title: Regulatory Characteristics of Vibrio vulnificus gbpA Gene Encoding a Mucin-binding Protein Essential for Pathogenesis.
abstract: Binding to mucin is the initial step for enteropathogens to establish pathogenesis. An open reading frame, gbpA, of Vibrio vulnificus was identified and characterized in this study. Compared with wild type, the gbpA mutant was impaired in binding to mucin-agar and the mucin-secreting HT29-methotrexate cells, and the impaired mucin binding was restored by the purified GbpA provided exogenously. The gbpA mutant had attenuated virulence and ability of intestinal colonization in a mouse model, indicating that GbpA is a mucin-binding protein and essential for pathogenesis of V. vulnificus. The gbpA transcription was growth phase-dependent, reaching a maximum during the exponential phase. The Fe-S cluster regulator (IscR) and the cyclic AMP receptor protein (CRP) coactivated, whereas SmcR, a LuxR homologue, repressed gbpA. The cellular levels of IscR, CRP, and SmcR were not significantly affected by one another, indicating that the regulator proteins function cooperatively to regulate gbpA rather than sequentially in a regulatory cascade. The regulatory proteins directly bind upstream of the gbpA promoter PgbpA. DNase I protection assays, together with the deletion analyses of PgbpA, demonstrated that IscR binds to two specific sequences centered at -164.5 and -106, and CRP and SmcR bind specifically to the sequences centered at -68 and -45, respectively. Furthermore, gbpA was induced by exposure to H2O2, and the induction appeared to be mediated by elevated intracellular levels of IscR. Consequently, the combined results indicated that IscR, CRP, and SmcR cooperate for precise regulation of gbpA during the V. vulnificus pathogenesis.
authors.affiliation: From the National Research Laboratory of Molecular Microbiology and Toxicology, Department of Agricultural Biotechnology, Center for Food Safety and Toxicology, Seoul National University, Seoul 151-921, South Korea.
authors.forename: Kyung Ku
authors.initials: KK
authors.lastname: Jang
authors.affiliation: From the National Research Laboratory of Molecular Microbiology and Toxicology, Department of Agricultural Biotechnology, Center for Food Safety and Toxicology, Seoul National University, Seoul 151-921, South Korea.
authors.forename: So Yeon
authors.initials: SY
authors.lastname: Gil
authors.affiliation: From the National Research Laboratory of Molecular Microbiology and Toxicology, Department of Agricultural Biotechnology, Center for Food Safety and Toxicology, Seoul National University, Seoul 151-921, South Korea.
authors.forename: Jong Gyu
authors.initials: JG
authors.lastname: Lim
authors.affiliation: From the National Research Laboratory of Molecular Microbiology and Toxicology, Department of Agricultural Biotechnology, Center for Food Safety and Toxicology, Seoul National University, Seoul 151-921, South Korea. Electronic address: choish@snu.ac.kr.
authors.forename: Sang Ho
authors.initials: SH
authors.lastname: Choi
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D005260:Female; D015964:Gene Expression Regulation, Bacterial; D007413:Intestinal Mucosa / Q000378:metabolism; D007422:Intestines / Q000382:microbiology / Q000473:pathology; D051379:Mice; D008813:Mice, Inbred ICR; D008969:Molecular Sequence Data; D009077:Mucins / Q000378:metabolism*; D009154:Mutation; D018384:Oxidative Stress; D011485:Protein Binding; D053038:Quorum Sensing; D024363:Transcription Initiation Site; D014735:Vibrio Infections / Q000378:metabolism* / Q000382:microbiology / Q000473:pathology*; D041261:Vibrio vulnificus / Q000235:genetics* / Q000502:physiology*
chemical_list: D001426:Bacterial Proteins; D009077:Mucins
keywords: CRP; GbpA; IscR; LuxR homologue; Vibrio vulnificus; bacterial adhesion; gene regulation; mucin; oxidative stress; quorum sensing