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The GO system prevents ROS-induced mutagenesis and killing in Pseudomonas aeruginosa.

ABSTRACT: Inactivation of the Pseudomonas aeruginosa mutM, mutY, or mutT gene conferred a 2.4-, 17.2-, or 38.1-fold increase in spontaneous mutation frequency, respectively. Importantly, the mutY and mutT strains each displayed a robust H(2)O(2)-induced mutation frequency. In addition, the mutM, mutY, and mutT mutations severely sensitized P. aeruginosa to killing by H(2)O(2), suggesting that these gene products act to repair one or more cytotoxic lesions in P. aeruginosa. Nucleotide sequence analysis of a fragment of the rpoB gene from rifampicin resistant mutM-, mutY-, and, mutT-deficient strains was consistent with this conclusion. These findings are discussed in terms of possible roles for mutM, mutY, and mutT in contributing to survival and mutagenesis of P. aeruginosa colonizing the airways of cystic fibrosis patients.

SUBMITTER: Sanders LH

PROVIDER: S-EPMC2863324 | biostudies-literature | 2009 May

REPOSITORIES: biostudies-literature

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The GO system prevents ROS-induced mutagenesis and killing in Pseudomonas aeruginosa.

Sanders Laurie H LH Sudhakaran Julee J Sutton Mark D MD

FEMS microbiology letters 20090310 1

Inactivation of the Pseudomonas aeruginosa mutM, mutY, or mutT gene conferred a 2.4-, 17.2-, or 38.1-fold increase in spontaneous mutation frequency, respectively. Importantly, the mutY and mutT strains each displayed a robust H(2)O(2)-induced mutation frequency. In addition, the mutM, mutY, and mutT mutations severely sensitized P. aeruginosa to killing by H(2)O(2), suggesting that these gene products act to repair one or more cytotoxic lesions in P. aeruginosa. Nucleotide sequence analysis of ...[more]

PMID: 19291074

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Project description:Cystic fibrosis (CF) patients chronically infected with both Pseudomonas aeruginosa and Staphylococcus aureus have worse health outcomes than patients who are monoinfected with either P. aeruginosa or S. aureus We showed previously that mucoid strains of P. aeruginosa can coexist with S. aureusin vitro due to the transcriptional downregulation of several toxic exoproducts typically produced by P. aeruginosa, including siderophores, rhamnolipids, and HQNO (2-heptyl-4-hydroxyquinoline N-oxide). Here, we demonstrate that exogenous alginate protects S. aureus from P. aeruginosa in both planktonic and biofilm coculture models under a variety of nutritional conditions. S. aureus protection in the presence of exogenous alginate is due to the transcriptional downregulation of pvdA, a gene required for the production of the iron-scavenging siderophore pyoverdine as well as the downregulation of the PQS (Pseudomonas quinolone signal) (2-heptyl-3,4-dihydroxyquinoline) quorum sensing system. The impact of exogenous alginate is independent of endogenous alginate production. We further demonstrate that coculture of mucoid P. aeruginosa with nonmucoid P. aeruginosa strains can mitigate the killing of S. aureus by the nonmucoid strain of P. aeruginosa, indicating that the mechanism that we describe here may function in vivo in the context of mixed infections. Finally, we investigated a panel of mucoid clinical isolates that retain the ability to kill S. aureus at late time points and show that each strain has a unique expression profile, indicating that mucoid isolates can overcome the S. aureus-protective effects of mucoidy in a strain-specific manner.IMPORTANCE CF patients are chronically infected by polymicrobial communities. The two dominant bacterial pathogens that infect the lungs of CF patients are P. aeruginosa and S. aureus, with ∼30% of patients coinfected by both species. Such coinfected individuals have worse outcomes than monoinfected patients, and both species persist within the same physical space. A variety of host and environmental factors have been demonstrated to promote P. aeruginosa-S. aureus coexistence, despite evidence that P. aeruginosa kills S. aureus when these organisms are cocultured in vitro Thus, a better understanding of P. aeruginosa-S. aureus interactions, particularly mechanisms by which these microorganisms are able to coexist in proximal physical space, will lead to better-informed treatments for chronic polymicrobial infections.

Project description:Pseudomonas aeruginosa is a human opportunistic pathogen that chronically infects the lungs of cystic fibrosis patients and is the leading cause of morbidity and mortality of people afflicted with this disease. A striking correlation between mutagenesis and the persistence of P. aeruginosa has been reported. In other well-studied organisms, error-prone replication by Y family DNA polymerases contributes significantly to mutagenesis. Based on an analysis of the PAO1 genome sequence, P. aeruginosa contains a single Y family DNA polymerase encoded by the dinB gene. As part of an effort to understand the mechanisms of mutagenesis in P. aeruginosa, we have cloned the dinB gene of P. aeruginosa and utilized a combination of genetic and biochemical approaches to characterize the activity and regulation of the P. aeruginosa DinB protein (DinB(Pa)). Our results indicate that DinB(Pa) is a distributive DNA polymerase that lacks intrinsic proofreading activity in vitro. Modest overexpression of DinB(Pa) from a plasmid conferred a mutator phenotype in both Escherichia coli and P. aeruginosa. An examination of this mutator phenotype indicated that DinB(Pa) has a propensity to promote C-->A transversions and -1 frameshift mutations within poly(dGMP) and poly(dAMP) runs. The characterization of lexA+ and DeltalexA::aacC1 P. aeruginosa strains, together with in vitro DNA binding assays utilizing cell extracts or purified P. aeruginosa LexA protein (LexA(Pa)), indicated that the transcription of the dinB gene is regulated as part of an SOS-like response. The deletion of the dinB(Pa) gene sensitized P. aeruginosa to nitrofurazone and 4-nitroquinoline-1-oxide, consistent with a role for DinB(Pa) in translesion DNA synthesis over N2-dG adducts. Finally, P. aeruginosa exhibited a UV-inducible mutator phenotype that was independent of dinB(Pa) function and instead required polA and polC, which encode DNA polymerase I and the second DNA polymerase III enzyme, respectively. Possible roles of the P. aeruginosa dinB, polA, and polC gene products in mutagenesis are discussed.

Dataset Information

The GO system prevents ROS-induced mutagenesis and killing in Pseudomonas aeruginosa.

Publications

The GO system prevents ROS-induced mutagenesis and killing in Pseudomonas aeruginosa.

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