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AcrAB multidrug efflux pump regulation in Salmonella enterica serovar Typhimurium by RamA in response to environmental signals.

Nikaido E, Yamaguchi A, Nishino K - J. Biol. Chem. (2008)

Bottom Line: Among these pumps, AcrAB is effective in generating drug resistance and has wide substrate specificity.Other regulators of acrAB such as MarA, SoxS, Rob, SdiA, and AcrR did not contribute to acrAB induction by indole in Salmonella.Our results suggest that RamA controls the Salmonella AcrAB-TolC multidrug efflux system through dual regulatory modes in response to environmental signals.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Membrane Biology, Institute of Scientific and Industrial Research, Osaka University, Osaka, Japan.

ABSTRACT
Salmonella enterica serovar Typhimurium has at least nine multidrug efflux pumps. Among these pumps, AcrAB is effective in generating drug resistance and has wide substrate specificity. Here we report that indole, bile, and an Escherichia coli conditioned medium induced the AcrAB pump in Salmonella through a specific regulator, RamA. The RamA-binding sites were located in the upstream regions of acrAB and tolC. RamA was required for indole induction of acrAB. Other regulators of acrAB such as MarA, SoxS, Rob, SdiA, and AcrR did not contribute to acrAB induction by indole in Salmonella. Indole activated ramA transcription, and overproduction of RamA caused increased acrAB expression. In contrast, induction of ramA was not required for induction of acrAB by bile. Cholic acid binds to RamA, and we suggest that bile acts by altering pre-existing RamA. This points to two different AcrAB regulatory modes through RamA. Our results suggest that RamA controls the Salmonella AcrAB-TolC multidrug efflux system through dual regulatory modes in response to environmental signals.

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Indole activation of acrAB expression through the RamA regulator. The data correspond to mean values from three independent experiments. Bars correspond to the standard deviation. Asterisks indicate statistically significant differences (*, p < 0.01) in the paired Student's t test. A, β-galactosidase levels in WT or ΔbaeSR cpxAR strains, carrying acrAB-lac and acrD-lac transcriptional fusions, grown in LB medium with (solid bars) or without (open bars) 2 mm indole. B, β-galactosidase levels were assayed in strains carrying the acrAB-lac transcriptional fusion in the WT (NKS505), ΔmarA (NES20), Δrob (NES29), ΔsoxS (NES28), ΔsdiA (NES35), ΔacrR (NES48), ΔmarA rob soxS sdiA acrR (NES55), and ΔramA (NES58) strains. Strains were grown in LB medium with (solid bars) or without (open bars) 2 mm indole.
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fig2: Indole activation of acrAB expression through the RamA regulator. The data correspond to mean values from three independent experiments. Bars correspond to the standard deviation. Asterisks indicate statistically significant differences (*, p < 0.01) in the paired Student's t test. A, β-galactosidase levels in WT or ΔbaeSR cpxAR strains, carrying acrAB-lac and acrD-lac transcriptional fusions, grown in LB medium with (solid bars) or without (open bars) 2 mm indole. B, β-galactosidase levels were assayed in strains carrying the acrAB-lac transcriptional fusion in the WT (NKS505), ΔmarA (NES20), Δrob (NES29), ΔsoxS (NES28), ΔsdiA (NES35), ΔacrR (NES48), ΔmarA rob soxS sdiA acrR (NES55), and ΔramA (NES58) strains. Strains were grown in LB medium with (solid bars) or without (open bars) 2 mm indole.

Mentions: Indole Induces acrAB Expression via the RamA Regulator—Among the multidrug efflux pumps, AcrAB plays a major role in the intrinsic resistance of Salmonella (25). Also, Hirakawa et al. (35) reported that the baeSR and cpxAR signal transduction system genes are required for indole induction of multidrug efflux pumps in E. coli. To identify the regulatory elements that induce acrAB in response to indole in Salmonella, we constructed a mutant that lacked baeSR and cpxAR. In the ΔbaeSR cpxAR mutant, the expression of acrAB was not significantly different from that in the wild-type (WT) strain; however, indole induction of acrD was significantly lower in the mutant compared with the WT strain (Fig. 2A). The result indicates that the BaeSR and CpxAR signal transduction systems are not involved in indole induction of acrAB, whereas they are required for acrD induction. Other regulators, marA, soxS, rob, sdiA, and acrR, have been previously reported to control acrAB expression in E. coli (27). With the exception of ramA, none significantly altered the indole induction of acrAB in Salmonella (Fig. 2B). The stimulatory effect of indole on acrAB expression was completely eliminated in the ΔramA mutant (Fig. 2B). The results indicate that the RamA regulator is required for indole induction of acrAB in Salmonella.


