Replication restart in UV-irradiated Escherichia coli involving pols II, III, V, PriA, RecA and RecFOR proteins.
In Escherichia coli, UV-irradiated cells resume DNA synthesis after a transient inhibition by a process called replication restart. To elucidate the role of several key proteins involved in this process, we have analysed the time dependence of replication restart in strains carrying a combination of mutations in lexA, recA, polB (pol II), umuDC (pol V), priA, dnaC, recF, recO or recR. We find that both pol II and the origin-independent primosome-assembling function of PriA are essential for the immediate recovery of DNA synthesis after UV irradiation. In their absence, translesion replication or 'replication readthrough' occurs approximately 50 min after UV and is pol V-dependent. In a wild-type, lexA+ background, mutations in recF, recO or recR block both pathways. Similar results were obtained with a lexA(Def) recF strain. However, lexA(Def) recO or lexA(Def) recR strains, although unable to facilitate PriA-pol II-dependent restart, were able to perform pol V-dependent readthrough. The defects in restart attributed to mutations in recF, recO or recR were suppressed in a recA730 lexA(Def) strain expressing constitutively activated RecA (RecA*). Our data suggest that in a wild-type background, RecF, O and R are important for the induction of the SOS response and the formation of RecA*-dependent recombination intermediates necessary for PriA/Pol II-dependent replication restart. In con-trast, only RecF is required for the activation of RecA that leads to the formation of pol V (UmuD'2C) and facilitates replication readthrough.
Rangarajan
Savithri
S
Department of Biological Sciences and Chemistry, Hedco Molecular Biology Laboratories, University of Southern California, Los Angeles, CA 90089-1340, USA.
Woodgate
Roger
R
Goodman
Myron F
MF
D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism / Q000528:radiation effects; D004257:DNA Polymerase II / Q000235:genetics / Q000378:metabolism; D004258:DNA Polymerase III / Q000235:genetics / Q000378:metabolism; D004261:DNA Replication / Q000528:radiation effects*; D004269:DNA, Bacterial / Q000096:biosynthesis; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics* / Q000528:radiation effects*; D029968:Escherichia coli Proteins; D009154:Mutation; D011938:Rec A Recombinases / Q000235:genetics / Q000378:metabolism; D051716:Replication Protein A; D012697:Serine Endopeptidases / Q000096:biosynthesis / Q000235:genetics / Q000528:radiation effects; D014466:Ultraviolet Rays; D000099160:LexA Repressor Protein
D001426:Bacterial Proteins; D004257:DNA Polymerase II; D004258:DNA Polymerase III; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D004259:DNA-Directed DNA Polymerase; D029968:Escherichia coli Proteins; D011938:Rec A Recombinases; D051716:Replication Protein A; D012697:Serine Endopeptidases; D000099160:LexA Repressor Protein; C081006:RecO protein, E coli; C489318:RecR protein, E coli; C489340:recF protein, E coli; C065831:RecR protein, Bacteria; C066218:recF protein, Bacteria; C120926:DNA polymerase V, E coli