2nd Feb - DNA Damage and Replication Stress responses Flashcards
What is a mutation?
Changes to the coding sequence of proteins
What is genetic instability?
When cells lose the ability to recognise damaged DNA
What is Microsatellite Instability?
Small subtle base changes leading to point mutations
What is chromosome instability?
Continous large scale loss or gain of whole chromosomes or parts of chromosomes. Associated with errors in chromosome segregation
What are chromosome translocations?
Breakage/fusion of chromosomes creating novel, uncontrolled genes. May be caused by eroded telomeres
What is copy number variation?
Loss or gain of multiple copies of genes in a particular region
What are the four main types of DNA repair and what do they repair?
Mismatch repair - errors in DNA replication
Nucleotide excision repair - environmental carcinogens
Base excision repair - damage due to cellular metabolism
Double strand break repair - repair caused by radiation
Outline the process of mismatch repair
- Error in newly made strand
- Binding of mismatch proofreading proteins (MSH1 and MLH1)
- DNA scanning detects nick in the new DNA strand
- Strand removed at the nick
- DNA synthesis to repair
Outline the process of nucleotide excision repair
- A helix distorting adduct is present
- Cleavage of the DNA fragment
- Filling in the fragment by pol delta or pol epsilon and processivity factor for DNAP delta (PCNA) and RPA
Outline the process of base excision repair
- Chemically altered base causes little helix distortion
- The base is cleaved away
- Deoxyribosephosphate cleaved away
- NT inserted and ligated by pol beta and DNA ligase
What are the two main forms of double strand break repair?
Homologous recombination based repiar
Non-homologous end joining
Outline the process of homologous recombination (HR) repair
- 2 sister chromatids
- 1 chromatid undergoes damage causing a double strand break - MRN recognises this
- Section removed
- Joint molecule is formed through Rad51, Rad52 and Rad 54
- DNAP and ligase fills the gaps
Outline the process of non-homologous end joining
- Ds DNA break
- Ku and DNAPkca recognise the break
- Ends are processed by Ku70/Ku80 and DNAPkca
- Stuck back together
Why is non-homologous end joining error prone?
Nucleotides are normally lost during the cut
What enzymes are required for HR?
Rad51 Rad 52 Rad 54 MRN complex BRCA1 BRCA2
What causes heriditary non-polyposis colorectal cancer (HNPCC)
Mutations in hMSH or hMLH (human mismatch repair enzymes) which are required to correct errors in DNA replication
What causes Xeroderma pigmentosa (XP)?
Mutations in the nucleotide excision repair enzymes (NER) lead to high susceptibility to skin cancers due to UV exposure
What causes Bloom’s and Werner’s syndrome?
Mutations in the RecQ family of DNA helicases required for DNA replication and repair –> premature ageing and high susceptibility to a wide variety of cancers
What are the effects of the DNA damage response?
RNA processing Replicase stability Apoptosis Transcription Cell cycle Energy Chromatin remodelling Repair
Why must cell cycle checkpoints arrest the cell very fast?
As DNA replication occurs rapidly
What is the main function of cell cycle arrest?
To allow the DNA repair enzymes time to repair the DNA
What are the key checkpoint mutations?
ATM and ATR
Chk1 and Chk2
p53
cdc25
What are ATM and ATR?
PI3K like enzymes (S/T kinase) that sense ds breaks
What are Chk1 and Chk2?
S/T kinases that are activated by ATM/ATR
What is cdc25?
The rate limiting enzyme in the cell cycle and target of checkpoint kinases
Outline the process of inhibition of the G1/S transition
- Activation of ATM and ATR
- Phosphorylation of p53, Mdm2 and Chk2
- Stabilisation of p53
- Increased p21 expression
- Cdc25 phosphorylation and degradation
- Failure to activate G1/S Cdks
Outline the process of inhibition of G2/M transition
- Activation of ATM and ATR
- Recruitment of BRCA1 and partners
- Phosphorylation of Chk1 and 2
- Block to Cdc25 activation in various pathways
- Failure to activate G2/M Cdks
What triggers the replication stress response?
Transcription and/or RNA hybrids
DNA lesions
Limiting Nts
DNA secondary structure
What is the major mediator of the replication stress response?
ATR
What is the cause of Li-fraumeni syndrome?
Mutations in p53 and Chk2
What is the cause of Ataxia-telangiectasia?
Mutations in ATM
What is the cause of AT like disorder (ATLD)?
Mutations in Mrell
What is the cause of familial breast and ovarian cancer?
Mutations in BRCA1 and BRCA2
What is the cause of Nijmegen breakage syndrome?
Mutations in Nbs1
What is the cause of Foncani’s Anaemia?
Mutations in BRCA2 and FANC genes
How does cisplatin cause DNA damage?
Induces DNA adducts
How does etoponide cause DNA damage?
Induces ds DNA breaks
What is combination therapy?
Using a DNA damaging agent and an inhibitor of checkpoint signalling
How do PARP inhibitors cause cancer cell death?
They exploit pre-existing defects in the DNA repair pathway. So tumours with homozygous BRCA mutations have an disfunctional HR repair pathway and therefore rely heavily on BER.
PARP is an essential enzyme of the BER process, thus PARP inhibitor leads to cell death
Normal cells have a functional HR pathway thus are unaffected
