Central to homologous recombination in eukaryotes is the RAD51 recombinase, which forms helical nucleoprotein filaments on single-stranded DNA (ssDNA) and catalyzes strand invasion with homologous duplex DNA. Various regulatory proteins assist this reaction including the RAD51 paralogs. We recently discovered that a RAD51 paralog complex from C. elegans, RFS-1/RIP-1, functions predominantly downstream of filament assembly by binding and remodeling RAD-51-ssDNA filaments to a conformation more proficient for strand exchange. Here, we demonstrate that RFS-1/RIP-1 acts by shutting down RAD-51 dissociation from ssDNA. Using stopped-flow experiments, we show that RFS-1/RIP-1 confers this dramatic stabilization by capping the 5' end of RAD-51-ssDNA filaments. Filament end capping propagates a stabilizing effect with a 5'→3' polarity approximately 40 nucleotides along individual filaments. Finally, we discover that filament capping and stabilization are dependent on nucleotide binding, but not hydrolysis by RFS-1/RIP-1. These data define the mechanism of RAD51 filament remodeling by RAD51 paralogs.

Clare Hall Laboratory The Francis Crick Institute South Mimms EN6 3LD UK

Department of Biochemistry and Molecular Biophysics Columbia University Medical Center New York City NY 10032 USA

Department of Biology Masaryk University 62500 Brno Czech Republic; International Clinical Research Center St Anne's University Hospital in Brno 62500 Brno Czech Republic

Department of Biology Masaryk University 62500 Brno Czech Republic; International Clinical Research Center St Anne's University Hospital in Brno 62500 Brno Czech Republic; National Centre for Biomolecular Research Masaryk University 62500 Brno Czech Republic

Electron Microscopy Science Technology Platform Lincoln's Inn Fields Laboratory The Francis Crick Institute London WC2A 3LY UK

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