title: Altered translesion synthesis in E. coli Pol V mutants selected for increased recombination inhibition.
abstract: Replication of damaged DNA, also termed as translesion synthesis (TLS), involves specialized DNA polymerases that bypass DNA lesions. In Escherichia coli, although TLS can involve one or a combination of DNA polymerases depending on the nature of the lesion, it generally requires the Pol V DNA polymerase (formed by two SOS proteins, UmuD' and UmuC) and the RecA protein. In addition to being an essential component of translesion DNA synthesis, Pol V is also an antagonist of RecA-mediated recombination. We have recently isolated umuD' and umuC mutants on the basis of their increased capacity to inhibit homologous recombination. Despite the capacity of these mutants to form a Pol V complex and to interact with the RecA polymer, most of them exhibit a defect in TLS. Here, we further characterize the TLS activity of these Pol V mutants in vivo by measuring the extent of error-free and mutagenic bypass at a single (6-4)TT lesion located in double stranded plasmid DNA. TLS is markedly decreased in most Pol V mutants that we analyzed (8/9) with the exception of one UmuC mutant (F287L) that exhibits wild-type bypass activity. Somewhat unexpectedly, Pol V mutants that are partially deficient in TLS are more severely affected in mutagenic bypass compared to error-free synthesis. The defect in bypass activity of the Pol V mutant polymerases is discussed in light of the location of the respective mutations in the 3D structure of UmuD' and the DinB/UmuC homologous protein Dpo4 of Sulfolobus solfataricus.
authors.affiliation: Institut de Génétique et Microbiologie, Bât. 409, Université Paris-Sud, F-91405, Orsay, France. suzanne.sommer@igmors.u-psud.fr
authors.forename: Suzanne
authors.initials: S
authors.lastname: Sommer
authors.forename: Olivier J
authors.initials: OJ
authors.lastname: Becherel
authors.forename: Geneviève
authors.initials: G
authors.lastname: Coste
authors.forename: Adriana
authors.initials: A
authors.lastname: Bailone
authors.forename: Robert P P
authors.initials: RP
authors.lastname: Fuchs
mesh_terms: D004247:DNA / Q000235:genetics / Q000378:metabolism; D004249:DNA Damage; D004261:DNA Replication; D004259:DNA-Directed DNA Polymerase / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins; D007266:Inhibition, Psychological; D009154:Mutation; D010957:Plasmids; D011938:Rec A Recombinases / Q000378:metabolism*; D011995:Recombination, Genetic; D016944:Sulfolobus / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D029968:Escherichia coli Proteins; D004247:DNA; D011938:Rec A Recombinases; C120926:DNA polymerase V, E coli; D004259:DNA-Directed DNA Polymerase