Abstract
RuvAB and RuvABC complexes catalyze branch migration and resolution of Holliday junctions (HJs) respectively. In addition to their action in the last steps of homologous recombination, they process HJs made by replication fork reversal, a reaction which occurs at inactivated replication forks by the annealing of blocked leading and lagging strand ends. RuvAB was recently proposed to bind replication forks and directly catalyze their conversion into HJs. We report here the isolation and characterization of two separation-of-function ruvA mutants that resolve HJs, based on their capacity to promote conjugational recombination and recombinational repair of UV and mitomycin C lesions, but have lost the capacity to reverse forks. In vivo and in vitro evidence indicate that the ruvA mutations affect DNA binding and the stimulation of RuvB helicase activity. This work shows that RuvA's actions at forks and at HJs can be genetically separated, and that RuvA mutants compromised for fork reversal remain fully capable of homologous recombination.
Highlights
DNA replication and recombination are two processes that are recognized as more closely connected than originally suspected
DNA replication is the process by which DNA strands are copied to ensure the transmission of the genetic material to daughter cells
The subject of the present study is RuvAB, a highly conserved bacterial complex used as the model enzyme for resolution of recombination intermediates, which we found to act at blocked forks
Summary
DNA replication and recombination are two processes that are recognized as more closely connected than originally suspected. The best-documented example is the bacterial PriA protein, which promotes replication restart (i) independently of recombination by its virtue of recognizing replication forks and (ii) during double-stranded DNA end recombinational repair by its virtue of recognizing Dloop structures (reviewed in [5]). Another example is the RuvAB complex, originally identified for its activity on Holliday junctions (HJs), four-DNA arm recombination intermediates (reviewed in [6,7]), and recently proposed to act on inactivated replication forks [8] (Figure 1)
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