title: A Polar and Nucleotide-Dependent Mechanism of Action for RAD51 Paralogs in RAD51 Filament Remodeling.
abstract: 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.
authors.affiliation: Clare Hall Laboratory, The Francis Crick Institute, South Mimms EN6 3LD, UK.
authors.forename: Martin R G
authors.initials: MRG
authors.lastname: Taylor
authors.affiliation: Department of Biology, Masaryk University, 62500 Brno, Czech Republic; International Clinical Research Center, St. Anne's University Hospital in Brno, 62500 Brno, Czech Republic.
authors.forename: Mário
authors.initials: M
authors.lastname: Špírek
authors.affiliation: Department of Biochemistry and Molecular Biophysics, Columbia University Medical Center, New York City, NY 10032, USA.
authors.forename: Chu
authors.initials: C
authors.lastname: Jian Ma
authors.affiliation: Electron Microscopy Science Technology Platform, Lincoln's Inn Fields Laboratory, The Francis Crick Institute, London WC2A 3LY, UK.
authors.forename: Raffaella
authors.initials: R
authors.lastname: Carzaniga
authors.affiliation: Clare Hall Laboratory, The Francis Crick Institute, South Mimms EN6 3LD, UK.
authors.forename: Tohru
authors.initials: T
authors.lastname: Takaki
authors.affiliation: Electron Microscopy Science Technology Platform, Lincoln's Inn Fields Laboratory, The Francis Crick Institute, London WC2A 3LY, UK.
authors.forename: Lucy M
authors.initials: LM
authors.lastname: Collinson
authors.affiliation: Department of Biochemistry and Molecular Biophysics, Columbia University Medical Center, New York City, NY 10032, USA.
authors.forename: Eric C
authors.initials: EC
authors.lastname: Greene
authors.affiliation: 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. Electronic address: lkrejci@chemi.muni.cz.
authors.forename: Lumir
authors.initials: L
authors.lastname: Krejci
authors.affiliation: Clare Hall Laboratory, The Francis Crick Institute, South Mimms EN6 3LD, UK. Electronic address: simon.boulton@crick.ac.uk.
authors.forename: Simon J
authors.initials: SJ
authors.lastname: Boulton
mesh_terms: D029742:Caenorhabditis elegans Proteins / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D007382:Intermediate Filaments / Q000235:genetics / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000378:metabolism; D011485:Protein Binding; D051135:Rad51 Recombinase / Q000235:genetics / Q000378:metabolism*; D059767:Recombinational DNA Repair
chemical_list: D029742:Caenorhabditis elegans Proteins; D002352:Carrier Proteins; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D046912:Multiprotein Complexes; C000602003:RFS-1 protein, C elegans; C000602004:RIP-1 protein, C elegans; D051135:Rad51 Recombinase; C495148:rad-51 protein, C elegans
keywords: DNA repair; Rad51; Rad51 paralogs; filaments; genome stability; homologous recombination