Structural basis for fork reversal and RAD51 regulation by the SCF ubiquitin ligase complex of F-box helicase 1.

Abstract

Replication fork reversal helps maintain genomic stability during replication stress. F-box helicase 1 (FBH1) catalyzes fork reversal and is an SCF (SKP-CUL1-F-box) E3 ubiquitin ligase that limits RAD51 association with chromatin. Here, we show that preferential binding of SCF^FBH1 to the lagging strand template at DNA fork structures stimulates helicase activity and is required for fork reversal. A cryo-EM structure of SCF^FBH1 bound to DNA representing a stalled fork reveals an intimate interaction between FBH1 and the fork junction. Disruption of this interface severely curtails fork reversal in vitro and replication progression in cells, providing a model for how ssDNA translocation by FBH1 facilitates annealing of parental DNA by a fundamentally different mechanism than the fork remodelers SMARCAL, HLTF, and ZRANB3. The structure provides a model for SCF^FBH1 disassembly of RAD51 filaments through translocation and ubiquitination, and implies that RAD51 is associated with the lagging strand at stalled forks.