DNA damage and repairDNA damage and repairWilliam Kaufmann31-325 LCCC [email protected] four R’s of human DNA metabolismMechanism of cytotoxicityof topoIpoisonsTopoII reactionsDNA strand scission by topoIIpoisonsSites of alkylation of DNABP adduct on DNAAn arrested repliconPathways of DNA repairO6-alkylguanine DNA alkyltransferasesuicide protein transfers methyl groupBase excision repairglycosylase, AP endonuclease, DNA polymerase, ligaseNucleotide excision repairtranscription-coupled and globalCross-link repairFanconi’s anemia genes and BRCA2Double-strand break repairHomologous recombination and non-homologous end- joiningHeteroduplex and mismatch repairPost-replication repair; translesion synthesis by pol eta, zeta, iota or kappaNucleotide excision repairPathways of repair of DNA double strand breaksNon-homologous end-joining; DNA-PK, Ku70/80, XRCC1, DNA ligase IV; immunoglobulin and TCR gene rearrangementHomologous recombination: rad51,52, XRCC2,3,4Crosslink repair requires Fanconi’sanemia genes and BRCA2Familial cancer syndromesAtaxia telangiectasia - cell cycle checkpoint function, DNA repair. ATM, NBS1, MRE-11Fanconi’s anemia - DNA repair. BRCA2 and 5 other genesHNPCC - mismatch repair, hMSH2, hMLH1, PMS2, hMSH6Xeroderma pigmentosum - nucleotide excision repair and post-replication repair. 8 XP genesFamilial breast cancer I - S and G2 checkpoint responses. BRCA1Li-Fraumeni syndrome – cell cycle checkpoint function and DNA repair. P53, Chk2Bloom’s syndrome, Werner syndrome, Rothmund-Thompson syndrome – chromosomal instability. Blm, Wrn, RecQSpindle assembly and polyploidization checkpointscontrolcolcemidNHF-neo NHF-E6P53-dependent G1 checkpoint functionATM, Nbs1, and Mre11 are required for the IR-induced S checkpoint.NHF10.000.050.100.150.200.250.300.350 102030Sham 1.5 GyAT M - /-0.000.100.200.300.400.500.600.700.800 102030Sham 1.5 GyNBS1 -/-0.000.100.200.300.400.500.600.700 102030Sham 1.5GyMre11 -/-0.000.100.200.300.400.500.600 102030Sham 1.5 GyNormalized 3H CPMFraction NumberATR is an essential checkpoint kinaseAtaxia telangiectasia- and rad3-relatedATR-null embryos die at day 3 with severe chromosomal damageExpression of kinase-inactive ATR overrides checkpoint responses to IR and UVCCell Cycle CheckpointsCell Cycle CheckpointsMMSSGG11GG22MMG1DelayG2DelayIRDNA DamageATM/ATRp53 CHK1 p21/14-3-3 CDC25C Cyclin B1 / CDC2UV IR DNA DamageATR ATM p53CHK2 p21Cyclin / CDK’sPlk-1Crm1Passive and active inhibition of DNA replication by UVCMechanisms and consequences of inhibition of DNA replication by UVCTTTTATRChk1DNA pol η/ζDNA dsbATMG2 checkpointReplicon initiationBlock pol α/δTTNERp53 p48UVC-induced chromosomal aberrationsEvolution of DNA damage during the cell cycleRecent models of carcinogenesis hold that deregulated DNA replication drives genetic instabilityBartkova et al., 2005Oncogene-induced senescence is associated with DNA re-replicationOIS = oncogene-induced senescenceAmplified globin
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