AcrAB multidrug efflux pump regulation in Salmonella enterica serovar Typhimurium by RamA in response to environmental signals.

Nikaido E, Yamaguchi A, Nishino K - J. Biol. Chem. (2008)

Indole activation of acrAB expression through the RamA regulator. The data correspond to mean values from three independent experiments. Bars correspond to the standard deviation. Asterisks indicate statistically significant differences (*, p < 0.01) in the paired Student's t test. A, β-galactosidase levels in WT or ΔbaeSR cpxAR strains, carrying acrAB-lac and acrD-lac transcriptional fusions, grown in LB medium with (solid bars) or without (open bars) 2 mm indole. B, β-galactosidase levels were assayed in strains carrying the acrAB-lac transcriptional fusion in the WT (NKS505), ΔmarA (NES20), Δrob (NES29), ΔsoxS (NES28), ΔsdiA (NES35), ΔacrR (NES48), ΔmarA rob soxS sdiA acrR (NES55), and ΔramA (NES58) strains. Strains were grown in LB medium with (solid bars) or without (open bars) 2 mm indole.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2527123&req=5

fig2: Indole activation of acrAB expression through the RamA regulator. The data correspond to mean values from three independent experiments. Bars correspond to the standard deviation. Asterisks indicate statistically significant differences (*, p < 0.01) in the paired Student's t test. A, β-galactosidase levels in WT or ΔbaeSR cpxAR strains, carrying acrAB-lac and acrD-lac transcriptional fusions, grown in LB medium with (solid bars) or without (open bars) 2 mm indole. B, β-galactosidase levels were assayed in strains carrying the acrAB-lac transcriptional fusion in the WT (NKS505), ΔmarA (NES20), Δrob (NES29), ΔsoxS (NES28), ΔsdiA (NES35), ΔacrR (NES48), ΔmarA rob soxS sdiA acrR (NES55), and ΔramA (NES58) strains. Strains were grown in LB medium with (solid bars) or without (open bars) 2 mm indole.
Mentions: Indole Induces acrAB Expression via the RamA Regulator—Among the multidrug efflux pumps, AcrAB plays a major role in the intrinsic resistance of Salmonella (25). Also, Hirakawa et al. (35) reported that the baeSR and cpxAR signal transduction system genes are required for indole induction of multidrug efflux pumps in E. coli. To identify the regulatory elements that induce acrAB in response to indole in Salmonella, we constructed a mutant that lacked baeSR and cpxAR. In the ΔbaeSR cpxAR mutant, the expression of acrAB was not significantly different from that in the wild-type (WT) strain; however, indole induction of acrD was significantly lower in the mutant compared with the WT strain (Fig. 2A). The result indicates that the BaeSR and CpxAR signal transduction systems are not involved in indole induction of acrAB, whereas they are required for acrD induction. Other regulators, marA, soxS, rob, sdiA, and acrR, have been previously reported to control acrAB expression in E. coli (27). With the exception of ramA, none significantly altered the indole induction of acrAB in Salmonella (Fig. 2B). The stimulatory effect of indole on acrAB expression was completely eliminated in the ΔramA mutant (Fig. 2B). The results indicate that the RamA regulator is required for indole induction of acrAB in Salmonella.

Bottom Line: Among these pumps, AcrAB is effective in generating drug resistance and has wide substrate specificity.Other regulators of acrAB such as MarA, SoxS, Rob, SdiA, and AcrR did not contribute to acrAB induction by indole in Salmonella.Our results suggest that RamA controls the Salmonella AcrAB-TolC multidrug efflux system through dual regulatory modes in response to environmental signals.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Membrane Biology, Institute of Scientific and Industrial Research, Osaka University, Osaka, Japan.

ABSTRACT
Salmonella enterica serovar Typhimurium has at least nine multidrug efflux pumps. Among these pumps, AcrAB is effective in generating drug resistance and has wide substrate specificity. Here we report that indole, bile, and an Escherichia coli conditioned medium induced the AcrAB pump in Salmonella through a specific regulator, RamA. The RamA-binding sites were located in the upstream regions of acrAB and tolC. RamA was required for indole induction of acrAB. Other regulators of acrAB such as MarA, SoxS, Rob, SdiA, and AcrR did not contribute to acrAB induction by indole in Salmonella. Indole activated ramA transcription, and overproduction of RamA caused increased acrAB expression. In contrast, induction of ramA was not required for induction of acrAB by bile. Cholic acid binds to RamA, and we suggest that bile acts by altering pre-existing RamA. This points to two different AcrAB regulatory modes through RamA. Our results suggest that RamA controls the Salmonella AcrAB-TolC multidrug efflux system through dual regulatory modes in response to environmental signals.

Show MeSH
Related in: MedlinePlus