title: The two faces of ToxR: activator of ompU, co-regulator of toxT in Vibrio cholerae.
abstract: ToxR of Vibrio cholerae directly activates the ompU promoter, but requires a second activator, TcpP to activate the toxT promoter. ompU encodes a porin, while toxT encodes the transcription factor, ToxT, which activates V. cholerae virulence genes including cholera toxin and the toxin co-regulated pilus. Using an ompU-sacB transcriptional fusion, toxR mutant alleles were identified that encode ToxR molecules defective for ompU promoter activation. Many toxR mutants defective for ompU activation affected residues involved in DNA binding. Mutants defective for ompU activation were also tested for activation of the toxT promoter. ToxR-F69A and ToxR-V71A, both in the α-loop of ToxR, were preferentially defective for ompU activation, with ToxR-V71A nearly completely defective. Six mutants from the ompU-sacB selection showed more dramatic defects in toxT activation than ompU activation. All but one of the affected residues map to the wing domain of the winged helix-turn-helix of ToxR. Some ToxR mutants preferentially affecting toxT activation had partial DNA-binding defects, and one mutant, ToxR-P101L, had altered interactions with TcpP. These data suggest that while certain residues in the α-loop of ToxR are utilized to activate the ompU promoter, the wing domain of ToxR contributes to both promoter binding and ToxR/TcpP interaction facilitating toxT activation.
authors.affiliation: Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
authors.forename: Sarah J
authors.initials: SJ
authors.lastname: Morgan
authors.forename: Suleyman
authors.initials: S
authors.lastname: Felek
authors.forename: Shilpa
authors.initials: S
authors.lastname: Gadwal
authors.forename: Nicole M
authors.initials: NM
authors.lastname: Koropatkin
authors.forename: Jeffrey W
authors.initials: JW
authors.lastname: Perry
authors.forename: Alyson B
authors.initials: AB
authors.lastname: Bryson
authors.forename: Eric S
authors.initials: ES
authors.lastname: Krukonis
mesh_terms: D018829:Adhesins, Bacterial / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D015964:Gene Expression Regulation, Bacterial; D019077:Helix-Turn-Helix Motifs; D008954:Models, Biological; D008958:Models, Molecular; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011487:Protein Conformation; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014734:Vibrio cholerae / Q000235:genetics / Q000502:physiology*
chemical_list: D018829:Adhesins, Bacterial; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D050505:Mutant Proteins; C096349:OmpU protein, Vibrio cholerae; C097244:TCPP protein, Vibrio cholerae; D014157:Transcription Factors; C499324:toxR protein, Vibrio cholerae; C075626:tcpN protein, Vibrio cholerae