Role of Saccharomyces Single-Stranded DNA-Binding Protein RPA in the Strand Invasion Step of Double-Strand Break Repair 英文参考文献.docVIP

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Role of Saccharomyces Single-Stranded DNA-Binding Protein RPA in the Strand Invasion Step of Double-Strand Break Repair 英文参考文献.doc

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o PL S BIOLOGY RoleofSaccharomycesSingle-Stranded DNA-BindingProteinRPAintheStrand InvasionStepofDouble-StrandBreakRepair Xuan Wang, James E.Haber* RosenstielCenterandDepartmentofBiology,BrandeisUniversity,Waltham,Massachusetts,UnitedStatesofAmerica Thesingle-strandedDNA(ssDNA)-bindingproteinreplicationproteinA(RPA)isessentialforbothDNAreplicationand recombination. Chromatin immunoprecipitation techniques were used to visualize the kinetics and extent of RPA binding following induction of a double-strand break (DSB) and during its repair by homologous recombination in yeast. RPA assembles at the HO endonuclease-cut MAT locus simultaneously with the appearance of the DSB, and bindingspreadsawayfromtheDSBas59to39exonucleaseactivitycreatesmoressDNA.RPAbindingprecedesbinding oftheRad51recombinationprotein.TheextentofRPAbindingisgreaterwhenRad51isabsent,supportingtheidea thatRad51displacesRPAfromssDNA.RPAplaysanimportantroleduringRAD51-mediatedstrandinvasionoftheMAT ssDNAintothedonorsequenceHML.Thereplication-proficientbutrecombination-defectiverfa1-t11(K45E)mutation in the large subunit of RPA is normal in facilitating Rad51 filament formation on ssDNA, but is unable to achieve synapsisbetweenMATandHML.Thus,RPAappearstoplayaroleinstrandinvasionaswellasinfacilitatingRad51 bindingtossDNA,possiblybystabilizingthedisplacedssDNA. homologue RecA to polymerize across regions that contain secondary structures (Shibata et al. 1980; West et al. 1981; Introduction Repair of double-strand breaks (DSBs) by homologous recombinationinvolvesthesearchforhomologytolocatean intact donor sequence. The search is successful when the broken DNA molecule basepairs with the homologous template, termed synapsis, and forms strand invasion intermediatesofrecombination.Inbuddingyeastandother higher eukaryotes, this process requires both the Rad51 strand exchange protein and the single-stranded DNA (ssDNA)-binding protein replication protein A (RPA) (Alani et al. 1992; Shinohara et al. 1992