title: RecA bundles mediate homology pairing between distant sisters during DNA break repair.
abstract: DNA double-strand break (DSB) repair by homologous recombination has evolved to maintain genetic integrity in all organisms. Although many reactions that occur during homologous recombination are known, it is unclear where, when and how they occur in cells. Here, by using conventional and super-resolution microscopy, we describe the progression of DSB repair in live Escherichia coli. Specifically, we investigate whether homologous recombination can occur efficiently between distant sister loci that have segregated to opposite halves of an E. coli cell. We show that a site-specific DSB in one sister can be repaired efficiently using distant sister homology. After RecBCD processing of the DSB, RecA is recruited to the cut locus, where it nucleates into a bundle that contains many more RecA molecules than can associate with the two single-stranded DNA regions that form at the DSB. Mature bundles extend along the long axis of the cell, in the space between the bulk nucleoid and the inner membrane. Bundle formation is followed by pairing, in which the two ends of the cut locus relocate at the periphery of the nucleoid and together move rapidly towards the homology of the uncut sister. After sister locus pairing, RecA bundles disassemble and proteins that act late in homologous recombination are recruited to give viable recombinants 1-2-generation-time equivalents after formation of the initial DSB. Mutated RecA proteins that do not form bundles are defective in sister pairing and in DSB-induced repair. This work reveals an unanticipated role of RecA bundles in channelling the movement of the DNA DSB ends, thereby facilitating the long-range homology search that occurs before the strand invasion and transfer reactions.
authors.affiliation: Department of Biochemistry, University of Oxford, Oxford OX1 3QU, UK.
authors.forename: Christian
authors.initials: C
authors.lastname: Lesterlin
authors.affiliation: Department of Biochemistry, University of Oxford, Oxford OX1 3QU, UK.
authors.forename: Graeme
authors.initials: G
authors.lastname: Ball
authors.affiliation: Department of Biochemistry, University of Oxford, Oxford OX1 3QU, UK.
authors.forename: Lothar
authors.initials: L
authors.lastname: Schermelleh
authors.affiliation: Department of Biochemistry, University of Oxford, Oxford OX1 3QU, UK.
authors.forename: David J
authors.initials: DJ
authors.lastname: Sherratt
mesh_terms: D023902:Chromosome Pairing; D002876:Chromosomes, Bacterial / Q000235:genetics / Q000378:metabolism; D053903:DNA Breaks, Double-Stranded; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D011938:Rec A Recombinases / Q000235:genetics / Q000378:metabolism*; D059767:Recombinational DNA Repair / Q000235:genetics
chemical_list: D004269:DNA, Bacterial; D004277:DNA, Single-Stranded; D029968:Escherichia coli Proteins; D050505:Mutant Proteins; D011938:Rec A Recombinases