB Repair (Holliday Junction) (4)? Where else do see this kind of recombination?

Answer 18

1) Use other chromosome as a template
2) Both strands replicate with DNA polymerase - D-loop formation
3) DNA interlinked = 4 way Holliday Junction
4) 2 ways you can split chromosomes, either to give you 2 original chromosomes with the copied regions, or can cause cross-over.

Happens in meiosis.

Question 19

Define cancer

Answer 19

Uncontrolled cell growth

Question 20

What are 5 important proteins/substances used for non-homology directed repair

Answer 20

Ku70/80
DNA-PKcs
Ligase IV
IXF
XRCC4

Question 21

Which two types of homologous DNA repair require DNA polymerase?

Answer 21

Synthesis dependent and DSB repair (Holliday Junction)

Question 22

What can cause nucleotide disincorporation?

Answer 22

Defect in exonuclease function of DNA polymerase

Question 23

How can a defect in BER lead to a SSB persisting? What could happen if a SSB persists at DNA replication? What disease is this implicated in?

Answer 23

Defect in BER could prevent ligation back together of a strand that has been broken to remove nucleotide misincorporation. If this single strand break persists until DNA replication, when the strand is replicated it will become a DSB and break away. Implicated in cancers - risk of genomic instability

Question 24

What does DNA damage repair prevent?

Answer 24

Genomic instability and cancer

Question 25

What is it called where thousands of clustered chromosomal rearrangements occur in a single event? When is it seen?

Answer 25

Chromothripsis, can be seen in early tumour development

Question 26

Give syndrome and therefore cancer arises from a defect in mismatch repair? Is this germline or somatic mutation in a gene?

Answer 26

Lynch Syndrome Colorectal Cancer Germline

Question 27

DNA replication stress and DNA damage can both cause ______

Answer 27

Cancer

Question 28

Whats the biggest risk factor for cancer and why?

Answer 28

Age due to mutation and accumulation (added mutation over time due to DNA replication stress and DNA damage)

Question 29

Whats Werner Syndrome and what causes it . Recessive or dominant?

Answer 29

Premature ageing due to Helices problem - DNA replication stress = ageing. Recessive

Question 30

A problem with mismatch repair could cause what 4 mutations?

Answer 30

A-G mismatch T-C mismatch Insertion Deletion

Question 31

A problem with BER repair could cause what 4 mutations?

Answer 31

Uracil accidentally in DNA 8-Oxyguanine mutations SSBs Abasic site

Question 32

A problem with NER repair could cause what 2 mutations?

Answer 32

Bulky adducts | Pyrimidine dimers

Question 33

A problem with Homology/recombinant DNA repair could cause what 2 mutations?

Answer 33

DSBs | Interstrand linkages

Question 34

What is crucial to locate DNA damage? Name one

Answer 34

Sensors - DNA-PKs

Question 35

what to ATM ATR and DNA-PKcs do?

Answer 35

Kinases phosphorylate downstream to cause either cell cycle arrest (and repair), senescence or apoptosis

Question 36

what is p53 an example of?

Answer 36

An effector protein - acts especially in apoptosis/senescence

Question 37

Name 4 things a transducer can do to downstream targets (1 is phosphorylation)

Answer 37

2) Ubiquitination 3) Sumoylation 4) Acetylation

Question 38

5 enzymes used in BER

Answer 38

``` Glycosylase Endonuclease Deoxyribophosphodiesterase DNA polymerase Ligase ```

Question 39

4 enzymes used in mismatch

Answer 39

Endonuclease Exonuclease DNA Polymerase Ligase

Question 40

Whats tumour heterogeneity and what's it's affect on treatment

Answer 40

Subclones of cells within a tumour. Some clones may have mutated and become resistant to treatment, leads to a tumour with different cell types that is able to proliferate.

Question 41

State a problem with current cancer treatments

Answer 41

Cause DNA damage so promote tumour evolution and treatment resistance.