title: Oxidative DNA damage defense systems in avoidance of stationary-phase mutagenesis in Pseudomonas putida.
abstract: Oxidative damage of DNA is a source of mutation in living cells. Although all organisms have evolved mechanisms of defense against oxidative damage, little is known about these mechanisms in nonenteric bacteria, including pseudomonads. Here we have studied the involvement of oxidized guanine (GO) repair enzymes and DNA-protecting enzyme Dps in the avoidance of mutations in starving Pseudomonas putida. Additionally, we examined possible connections between the oxidative damage of DNA and involvement of the error-prone DNA polymerase (Pol)V homologue RulAB in stationary-phase mutagenesis in P. putida. Our results demonstrated that the GO repair enzymes MutY, MutM, and MutT are involved in the prevention of base substitution mutations in carbon-starved P. putida. Interestingly, the antimutator effect of MutT was dependent on the growth phase of bacteria. Although the lack of MutT caused a strong mutator phenotype under carbon starvation conditions for bacteria, only a twofold increased effect on the frequency of mutations was observed for growing bacteria. This indicates that MutT has a backup system which efficiently complements the absence of this enzyme in actively growing cells. The knockout of MutM affected only the spectrum of mutations but did not change mutation frequency. Dps is known to protect DNA from oxidative damage. We found that dps-defective P. putida cells were more sensitive to sudden exposure to hydrogen peroxide than wild-type cells. At the same time, the absence of Dps did not affect the accumulation of mutations in populations of starved bacteria. Thus, it is possible that the protective role of Dps becomes essential for genome integrity only when bacteria are exposed to exogenous agents that lead to oxidative DNA damage but not under physiological conditions. Introduction of the Y family DNA polymerase PolV homologue rulAB into P. putida increased the proportion of A-to-C and A-to-G base substitutions among mutations, which occurred under starvation conditions. Since PolV is known to perform translesion synthesis past damaged bases in DNA (e.g., some oxidized forms of adenine), our results may imply that adenine oxidation products are also an important source of mutation in starving bacteria.
authors.affiliation: Department of Genetics, Institute of Molecular and Cell Biology, Tartu University and Estonian Biocentre, 23 Riia Street, 51010 Tartu, Estonia.
authors.forename: Signe
authors.initials: S
authors.lastname: Saumaa
authors.forename: Andres
authors.initials: A
authors.lastname: Tover
authors.forename: Mariliis
authors.initials: M
authors.lastname: Tark
authors.forename: Radi
authors.initials: R
authors.lastname: Tegova
authors.forename: Maia
authors.initials: M
authors.lastname: Kivisaar
mesh_terms: D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000235:genetics / Q000502:physiology; D015169:Colony Count, Microbial; D004249:DNA Damage; D045647:DNA Glycosylases / Q000235:genetics / Q000502:physiology; D004260:DNA Repair / Q000502:physiology*; D045643:DNA Repair Enzymes / Q000235:genetics / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology; D004259:DNA-Directed DNA Polymerase / Q000502:physiology; D017353:Gene Deletion; D016296:Mutagenesis; D009154:Mutation; D016958:Pseudomonas putida / Q000235:genetics / Q000502:physiology*; D011995:Recombination, Genetic
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C083911:DPS protein, Bacteria; D004259:DNA-Directed DNA Polymerase; D045647:DNA Glycosylases; C061738:mutY adenine glycosylase; D045643:DNA Repair Enzymes