title: Cooperative regulation of the Vibrio vulnificus nan gene cluster by NanR protein, cAMP receptor protein, and N-acetylmannosamine 6-phosphate.
abstract: The nan cluster of Vibrio vulnificus, a food-borne pathogen, consists of two divergently transcribed operons, nanT(PSL)AR and nanEK nagA, required for transport and catabolism of N-acetylneuraminic acid (Neu5Ac). A mutation of nanR abolished the extensive lag phase observed for the bacteria growing on Neu5Ac and increased transcription of nanT(P) and nanE, suggesting that NanR is a transcriptional repressor of both nan operons. Intracellular accumulation of Neu5Ac was dependent on the carbon source, implying that the nan operons are also subject to catabolite repression. Hence, cAMP receptor protein (CRP) appeared to activate and repress transcription of nanT(PSL)AR and nanEK nagA, respectively. Direct bindings of NanR and CRP to the nanT(P)-nanE intergenic DNA were demonstrated by EMSA. Two adjacent NanR-binding sites centered at +44.5 and -10 and a CRP-binding site centered at -60.5 from the transcription start site of nanT(P) were identified by DNase I protection assays. Mutagenesis approaches, in vitro transcription, and isothermal titration calorimetry experiments demonstrated that N-acetylmannosamine 6-phosphate specifically binds to NanR and functions as the inducer of the nan operons. The combined results propose a model in which NanR, CRP, and N-acetylmannosamine 6-phosphate cooperate for precise adjustment of the expression level of the V. vulnificus nan cluster.
authors.affiliation: National Research Laboratory of Molecular Microbiology and Toxicology, Department of Agricultural Biotechnology, Center for Food Safety and Toxicology, and Research Institute for Agriculture and Life Sciences, Seoul National University, Seoul 151-921, South Korea.
authors.forename: Byoung Sik
authors.initials: BS
authors.lastname: Kim
authors.forename: Jungwon
authors.initials: J
authors.lastname: Hwang
authors.forename: Myung Hee
authors.initials: MH
authors.lastname: Kim
authors.forename: Sang Ho
authors.initials: SH
authors.lastname: Choi
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D002373:Cyclic AMP Receptor Protein / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000378:metabolism; D005798:Genes, Bacterial / Q000235:genetics*; D006595:Hexosamines / Q000378:metabolism*; D005810:Multigene Family / Q000235:genetics*; D009154:Mutation; D019158:N-Acetylneuraminic Acid / Q000378:metabolism; D009838:Oligodeoxyribonucleotides / Q000235:genetics / Q000378:metabolism; D009876:Operon / Q000235:genetics; D010710:Phosphates / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D013379:Substrate Specificity; D013403:Sugar Phosphates / Q000378:metabolism*; D014158:Transcription, Genetic; D041261:Vibrio vulnificus / Q000235:genetics* / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D002373:Cyclic AMP Receptor Protein; D004269:DNA, Bacterial; D006595:Hexosamines; C569296:N-acetylmannosamine 6-phosphate; D009838:Oligodeoxyribonucleotides; D010710:Phosphates; D013403:Sugar Phosphates; D003850:Deoxyribonuclease I; D019158:N-Acetylneuraminic Acid