Evidence for Different Pathways during Horizontal Gene Transfer in Competent Bacillus subtilis Cells 英文参考文献.docVIP

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Evidence for Different Pathways during Horizontal Gene Transfer in Competent Bacillus subtilis Cells 英文参考文献.doc

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Evidence for Different Pathways during Horizontal Gene Transfer in Competent Bacillus subtilis Cells 英文参考文献

EvidenceforDifferentPathwaysduringHorizontalGene TransferinCompetentBacillussubtilisCells DawitKidane1.,Begon?aCarrasco2.,CandelaManfredi2,KatharinaRothmaier1,SilviaAyora2 ,Serkalem Tadesse1,JuanC.Alonso2*,PeterL.Graumann1* 1Mikrobiologie,Fakulta¨tfu¨rBiologie,Universita¨tFreiburg,Freiburg,Germany,2DepartmentofMicrobialBiotechnology,CentroNacionaldeBiotecnolog?′a,CSIC,Campus UniversidadAuto′nomadeMadrid,Madrid,Spain Abstract CytologicalandgeneticevidencesuggeststhattheBacillussubtilisDNAuptakemachinerylocalizesatasinglecellpoleand takes up single-stranded (ss) DNA. The integration of homologous donor DNA into the recipient chromosome requires RecA, while plasmid establishment, which is independent of RecA, requires at least RecO and RecU. RecA and RecN colocalize at the polar DNA uptake machinery, from which RecA forms filamentous structures, termed threads, in the presence of chromosomal DNA. We show that the transformation of chromosomal and of plasmid DNA follows distinct pathways.IntheabsenceofDNA,RecUaccumulatedatasinglecellpoleincompetentcells,dependentonRecA.Upon addition of any kind of DNA, RecA formed highly dynamic thread structures, which rapidly grew and shrank, and RecU dissipatedfromthepole.RecOvisibly accumulatedatthecellpoleonlyuponadditionofplasmidDNA,and,toalesser degree,ofphageDNA,butnotofchromosomalDNA.RecOaccumulationwasweaklyinfluencedbyRecN,butnotbyRecA. RecO annealed ssDNA complexed with SsbA in vitro, independent of any nucleotide cofactor. The DNA end-joining Ku proteinwasalsofoundtoplayaroleinviralandplasmidtransformation.Ontheotherhand,transfectionwithSPP1phage DNArequiredfunctionsfrombothchromosomalandplasmidtransformationpathways.Thefindingsshowthatcompetent bacterial cells possess a dynamic DNA recombination machinery that responds in a differential manner depending if enteringDNAshowshomologywithrecipientDNAorhasself-annealingpotential.TransformationwithchromosomalDNA only requires RecA, which forms dynamic filamentous structures th