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 DNA metabolism, including repair, recombination, and recombination-dependent replication. UvsW has also been implicated in the repression of origin-dependent replication at late times of infection, when UvsW is normally synthesized. Two well-characterized T4 origins, ori(uvsY) and ori(34), are believed to initiate replication through an R-loop mechanism. Here we provide both in vivo and in vitro evidence that UvsW is an RNA-DNA helicase that catalyzes the dissociation of RNA from origin R-loops. Two-dimensional gel analyses show that the replicative intermediates formed at ori(uvsY) persist longer 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 these replicative intermediates. Using a synthetic origin R-loop, we also demonstrate that purified UvsW functions as a helicase that efficiently dissociates RNA from R-loops. These and previous results from a number of studies provide strong evidence that UvsW is a molecular switch that allows T4 replication to progress from a mode that initiates from R-loops at origins to a mode that initiates from D-loops formed by recombination proteins.
Department of Microbiology, Duke University Medical Center, Durham, North Carolina 27710, USA.
K C
KC
Dudas
K N
KN
Kreuzer
D017122:Bacteriophage T4 / Q000235:genetics / Q000502:physiology*; D001483:Base Sequence; D004265:DNA Helicases / Q000378:metabolism*; D017931:DNA Primers / Q000235:genetics; D004261:DNA Replication / Q000235:genetics / Q000502:physiology; D004926:Escherichia coli / Q000821:virology; D005814:Genes, Viral; D009154:Mutation; D009690:Nucleic Acid Conformation; D012367:RNA, Viral / Q000737:chemistry / Q000378:metabolism; D018741:Replication Origin; D014764:Viral Proteins / Q000378:metabolism*; D014779:Virus Replication / Q000235:genetics / Q000502:physiology
D017931:DNA Primers; D012367:RNA, Viral; D014764:Viral Proteins; D004265:DNA Helicases; C107414:UvsW protein, Bacteriophage T4