Increased mutability by oxidative stress in OxyR-deficient Escherichia coli and Salmonella typhimurium cells: clonal occurrence of the mutants during growth on … (original) (raw)
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Mutation Research Letters, 1995
Escherichia coli and Salmonella typhimurium strains deficient in the OxyR-regulated adaptive response to oxidative stress were used to study the mode in which spontaneous SOS-dependent mutations are generated in a distressed bacterial population. When assayed on supplemented selective medium, the E. coli strain IC3821 (trpE65), carrying the AoxyR30 mutation and containing the plasmid pRW144 (mucA/B), showed a frequency of spontaneous Trp ÷ revertants similar to that of the oxyR + control. Instead, the IC3821 strain exhibited an enhancement in the clonal occurrence of spontaneous revertants arising at random during growth on a nonselective medium. A similar enhancement was observed for the S. typhimurium strain TA4125 (hisG428 AoxyR2). The mutator effect observed in oxyRcells would be induced by an increased background of reactive oxygen species; it provides a model for studying the mutability of a cell population constantly exposed to mutation-inducing agents. In the IC3821 strain, revertants were induced by t-butyl hydroperoxide with higher efficiency than in oxyR +. We suggest that strain IC3821 could be useful for the detection of SOS-dependent mutagenesis induced by chemical oxidants.
The EMBO journal, 1988
Mutations that suppressed the H2O2 sensitivity of Escherichia coli oxyR- strains caused elevated levels of one three enzymes that destroy organic and hydrogen peroxides: catalase-hydroperoxidase I (the katG gene product), catalase-hydroperoxidase II (controlled by katEF) or alkyl hydroperoxide reductase (specified by the ahp genes). The continuous high-level expression of any one of these enzymes also conferred resistance in an oxyR deletion mutant against other compounds such as N-ethylmaleimide and the superoxide-generator menadione. Overproduction of alkyl hydroperoxide reductase, but not of the catalases, gave resistance to the organic oxidant cumene hydroperoxide. The E. coli delta oxyR strains also exhibited a strongly elevated frequency of spontaneous mutagenesis, as reported for such mutants in Salmonella typhimurium. This mutagenesis was greatly diminished by the individual overexpression of these scavenging enzymes. All of these phenotypes--enzyme overproduction, resistanc...
Mutation Research-fundamental and Molecular Mechanisms of Mutagenesis, 1995
The Escherichia coli strain IC3821, a ΔoxyR derivative of WP2 uvrA trpE65, was more sensitive to mutagenicity promoted by t-butyl hydroperoxide and cumene hydroperoxide than the isogenic oxyR+ control. Mutagenicity of menadione, a redox cycling quinone, was clearly detected in the ΔoxyR strain, whereas only a slight mutagenic response was observed in the oxyR+ strain. Plumbagin, another quinone structurally similar to menadione, was not mutagenic to any of the strains. These mutagenic responses appeared to involve the SOS processing of oxidative DNA lesions and were mediated by proteins more efficiently than by . In cells lacking mutagenesis proteins, induction of SOS-independent mutations by the two alkyl hydroperoxides required a deficiency in the MutY DNA glycosylase and was increased by the presence of the ΔoxyR mutation. In contrast, the two quinones assayed were unable to induce SOS-independent mutations in the MutY-deficient strains.
Malaysian Journal of Microbiology, 2007
Superoxide dismutase (SOD) is widely distributed in organisms and alleviates toxicity of reactive oxygen species (ROS) formed inside cells. It was found that sublethal oxidative stress derived from photoexcited TiO 2 exerted a simulative effect on the growth of SOD null mutant of Escherichia coli (IM303) with reduction in intracellular ROS level. DNA microarray analysis was then carried out to compare gene expression between IM303 cells with and without the oxidative stress. From the DNA micro array data, yfiD, yggB and yggE were selected as genes up-regulated under the oxidative condition and then cloned into a pUC 19 plasmid. The original pUC 19 and constructed plasmids were introduced into E. coli MM294 and the transformants were cultivated in M9 medium with paraquat. Among these transformants, intracellular ROS content was the lowest in the cells carrying yggE gene and maximum specific growth rate of those cells was also higher than that of control cells with pUC 19. These results suggest that yggE gene product has an ROS-scavenging function in the cells of E. coli cells exposed to an oxidative stress and improves the efficiency of cellular growth.
Journal of Bacteriology, 1997
Escherichia coli produces an inducible set of proteins that protect the cell from exogenous peroxide stress. A subset of these genes is induced by hydrogen peroxide and is controlled at the transcriptional level by the OxyR protein. To identify additional genes involved in protection from hydrogen peroxide, a library of random transcriptional fusions of placMu53 was screened for hydrogen peroxide sensitivity and 27 such mutants were identified. These fusions were transduced into nonlysogenic strains to ensure that the phenotypes observed were the result of a single mutation. The mutants were grouped into three classes based on the expression of the lacZ fusion during growth in oxyR ؉ and ⌬oxyR backgrounds. The expression of the lacZ fusion in 8 mutants was independent of OxyR, 10 mutants required OxyR for expression, and 6 mutants showed reduced levels of expression in the presence of OxyR. OxyR dependence varied from 2-to 50-fold in these mutants. The OxyR-dependent phenotype was complemented by a plasmid-borne copy of oxyR gene in all mutants. Three mutants exhibited dual regulation by OxyR and RpoS. We sequenced the fusion junctions of several of these mutants and identified the genetic loci responsible for the hydrogen peroxide-sensitive (hps) phenotype. In this study, we report the identification of several genes that require OxyR for expression, including hemF (encoding coproporphyrinogen III oxidase), rcsC (encoding a sensor-regulator protein of capsular polysaccharide synthesis genes), and an open reading frame, f497, that is similar to arylsulfatase-encoding genes.
Nucleic acids research, 2014
In Escherichia coli, an increase in the ATP bound form of the DnaA initiator protein results in hyperinitiation and inviability. Here, we show that such replication stress is tolerated during anaerobic growth. In hyperinitiating cells, a shift from anaerobic to aerobic growth resulted in appearance of fragmented chromosomes and a decrease in terminus concentration, leading to a dramatic increase in ori/ter ratio and cessation of cell growth. Aerobic viability was restored by reducing the level of reactive oxygen species (ROS) or by deleting mutM (Fpg glycosylase). The double-strand breaks observed in hyperinitiating cells therefore results from replication forks encountering single-stranded DNA lesions generated while removing oxidized bases, primarily 8-oxoG, from the DNA. We conclude that there is a delicate balance between chromosome replication and ROS inflicted DNA damage so the number of replication forks can only increase when ROS formation is reduced or when the pertinent re...
Journal of Bacteriology
OxyR is a redox-sensitive transcriptional regulator of the LysR family which activates the expression of genes important for the defense against hydrogen peroxide in Escherichia coli and Samonella typhimurium. OxyR is sensitive to oxidation and reduction, and only oxidized OxyR is able to activate transcription of its target genes. Using site-directed mutagenesis, we found that one cysteine residue (C-199) is critical for the redox sensitivity of OxyR, and a C-199-->S mutation appears to lock the OxyR protein in the reduced form. We also used a random mutagenesis approach to isolate eight constitutively active mutants. All of the mutations are located in the C-terminal half of the protein, and four of the mutations map near the critical C-199 residue. In vivo as well as in vitro transcription experiments showed that the constitutive mutant proteins were able to activate transcription under both oxidizing and reducing conditions, and DNase I footprints showed that this activation ...