title: Transcriptional analysis of the grlRA virulence operon from Citrobacter rodentium.
abstract: The locus for enterocyte effacement (LEE) is the virulence hallmark of the attaching-and-effacing (A/E) intestinal pathogens, namely, enteropathogenic Escherichia coli, enterohemorrhagic E. coli, and Citrobacter rodentium. The LEE carries more than 40 genes that are arranged in several operons, e.g., LEE1 to LEE5. Expression of the various transcriptional units is subject to xenogeneic silencing by the histone-like protein H-NS. The LEE1-encoded regulator, Ler, plays a key role in relieving this repression at several major LEE promoters, including LEE2 to LEE5. To achieve appropriate intracellular concentrations of Ler in different environments, A/E pathogens have evolved a sophisticated regulatory network to control ler expression. For example, the LEE-encoded GrlA and GrlR proteins work as activator and antiactivator, respectively, of ler transcription. Thus, control of the transcriptional activities of the LEE1 (ler) promoter and the grlRA operon determines the rate of transcription of all of the LEE-encoded virulence factors. To date, only a single promoter has been identified for the grlRA operon. In this study, we showed that the non-LEE-encoded AraC-like regulatory protein RegA of C. rodentium directly stimulates transcription of the grlRA promoter by binding to an upstream region in the presence of bicarbonate ions. In addition, in vivo and in vitro transcription assays revealed a sigma(70) promoter that is specifically responsible for transcription of grlA. Expression from this promoter was strongly repressed by H-NS and its paralog StpA but was activated by Ler. DNase I footprinting demonstrated that Ler binds to a region upstream of the grlA promoter, whereas H-NS interacts specifically with a region extending from the grlA core promoter into its coding sequence. Together, these findings provide new insights into the environmental regulation and differential expressions of the grlR and grlA genes of C. rodentium.
authors.affiliation: Department of Microbiology and Immunology, the University of Melbourne, Parkville, Victoria 3010, Australia.
authors.forename: Marija
authors.initials: M
authors.lastname: Tauschek
authors.forename: Ji
authors.initials: J
authors.lastname: Yang
authors.forename: Dianna
authors.initials: D
authors.lastname: Hocking
authors.forename: Kristy
authors.initials: K
authors.lastname: Azzopardi
authors.forename: Aimee
authors.initials: A
authors.lastname: Tan
authors.forename: Emily
authors.initials: E
authors.lastname: Hart
authors.forename: Judyta
authors.initials: J
authors.lastname: Praszkier
authors.forename: Roy M
authors.initials: RM
authors.lastname: Robins-Browne
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D002874:Chromosome Mapping; D044082:Citrobacter rodentium / Q000235:genetics* / Q000378:metabolism* / Q000472:pathogenicity; D004269:DNA, Bacterial; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology; D008969:Molecular Sequence Data; D009876:Operon / Q000235:genetics*; D017693:Sodium Bicarbonate; D024363:Transcription Initiation Site; D014158:Transcription, Genetic; D014774:Virulence
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; C072722:RegA protein, Bacteria; D017693:Sodium Bicarbonate
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