A polar and nucleotide-dependent mechanism of action for RAD51 paralogs in RAD51 filament remodeling
journal contributionposted on 02.10.2020, 14:42 by Martin RG Taylor, Mário Špírek, Chu Jian Ma, Raffaella Carzaniga, Tohru Takaki, Lucy M Collinson, Eric C Greene, Lumir Krejci, Simon J Boulton
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.
DNA repairRad51Rad51 paralogsfilamentsgenome stabilityhomologous recombinationCaenorhabditis elegans ProteinsCarrier ProteinsDNA, Single-StrandedDNA-Binding ProteinsIntermediate FilamentsMultiprotein ComplexesProtein BindingRad51 RecombinaseRecombinational DNA RepairBoulton FC001048EM06 Biological Sciences11 Medical and Health SciencesDevelopmental Biology