FBH1 helicase disrupts RAD51 filaments in vitro and modulates homologous recombination in mammalian cells

TY - JOUR

T1 - FBH1 helicase disrupts RAD51 filaments in vitro and modulates homologous recombination in mammalian cells

AU - Simandlova, Jitka

AU - Zagelbaum, Jennifer

AU - Payne, Miranda J

AU - Chu, Wai Kit

AU - Shevelev, Igor

AU - Hanada, Katsuhiro

AU - Chatterjee, Sujoy

AU - Reid, Dylan A

AU - Liu, Ying

AU - Janscak, Pavel

AU - Rothenberg, Eli

AU - Hickson, Ian D

PY - 2013/11/22

Y1 - 2013/11/22

N2 - Efficient repair of DNA double strand breaks and interstrand cross-links requires the homologous recombination (HR) pathway, a potentially error-free process that utilizes a homologous sequence as a repair template. A key player in HR is RAD51, the eukaryotic ortholog of bacterial RecA protein. RAD51 can polymerize on DNA to form a nucleoprotein filament that facilitates both the search for the homologous DNA sequences and the subsequent DNA strand invasion required to initiate HR. Because of its pivotal role in HR, RAD51 is subject to numerous positive and negative regulatory influences. Using a combination of molecular genetic, biochemical, and single-molecule biophysical techniques, we provide mechanistic insight into the mode of action of the FBH1 helicase as a regulator of RAD51-dependent HR in mammalian cells. We show that FBH1 binds directly to RAD51 and is able to disrupt RAD51 filaments on DNA through its ssDNA translocase function. Consistent with this, a mutant mouse embryonic stem cell line with a deletion in the FBH1 helicase domain fails to limit RAD51 chromatin association and shows hyper-recombination. Our data are consistent with FBH1 restraining RAD51 DNA binding under unperturbed growth conditions to prevent unwanted or unscheduled DNA recombination.

AB - Efficient repair of DNA double strand breaks and interstrand cross-links requires the homologous recombination (HR) pathway, a potentially error-free process that utilizes a homologous sequence as a repair template. A key player in HR is RAD51, the eukaryotic ortholog of bacterial RecA protein. RAD51 can polymerize on DNA to form a nucleoprotein filament that facilitates both the search for the homologous DNA sequences and the subsequent DNA strand invasion required to initiate HR. Because of its pivotal role in HR, RAD51 is subject to numerous positive and negative regulatory influences. Using a combination of molecular genetic, biochemical, and single-molecule biophysical techniques, we provide mechanistic insight into the mode of action of the FBH1 helicase as a regulator of RAD51-dependent HR in mammalian cells. We show that FBH1 binds directly to RAD51 and is able to disrupt RAD51 filaments on DNA through its ssDNA translocase function. Consistent with this, a mutant mouse embryonic stem cell line with a deletion in the FBH1 helicase domain fails to limit RAD51 chromatin association and shows hyper-recombination. Our data are consistent with FBH1 restraining RAD51 DNA binding under unperturbed growth conditions to prevent unwanted or unscheduled DNA recombination.

KW - Animals

KW - Cells, Cultured

KW - Chromatin

KW - DNA

KW - DNA Helicases

KW - DNA-Binding Proteins

KW - Embryonic Stem Cells

KW - F-Box Proteins

KW - Homologous Recombination

KW - Humans

KW - Mice

KW - Multienzyme Complexes

KW - Protein Binding

KW - Rad51 Recombinase

U2 - 10.1074/jbc.M113.484493

DO - 10.1074/jbc.M113.484493

M3 - Journal article

C2 - 24108124

SN - 0021-9258

VL - 288

SP - 34168

EP - 34180

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

IS - 47

ER -