Cancer and DNA Repair Pathways Flashcards
1
Q
Genome instability
A
- germ cells
- somatic cells
- T and B cells
2
Q
Germ cell genome instability
A
- meiotic homologous recombination
3
Q
T and B cell genome instability
A
- VDJ recombination
- class switching
4
Q
Somatic cell genome instability
A
- 10^16 mitotic divisions
5
Q
Mutation accumulation is correlated with
A
- cancer
- neurological degeneration
6
Q
Causes of genome instability can be
A
- endogenous
- exogenous
7
Q
Endogenous causes of genome instability
A
- spontaneous (5meC deamination)
- reactive metabolites (aldehydes)
- free radical enzymes (APOBEC, topoisomerases)
- faulty replication
- defective repair
8
Q
Exogenous causes of genome instability
A
- radiation therapy
- sunlight
- carcinogens (tobacco smoke)
- chemotherapy
- retroviral integration
- viral infection (EBV)
9
Q
Structure
A
- genome instability
- Types of DNA damage and genome instability
3.
10
Q
Coping with DNA damage
A
- repair (unique)
- cell cycle checkpoints
- apoptosis
11
Q
apoptotic defects
A
result in mutation accumulation
12
Q
Describe the correlation between genetic instability and cancer
A
- cancer incidence is correlated with age
- somatic cancer genomes are highly mutated
- DNA repair and replication defects are correlated with a cancer predisposition
- cancer karyotypes are abnormal
13
Q
How can we determine which mutational processes are important in cancer?
A
- use mutational signatures
- 6x possible bp substitutions
14
Q
How important is DNAR?
A
approximately 5% of proteins take part in this process
15
Q
How do study DNAR using yeast?
A
- create control and mutagenised plates
- tag DNAR factors with GFP
- IR
- live cell confocal analysis
- study protein recruitment
- gene cloning and complementation analysis can reveal genetic underpinnings
16
Q
DNA repair pathways
A
- direct reversal of DNA damage
- base excision repair
- nucleotide excision repair
- ribonucleotide excision repair
- mismatch repair
- interstrand crosslink repair
- non-homologous end-joining
- homologous recombination
17
Q
Direct reversal of DNA damage
A
- error-free
- limited to a small number of lesions
18
Q
cyclobutane pyrimidine dimers +
A
pyrimidine pyrimidine products - (specific photolyase, light, FADH-)-> DNA
19
Q
cyclobutane pyrimidine dimers and pyrimidine pyrimidine products
A
- UV photoproducts
- covalent bonds form between adjacent bases
20
Q
6meG repair
A
-(alkyltransferase)-> DNA
21
Q
How does the alkyltransferase work in direct reversal of DNA damage?
A
- methyl group transferred to enzyme cysteine
- suicide enzyme (irreversible damage results in degradation)
22
Q
6meG
A
- mutagenic
- can pair with T in replication
23
Q
Base Excision Repair (BER)
A
- designated enzymes to recognise sponataneous damage
- e.g. deamination, methylation, oxidation (uracil, 8-oxoG)
- essential
24
Q
General BER
A
1) DNA glycosylase binds to minor groove
2) kinks DNA @ damage sit
3) cuts glycosidic bond
4) flips out damaged base into an abasic site
5) abasic site processed to make a nick
25
Short Patch BER
1) DNA glycosylase binds to minor groove
2) kinks DNA @ damage sit
3) cuts glycosidic bond
4) flips out damaged base into an abasic site
5) abasic site processed to make a nick
6) nick processed to generate 3'OH
7) abasic nucleotide removed
8) Pol beta replaces damaged nucleotide
9) DNA ligase fills the nick
26
Long Patch BER
1) DNA glycosylase binds to minor groove
2) kinks DNA @ damage sit
3) cuts glycosidic bond
4) flips out damaged base into an abasic site
5) abasic site processed to make a nick
6) nick processed to generate 3'OH
7) abasic nucleotide removed
8) Pol beta synthesises several nucleotides from the 3' OH
9) damaged DNA is displaced as a flap
10) Fen1 cleaves the flap
11) DNA ligase fills the nick
27
NER
Case Study: Xeroderma pigmentosum
28
Xeroderma pigmentosum
- early-onset skin cancer (x2000 increase)
- rare survival to middle age
29
Global genome NER
- detects a wide range of damage
- bulky adducts/double helix destabilising (e.g. UV thymidine dimer)
- non-essential
30
Global genome NER process
1) damage recognition
2) XPC-Rad23b binds to undamaged, complementary strand
3) TFIIH helicase interacts with XPC (separates strand)
4) XPD tracks to lesion
5) XPA, RPA and XPG recruited (makes 5' incision)
6) DNA synthesis replaces damaged strand
8) XPG makes 3' incision (damaged fragment released)
9) ligation completes repair
31
Transcription-coupled NER - the basis
ensures efficient repair of transcribed strand
32
Transcription-coupled NER
- RNA Pol-II recognises damage
- CSB recruited
- CSB-dependent recruitment of NER + synthesis factors (e.g. CSA-cullin complex)
- CSA Ub-ligase complex + UBD of CSB triggers damage excision
- repair synthesis fills the gap
33
ICL repair
-
34
ICL toxicity arises due to
blocked DNA replication and transcription caused by endogenous metabolites (aldehydes) and chemotherapeutics (sulphur, nitrogen mustards)
35
cisplatin is used to treat
- lymphomas
- leukaemias
- sarcomas
- head
- neck
- breast
- cervical
- ovarian
- testicular
36
How does cisplatin work?
1) H2O reacts with cisplatin to form an OH2 bond (displacing Cl)
2) the OH2 reacts with DNA (at G/A N7 position) to form a monoadduct residue
3) a second reaction event displaces the OH2, resulting in interstrand crosslinking
37
ICL repair case study
Fanconi anaemia
38
Fanconi anaemia
- familial aplastic anaemia
- defective ICL repair
- > 20 genes (core FA protein complex)
- short stature
- hypogonadism
- skin pigmentation
- bone marrow failure (unrepaired ICL-induced apoptosis)
- haematological + other cancers (via chromosomal translocation)
39
FA ICL repair pathways
- conserved in vertebrates
- simpler in other eukaryotes
40
ICL Repair:
1) ICL blocks replication fork
2) TRAIP Ub-ligase modifies Mcm helicase
3) endonucleases nick either side of the ICL (breaks fork)
4) TLS polymerase: cut-end resection
5) resected cut ends invade intact strand (forms DHJ)
6) dissolution
7) damaged base removed by BER/NER/HR
41
Give an example of an FA-complex gene
BRCA2
