title: Bile-mediated activation of the acrAB and tolC multidrug efflux genes occurs mainly through transcriptional derepression of ramA in Salmonella enterica serovar Typhimurium.
abstract: OBJECTIVE
  In Salmonella Typhimurium, the genes encoding the AcrAB-TolC multidrug efflux system are mainly regulated by the ramRA locus, composed of the divergently transcribed ramA and ramR genes. The acrAB and tolC genes are transcriptionally activated by RamA, the gene for which is itself transcriptionally repressed by RamR. Previous studies have reported that bile induces acrAB in a ramA-dependent manner, but none provided evidence for an induction of ramA expression by bile. Therefore, the objective of this study was to clarify the regulatory mechanism by which bile activates acrAB and tolC.
  
  
  METHODS
  qRT-PCR was used to address the effects of bile (using choleate, an ox-bile extract) on the expression of ramA, ramR, acrB and tolC. Electrophoretic mobility shift assays and surface plasmon resonance experiments were used to measure the effect of bile on RamR binding to the ramA promoter (PramA) region.
  
  
  RESULTS
  We show that ramA is transcriptionally activated by bile and is strictly required for the bile-mediated activation of acrB and tolC. Additionally, bile is shown to specifically inhibit the binding of RamR to the PramA region, which overlaps the putative divergent ramR promoter, thereby explaining our observation that bile also activates ramR transcription.
  
  
  CONCLUSIONS
  We propose a regulation model whereby the bile-mediated activation of the acrAB and tolC multidrug efflux genes occurs mainly through the transcriptional derepression of the ramA activator gene.
authors.affiliation: INRA, UMR1282 Infectiologie et Santé Publique, F-37380 Nouzilly, France Université François Rabelais de Tours, UMR1282 Infectiologie et Santé Publique, F-37000 Tours, France.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Baucheron
authors.affiliation: Laboratory of Microbiology and Infectious Diseases, Institute of Scientific and Industrial Research, Osaka University, Ibaraki, Osaka, Japan.
authors.forename: Kunihiko
authors.initials: K
authors.lastname: Nishino
authors.affiliation: INRA, UMR1282 Infectiologie et Santé Publique, F-37380 Nouzilly, France Université François Rabelais de Tours, UMR1282 Infectiologie et Santé Publique, F-37000 Tours, France.
authors.forename: Isabelle
authors.initials: I
authors.lastname: Monchaux
authors.affiliation: INRA, UMR7247, Plateforme d'Analyse Intégrative des Biomolécules et de Phénomique des Animaux d'Intérêt Bio-agronomique, Nouzilly, France.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Canepa
authors.affiliation: INRA, UMR7247, Plateforme d'Analyse Intégrative des Biomolécules et de Phénomique des Animaux d'Intérêt Bio-agronomique, Nouzilly, France.
authors.forename: Marie-Christine
authors.initials: MC
authors.lastname: Maurel
authors.affiliation: Centre de Biophysique Moléculaire CNRS, UPR4301, Orléans, France.
authors.forename: Franck
authors.initials: F
authors.lastname: Coste
authors.affiliation: Centre de Biophysique Moléculaire CNRS, UPR4301, Orléans, France.
authors.forename: Alain
authors.initials: A
authors.lastname: Roussel
authors.affiliation: INRA, UMR1282 Infectiologie et Santé Publique, F-37380 Nouzilly, France Université François Rabelais de Tours, UMR1282 Infectiologie et Santé Publique, F-37000 Tours, France.
authors.forename: Axel
authors.initials: A
authors.lastname: Cloeckaert
authors.affiliation: INRA, UMR1282 Infectiologie et Santé Publique, F-37380 Nouzilly, France Université François Rabelais de Tours, UMR1282 Infectiologie et Santé Publique, F-37000 Tours, France etienne.giraud@tours.inra.fr.
authors.forename: Etienne
authors.initials: E
authors.lastname: Giraud
mesh_terms: D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000378:metabolism*; D001646:Bile / Q000378:metabolism*; D002352:Carrier Proteins / Q000096:biosynthesis*; D004269:DNA, Bacterial / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects*; D006801:Humans; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000096:biosynthesis* / Q000378:metabolism*; D011485:Protein Binding; D060888:Real-Time Polymerase Chain Reaction; D012486:Salmonella typhimurium / Q000187:drug effects* / Q000235:genetics*; D020349:Surface Plasmon Resonance; D015534:Trans-Activators / Q000096:biosynthesis* / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; D004269:DNA, Bacterial; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D015534:Trans-Activators; C493251:AcrAB-TolC protein, Salmonella enterica; C531999:RamA protein, Salmonella typhimurium; C000596674:RamR protein, Salmonella typhimurium
keywords: promoters; ramR; surface plasmon resonance; transcriptional regulation