R7 DNA damage and repair

Question 1

What is a DNA lesion ?

Answer 1

breaks or other chemical changes that affect the structure of the helix, ultimately preventing transcription

Question 2

What are some examples of DNA lesions ?

Answer 2

ss breaks, ds breaks, bulky adduct (covalently linked), interstrand crosslink, base mismatch, base alkylation

Question 2

What effect can UV have on DNA ?

Answer 2

cause thymines to form dimers (bond to one another)

Question 3

What effect can ionising radiation have on DNA ?

Answer 3

ss and ds breaks

Question 4

What effect can alcohol have on DNA ?

Answer 4

interstrand crosslinks

Question 5

What effect can oxidative damage have on DNA ?

Answer 5

oxidised bases, ROS - reactive oxygen species

Question 6

What effect can mechanical stress have on DNA ?

Answer 6

ds breaks during mitsosis

Question 7

What process is the biggest source of DNA lesions

Answer 7

DNA replication

Question 8

How does the unpacking of chromatin affect the chances of lesions ?

Answer 8

Increases, as the proteins in chromatin protect from UV and IR

Question 9

How can DNA polymerase increase the chance of lesions occurring ?

Answer 9

May miss or add nucleotides or mismatch them, damaged nucleotides can be used, ribonucleotides are incorporated or not removed from RNA primers

Question 10

How do unpaired bases affect the chances of lesions occurring ?

Answer 10

they are exposed to bases and ROS

Question 11

How is ssDNA affected by lesions ?

Answer 11

more prone to breakage due to mechanical stress

Question 12

What happens if lesions are not repaired or repaired incorrectly ?

Answer 12

Mutation(s) will occur

Question 13

What type of lesions does the cell recognise ?

Answer 13

DNA free ends (not telomeres), ssDNA accumulation and persistence, base pairing incorrectly, the bases not being A,T,G,C, not having deoxyribose

Question 14

How is an extra nucleotide base repaired by the cell

Answer 14

Nuclease removes the structural damage then polymerase fills the gap then ligase seals the nick

Question 15

How is a thymine dimer repaired by the cell ?

Answer 15

Photolyase (light-activated enzymes) cleaves the bond between thymines during times of high UV

Question 16

How does an extra nucleotide/ mismatch affect the replication process ?

Answer 16

they will be paired up on one strand leading to a mutation, this would be the same for mismatch

Question 17

How may a RNA nucleotide affect the replication process?

Answer 17

cause mismatch in the daughter strand

Question 18

How may an oxidised guanine affect the replication process?

Answer 18

oxo-G can pair with A, when repaired by cut and patch the G will be removed and replaced with a T, leading to a mutation in both strands

Question 19

What is the cut and patch mechanism ?

Answer 19

Nuclease removes a nucleotide, polymerase adds one, ligase seals the nick

Question 20

How can a dsBreak be caused?

Answer 20

broken replication fork, unseperated sister chromatids in mitosis, ionising radiation, some chemicals (chemotherapeutic)

Question 21

How is an ICL caused ?

Answer 21

endogenous metabolites (acetaldehyde), chemicals (chemotherapeutic drugs)

Question 22

How is a dsbreak repaired ?

Answer 22

Ligation of broken ends, homologous recombination (the breakage is compared to a non-broken homologous chromosome and copied)

Question 23

What can ligation of a dsbreak cause ?

Answer 23

addition or deletion at the site of breakages which is error prone

Question 24

What can homologous recombination cause ?

Answer 24

Erroneous repair

Question 25

What is erroneous repair in terms of DSB ?

Answer 25

- telomeres added to a break (causing the loss of the acentromeric arm and a shorter chromosome), - reciprocal translation - the broken end joins to another strand - dicentric chromosomes - another chromosome binds to the broken end, making an unstable double chromosome, the chromosome then has 2 centromeres so the spindle fibres pull at them both, leading to more breakages

Question 26

What is GCR ?

Answer 26

Gross chromosomal rearrangement: - large deletions, large insertions, large inversions, translocation arises from erroneous DSB repair

Question 27

what is aneuploidy ?

Answer 27

more than one abnormal chromosome

Question 28

What is common in cancer cells that may be caused by erroneous dsb repair ?

Answer 28

aneuploidy, GCR

Question 29

How is ICLs repaired ?

Answer 29

they cannot be transcribed as the DNA strands cannot be separated in proliferating cells it may block sister chromatid segregation in mitosis may be converted to DSB then repaired as a DSB sometimes induced in anti-cancerous therapy

Question 30

What is the bacterial SOS response ?

Answer 30

During serious DNA damage, ssDNA accumulates and activates SOS regulons This causes: - replication to continue, cell division inhibitors are activated, filamentous cells with many chromosome copies are made - homologous recombination genes activated - trans lesion synthesis (TLS) DNA polymerase genes activated allowing for mutagenesis (gene mutation in base order). the polymerase uses the damage template strand adding the wrong bases - delayed cell division allows time to repair DNA before dividing

Question 31

What is mutagenesis ?

Answer 31

The change in base order leading to a mutant forming

Question 32

Where are the DNA damage checkpoints in the cell cycle?

Answer 32

G1 - so DSB cannot go into S Intra-S checkpoints - replication fork problems means the late origin is not fired G2 - there is ssDNA, mitosis is then blocked

Question 33

What is the DNA damage response based on ?

Answer 33

PTMs, transcription activation plays a minor role

Question 34

Why is blocking the next stage of the cell cycle important ?

Answer 34

Allows time for repair

Question 35

What two processes must happen before a mutation is acquired ?

Answer 35

lesion formation, escaping repair

Question 36

How can we work on the probability of mutations?

Answer 36

multiple the probability of lesions, and probability of escaping correct repair external stimuli may change the probability, UV increases the chance of lesions

Question 37

How can mutation rates be increased ?

Answer 37

Exposure to high UV, then placed in the dark so photolyase isn't activated

Question 38

what can increase the chance of acquiring lesions ?

Answer 38

- DNA polymerase loses its proofreading activity - Wrong nucleotide incorporated - Helicase loses function, forks break more often - Mitochondria are leaking, more ROS in the nucleus

Question 39

What increases the probability of escaping repair ?

Answer 39

- Mutations lead to higher mutation rates due to higher probability of unrepaired or incorrectly repaired damage. - Mutations in DNA damage checkpoint genes - Mutations in chromosome segregation genes (mitotic spindles)

Question 40

What is somatic mosaicism ?

Answer 40

the acquiring of mutations in somatic cells leading to only some areas being affected - cannot be inherited BRCA is an example of this

Question 41

Why might a replication fork be stalled ?

Answer 41

DNA damage (ss breaks, ds breaks, abbducts), G-quadruplexes (lots of G), fork collision, not enoguh free nucleotides, helicase and polymerase loose function, R-loops (RNA, DNA hybrids), failure for topioisomerase, lack of checkpoints, DNA-binding proteins, crosslinks