title: The LuxR-type regulator VpsT negatively controls the transcription of rpoS, encoding the general stress response regulator, in Vibrio cholerae biofilms.
abstract: Cholera is a waterborne diarrheal disease caused by Vibrio cholerae strains of serogroups O1 and O139. Expression of the general stress response regulator RpoS and formation of biofilm communities enhance the capacity of V. cholerae to persist in aquatic environments. The transition of V. cholerae between free-swimming (planktonic) and biofilm life-styles is regulated by the second messenger cyclic di-GMP (c-di-GMP). We previously reported that increasing the c-di-GMP pool by overexpression of a diguanylate cyclase diminished RpoS expression. Here we show that c-di-GMP repression of RpoS expression is eliminated by deletion of the genes vpsR and vpsT, encoding positive regulators of biofilm development. To determine the mechanism of this regulation, we constructed a strain expressing a vpsT-FLAG allele from native transcription and translation signals. Increasing the c-di-GMP pool induced vpsT-FLAG expression. The interaction between VpsT-FLAG and the rpoS promoter was demonstrated by chromatin immunoprecipitation. Furthermore, purified VpsT interacted with the rpoS promoter in a c-di-GMP-dependent manner. Primer extension analysis identified two rpoS transcription initiation sites located 43 bp (P1) and 63 bp (P2) upstream of the rpoS start codon. DNase I footprinting showed that the VpsT binding site at the rpoS promoter overlaps the primary P1 transcriptional start site. Deletion of vpsT significantly enhanced rpoS expression in V. cholerae biofilms that do not make HapR. This result suggests that VpsT and c-di-GMP contribute to the transcriptional silencing of rpoS in biofilms prior to cells entering the quorum-sensing mode.
authors.affiliation: Morehouse School of Medicine, Department of Microbiology, Biochemistry and Immunology, Atlanta, Georgia, USA.
authors.forename: Hongxia
authors.initials: H
authors.lastname: Wang
authors.forename: Julio C
authors.initials: JC
authors.lastname: Ayala
authors.forename: Jorge A
authors.initials: JA
authors.lastname: Benitez
authors.forename: Anisia J
authors.initials: AJ
authors.lastname: Silva
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D018441:Biofilms / Q000254:growth & development*; D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism*; D013312:Stress, Physiological / Q000502:physiology; D014158:Transcription, Genetic / Q000502:physiology; D014734:Vibrio cholerae / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*
chemical_list: D001426:Bacterial Proteins; D012808:Sigma Factor; C069567:sigma factor KatF protein, Bacteria; C062025:bis(3',5')-cyclic diguanylic acid; D006152:Cyclic GMP