Lecture 22 - DNA damage and repair Flashcards
Wht uis unique about DNA to other biological macromolecule ?
It is the only one repaired, everthing else is replaced
What are the consequnces of DNA damage ?
DNA Damage -> DNA Damage to predominantly non-dividing cells -> Blocking of Transcriptionm -> reduced gene expression -> functional decline of tissues and organs = Aging
DNA Damage -> DNA Dmage to cells that proliferate -> Errors of Replication of repair -> Cell death
DNA Damage -> DNA Dmage to cells that proliferate -> Errors of Replication of repair -> mutations -> Pre-malignant field defect -> Cancer
What are the 2 different ways cells can be under attack? Give examples
Endogenous - spontaeneuos in cell i.e hydrolysis, oxygen species, by-products of metabolism
Exogenous - rxns with molecules from outside the cell i.e UV, X-ray, carcinogens, chemotherapues
What are the differences in types of DNA damage between Endogenous and Exgenous DNA?
Endogenous:
Depurination (Abasic sites)
Deamination
Methylation
Replication errors
(These effect ONE strand of the DNA helix)
Exogenous:
Pyrimidine dimers
Double strand breaks
Interstrand crosslinks
(Pyriminidine affects one strand the other effect two)
What is Deamination ?
Removal of the amino group by hydrolysis.
Results in changes to the DNA bases
Cytosine removes NH2 molecule via hydrolysis, oxygen attaches creating Uracil
amino acid C is deanimated from C to U
This is a CG-TA point mutation
What are the 2 different types of point mutations?
- Trabnsitions - A to G or C to T. Less likely to result in aa substitutions
- Transversions - A to C or T
G to C or T
C to A or G
T to A or G
What is depurination ?
When the whole base is removed.
N-glycosidic bond is a common substrate for hydrolysis left with an basic site.
Happen around 20,000 per genome a day, most common at purine bases
What are the consequences of Depurination?
Frame-shift mutation
You will have one accurately syntheised strand but one has a deleted A-T nucleotide.
Frame shift mutations generate missense proteins that doesn’t function properly
e.g. sickle cell anemia single point mutation in beat haemoglobin gene causes sickle shape, cannot function
How does UV light damage DNA?
DNA doesnt have flexibitly, structure is distorted.
How does UV light cause interstrand DNA crosslinks and DNA-protein crosslinks?
INsterstrand crosslinks - incorrect bases pair (DNA cant unwind during replication)
DNA-protein crosslinks - form on one strand
Highly toxic
Give examples of what causes single strand break inducers that affect phosphate in DNA
Reactive oxygen specices
Hydroxyurea
Camptothecin
Give examples of what causes double strand break inducers that affect phosphate in DNA
X-rays
Ionising radiation
Topoisomerase II inhibitors
What is Base excision repair ?
Repairs base damage using base flippping strategy to identify errors.
e.g. abasic sites and deamination
What is Base excision repair ?
Repairs base damage using base flippping strategy to identify errors.
e.g. abasic sites and deamination
What is Nucleotide excision repair?
Repairs damage when moprthan one base is involved
e.g. pryrimindine dimers caused by UV light
Invovles the excision of short pathces of single stranded DNA to remove the affected bases.
What can Translesional DNA Polymerases do?
They can replicate highly damaged DNA
What do Translesional DNA Polymerases lack?
Precision in template recognition and substrate base choice
Exonucleolytic proof-reading activity
What do Translesional DNA Polymerases cause?
Causes most base substitution and single nucleotide deletion mutations. Quite error prone.
What is double strand break repair?
2 mechanisms exist to repair double strand breaks
There are 2 mechanims
1. Non home ologous end joining (NHEJ)
2. Homologous Recombination
What are the steps in Non homologous end joining?
Error prone
restricted to G1 phase
Usuallu results in loss of nucleoides surrounding the breaksite
Quick process
What happens in Homologous Recombination?
Error-free repair
Occurs only in S-phase
Uses intact sister chromatid as a template
Double strand break is accurately repaired
Whare are the 3 places in cell cycle where DNA damage is detected and acted upon to stop the cycle?
G1 checkpoint - NHEJ
Entry to S-phase
Entry into mitosis
and check for chromosome
How is damage detected?
Protein kinases ATM/ATR activated and associate at the damaged site
This activates other kinases to block cell cycle. Chk1/Chk2 kinases are activated
p53 is phosphorylated and stabalised in order to bind to p21 (inhibitor of cyclin-dependent kniases)
p21 renders G1/S-CDK and S-CDK complexes INACTIVE
Prevents cell cycle progression
What is the disease Xeroderma Pigmentosum ?
Predisposition disease - Xeroderma Pigmentosum
Autosomal recessive disease
High risk of skin cancer
Associated with a defect in nucleotide excision repair (UV damage)
What are BRCA 1/2 genes?
Invovled in homologous recombination
80-90% of all inherited breast cancer contain BRCA 1/2
How do BRCA2 genes cause cancer?
BRCA2 deficient cells cause
genomic instability
DNA damage sensitivity agents
defective in
