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NsaRS is a cell-envelope-stress-sensing two-component system of Staphylococcus aureus.

ABSTRACT: Staphylococcus aureus possesses 16 two-component systems (TCSs), two of which (GraRS and NsaRS) belong to the intramembrane-sensing histidine kinase (IM-HK) family, which is conserved within the firmicutes. NsaRS has recently been documented as being important for nisin resistance in S. aureus. In this study, we present a characterization of NsaRS and reveal that, as with other IM-HK TCSs, it responds to disruptions in the cell envelope. Analysis using a lacZ reporter-gene fusion demonstrated that nsaRS expression is upregulated by a variety of cell-envelope-damaging antibiotics, including phosphomycin, ampicillin, nisin, gramicidin, carbonyl cyanide m-chlorophenylhydrazone and penicillin G. Additionally, we reveal that NsaRS regulates a downstream transporter NsaAB during nisin-induced stress. NsaS mutants also display a 200-fold decreased ability to develop resistance to the cell-wall-targeting antibiotic bacitracin. Microarray analysis reveals that the transcription of 245 genes is altered in an nsaS mutant, with the vast majority being downregulated. Included within this list are genes involved in transport, drug resistance, cell envelope synthesis, transcriptional regulation, amino acid metabolism and virulence. Using inductively coupled plasma-MS we observed a decrease in intracellular divalent metal ions in an nsaS mutant when grown under low abundance conditions. Characterization of cells using electron microscopy reveals that nsaS mutants have alterations in cell envelope structure. Finally, a variety of virulence-related phenotypes are impaired in nsaS mutants, including biofilm formation, resistance to killing by human macrophages and survival in whole human blood. Thus, NsaRS is important in sensing cell damage in S. aureus and functions to reprogram gene expression to modify cell envelope architecture, facilitating adaptation and survival.

SUBMITTER: Kolar SL

PROVIDER: S-EPMC3167884 | biostudies-literature | 2011 Aug

REPOSITORIES: biostudies-literature

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NsaRS is a cell-envelope-stress-sensing two-component system of Staphylococcus aureus.

Kolar Stacey L SL Nagarajan Vijayaraj V Oszmiana Anna A Rivera Frances E FE Miller Halie K HK Davenport Jessica E JE Riordan James T JT Potempa Jan J Barber David S DS Koziel Joanna J Elasri Mohamed O MO Shaw Lindsey N LN Shaw Lindsey N LN

Microbiology (Reading, England) 20110512 Pt 8

Staphylococcus aureus possesses 16 two-component systems (TCSs), two of which (GraRS and NsaRS) belong to the intramembrane-sensing histidine kinase (IM-HK) family, which is conserved within the firmicutes. NsaRS has recently been documented as being important for nisin resistance in S. aureus. In this study, we present a characterization of NsaRS and reveal that, as with other IM-HK TCSs, it responds to disruptions in the cell envelope. Analysis using a lacZ reporter-gene fusion demonstrated th ...[more]

PMID: 21565927

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Project description:S. aureus possesses 16 two-component systems (TCS), two of which (GraRS & NsaRS) belong to the intramembrane-sensing histidine kinase (IM-HK) family, conserved within the firmicutes. NsaRS has recently been documented as being important for nisin-resistance in S. aureus. In this study we present a detailed characterization of NsaRS and reveal that, as with other IM-HK TCS, it responds to disruptions in cell-wall stability. Analysis using a lacZ reporter-gene fusion demonstrated that nsaRS expression is upregulated by a variety of cell-wall damaging antibiotics, including Phosphomycin, Ampicillin, Nisin and Penicillin G. Additionally; we reveal that NsaRS regulates a downstream transporter, NsaAB, during nisin-induced stress. NsaS mutants also display a 200-fold decreased ability to develop resistance to the cell-wall targetting antibiotic bacitracin. Microarray analysis reveals the transcription of 245 genes is altered in a nsaS mutant, with the vast majority down-regulated. Included within this list are genes involved in transport, drug-resistance, cell-envelope synthesis, transcriptional regulation, amino acid metabolism and virulence. Using ICP-MS we observed a decrease in intracellular divalent metal ions in an nsaS mutant when grown under low affinity conditions. Characterization of cells using electron microscopy reveals that nsaS mutants have alterations in both cell-wall and cell-envelope structure. Finally, a variety of virulence related phenotypes are impaired in nsaS mutants, including biofilm formation, resistance to killing by human macrophages and survival in whole human blood. Thus NsaRS is important in sensing cell-wall instability in S. aureus, and functions to reprogram gene expression to modify cell-envelope architecture, facilitating adaptation and survival. We did four hybridizations for this experiment, including a biological replicate and a dye-swap experiment for each replicate to account for dye-bias. Spots flagged as empty or bad were excluded and the raw data from each slide was normalized using LOWESS method, with background correction. The data from the replicates were combined (using the median value) and a one-sample t-test was performed. The volcano plot was used with a fold change cut-off of >= 2 and a p-value of <0.05, to filter the genes that were differentially expressed.

Project description:UnlabelledStaphylococcus aureus is both a commensal and a pathogen of the human host. Survival in the host environment requires resistance to host-derived nitric oxide (NO·). However, S. aureus lacks the NO·-sensing transcriptional regulator NsrR that is used by many bacteria to sense and respond to NO·. In this study, we show that S. aureus is able to sense and respond to both NO· and hypoxia by means of the SrrAB two-component system (TCS). Analysis of the S. aureus transcriptome during nitrosative stress demonstrates the expression of SrrAB-dependent genes required for cytochrome biosynthesis and assembly (qoxABCD, cydAB, hemABCX), anaerobic metabolism (pflAB, adhE, nrdDG), iron-sulfur cluster repair (scdA), and NO· detoxification (hmp). Targeted mutations in SrrAB-regulated loci show that hmp and qoxABCD are required for NO· resistance, whereas nrdDG is specifically required for anaerobic growth. We also show that SrrAB is required for survival in static biofilms, most likely due to oxygen limitation. Activation by hypoxia, NO·, or a qoxABCD quinol oxidase mutation suggests that the SrrAB TCS senses impaired electron flow in the electron transport chain rather than directly interacting with NO· in the manner of NsrR. Nevertheless, like NsrR, SrrAB achieves the physiological goals of selectively expressing hmp in the presence of NO· and minimizing the potential for Fenton chemistry. Activation of the SrrAB regulon allows S. aureus to maintain energy production and essential biosynthetic processes, repair damage, and detoxify NO· in diverse host environments.ImportanceThe Hmp flavohemoglobin is required for nitric oxide resistance and is widely distributed in bacteria. Hmp expression must be tightly regulated, because expression under aerobic conditions in the absence of nitric oxide can exacerbate oxidative stress. In most organisms, hmp expression is controlled by the Fe-S cluster-containing repressor NsrR, but this transcriptional regulator is absent in the human pathogen Staphylococcus aureus. We show here that S. aureus achieves hmp regulation in response to nitric oxide and oxygen limitation by placing it under the control of the SrrAB two-component system, which senses reduced electron flow through the respiratory chain. This provides a striking example of convergent evolution, in which the common physiological goals of responding to nitrosative stress while minimizing Fenton chemistry are achieved by distinct regulatory mechanisms.

Dataset Information

NsaRS is a cell-envelope-stress-sensing two-component system of Staphylococcus aureus.

Publications

NsaRS is a cell-envelope-stress-sensing two-component system of Staphylococcus aureus.

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