A Polar and Nucleotide-Dependent Mechanism of Action for RAD51 Paralogs in RAD51 Filament Remodeling

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Authors

TAYLOR Martin R.G. ŠPÍREK Mário MA Chu Jian CARZANIGA Raffaella TAKAKI Tohru COLLINSON Lucy M. GREENE Eric C. KREJČÍ Lumír BOULTON Simon J.

Year of publication 2016
Type Article in Periodical
Magazine / Source Molecular Cell
MU Faculty or unit

Faculty of Medicine

Citation
Doi http://dx.doi.org/10.1016/j.molcel.2016.10.020
Field Genetics and molecular biology
Keywords SINGLE-STRANDED-DNA; INDUCED FLUORESCENCE ENHANCEMENT; CANCER TUMOR-SUPPRESSOR; REPLICATION PROTEIN-A; HOMOLOGOUS RECOMBINATION; SACCHAROMYCES-CEREVISIAE; MAMMALIAN-CELLS; COMPLEX; REPAIR; PROMOTES
Description 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 50 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.
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