UvsW protein regulates bacteriophage T4 origin-dependent replication by unwinding R-loops

The UvsW protein of bacteriophage T4 is involved in many aspects of phage D NA metabolism, including repair, recombination, and recombination-dependent replication. UvsW has also been implicated in the repression of origin-dep endent replication at late times of infection, when UvsW is normally synthe sized. Two well-characterized T4 origins, ori(uvsY) and ori(34), are believ ed to initiate replication through an R-loop mechanism. Here we provide bot h in vivo and in vitro evidence that UvsW is an RNA-DNA helicase that catal yzes the dissociation of RNA from origin R-loops, Two-dimensional gel analy ses show that the replicative intermediates formed at ori(uvsY) persist lon ger in a uvsW mutant infection than in a wild-type infection. In addition, the inappropriate early expression of UvsW protein results in the loss of t hese replicative intermediates. Using a synthetic origin R-loop, we also de monstrate that purified UvsW functions as a helicase that efficiently disso ciates RNA from R-loops, These and previous results from a number of studie s provide strong evidence that UvsW is a molecular switch that allows T4 re plication to progress from a mode that initiates from R-loops at origins to a mode that initiates from D-loops formed by recombination proteins.