DNA Repair Repair and Recombination Mutation frequencies 235
DNA Repair
Repair, and Recombination
Mutation frequencies 235, 320
Hallmarks of cancer
Simplified view of the DNA damage response ATM-Ataxia telangiectasia. Human atm-/- has progeroid appearance, immunodeficinecy , cerebellar degradation and cancer (mice are similar with less progeroid)
Checkpoint
• E. Coli SOS system Emergency response that allow the insertion of non specific base! In bacteria its Din. B, Umu. C and Umu. D that encode for pol IV and V, the error prone polymerase that incorporate an arbitrary base across to the damaged base.
Regulation of Rec. A by Lex in E. coli
Pol eta/iota
Direct reversal By photolyase
Base excision repair • Start with glycosylase that break the sugarbase bond* and release the altred base and generating apurinic or apyrimidinic sites. * For example guanine +EMS -> 0 -6 -ethylgunine (can mispaired with Thymine)
Nucleotide excision repair (NER) in bacteria
Nucleotide excision repair (NER) in yeast
XPD and XPB – helicase XPG and XPF - endonuclase
Lesions are initially recognized, either by a translocating RNA polymerase (RNAP) (for transcription-coupled repair (TCR); left) or through the binding of the lesion sensor DNA damage-binding-2 (DDB 2;
Cockayne syndrome • Mutation in CSA or CSB causes to retinal degeneration, hearing impairment, ataxia and photosensitivity • Die very young
Tricothiodystrophy (TTD) • Mutations in XPD (the helicase)
TTD Human shows photosensetivity, brittle hair, skin defects and short life span. XPD -/- mice show also wasting, scoliosis, osteophoresis and melanocyte loss. 3 m WT 15 m WT TTD
Heterogeneous ovarian dysfunction
Mismatch repair in bacteria
Mismatch repair (NER) in human
Double strand breaks repair
How to detect DSB’s? g. H 2 AX Commet Assay
Two options of repair
BRCA 1 • Possess ubiquitin ligase activity. • Phosphorylated by ATM/ATR. • BRCA 1 deficiency leads to impaired HR and hypersensitivity to DNA damaging agents and genome instability. • Other functions including transcription, G 2/M checkpoint etc.
Loss of heterozygosity
BRCA 1
During the pachytene stage
Meiotic Recombination
Synaptonemal complex
Pulsed field gel electrophoresis
DSBs mapping in yeast Probe
End products of recombination
Crossover vs. gene conversion
Spo 11
Gene conversion 4: 4 6: 2 2: 6 5: 3 3: 5 A A A a A A a A a a a a Aberrant ratio
- Slides: 83