RAD50 Is Required for Efficient Initiation of Resection and Recombinational Repair at Random, γ-Induced Double-Strand Break Ends 英文参考文献.docVIP

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RAD50 Is Required for Efficient Initiation of Resection and Recombinational Repair at Random, γ-Induced Double-Strand Break Ends 英文参考文献.doc

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RAD50 Is Required for Efficient Initiation of Resection and Recombinational Repair at Random, γ-Induced Double-Strand Break Ends 英文参考文献

RAD50IsRequiredforEfficientInitiationofResection andRecombinationalRepairatRandom,c-Induced Double-StrandBreakEnds JimWestmoreland1,WenjianMa1,YanYan1,KellyVanHulle2,AnnaMalkova2,MichaelA.Resnick1* 1ChromosomeStabilitySection,NationalInstituteofEnvironmentalHealthSciences,NationalInstitutesofHealth,ResearchTrianglePark,NorthCarolina,UnitedStatesof America,2BiologyDepartment,IndianaUniversity–PurdueUniversity,Indianapolis,Indiana,UnitedStatesofAmerica Abstract ResectionofDNAdouble-strandbreak(DSB)endsisgenerallyconsideredacriticaldeterminantinpathwaysofDSBrepair andgenomestability.Unlikeforenzymaticallyinducedsite-specificDSBs,littleisknownaboutprocessingofrandom‘‘dirty- ended’’DSBscreatedbyDNAdamagingagentssuchasionizingradiation.Herewepresentanovelsystemformonitoring earlyeventsintherepairofrandomDSBs,basedonourfindingthatsingle-strandtailsgeneratedbyresectionattheendsof largemoleculesinbuddingyeastdecreasesmobilityduringpulsedfieldgelelectrophoresis(PFGE).Weutilizedthis‘‘PFGE- shift’’tofollowthefateofbothendsoflinearmoleculesgeneratedbyasinglerandomDSBincircularchromosomes.Within 10min after c-irradiation of G2/M arrested WT cells, there is a near-synchronous PFGE-shift of the linearized circular molecules,correspondingtoresectionofafewhundredbases.Resectionattheradiation-inducedDSBscontinuessothat bythetimeofsignificantrepairofDSBsat1hrthereisabout1–2kbresectionperDSBend.ThePFGE-shiftiscomparablein WTandrecombination-defectiverad52andrad51strainsbutsomewhatdelayedinexo1mutants.However,inrad50and mre11nullmutantstheinitiationandgenerationofresectedendsatradiation-inducedDSBendsisgreatlyreducedinG2/M. Thus, the Rad50/Mre11/Xrs2 complex is responsible for rapid processing of most damaged ends into substrates that subsequentlyundergorecombinationalrepair.AsimilarrequirementwasfoundforRAD50inasynchronouslygrowingcells. Amongthefewmoleculesexhibitingshiftintherad50mutant,theresidualresectionisconsistentwithresectionatonly oneoftheDSBends.Surprisingly,within1hrafterirra