title: A member of the cAMP receptor protein family of transcription regulators in Mycobacterium tuberculosis is required for virulence in mice and controls transcription of the rpfA gene coding for a resuscitation promoting factor.
abstract: Deletion of gene Rv3676 in Mycobacterium tuberculosis coding for a transcription factor belonging to the cAMP receptor protein (CRP) family caused growth defects in laboratory medium, in bone marrow-derived macrophages and in a mouse model of tuberculosis. Transcript profiling of M. tuberculosis grown in vitro identified 16 genes with significantly altered expression in the mutant compared with the wild type. Analysis of the DNA sequences upstream of the corresponding open reading frames revealed that 12 possessed sequences related to a consensus CRP binding site that could represent the sites of action of Rv3676. These included rpfA, lprQ, whiB1 and ahpC among genes with enhanced expression in the wild type, and Rv3616c-Rv3613c, Rv0188 and lipQ among genes exhibiting enhanced expression in the mutant. The activity of an rpfA::lacZ promoter fusion was lowered in the Rv3676 mutant and by mutation of the predicted Rv3676 binding site. Moreover, the product of Rv3676 (isolated as a TrxA fusion protein) interacted specifically with the rpfA promoter, and binding was inhibited by mutation of the Rv3676 site. Although Rv3676 retains four of the six amino acid residues that bind cAMP in Escherichia coli CRP addition of cAMP did not enhance Rv3676 binding at the rpfA promoter in vitro. In summary, it has been shown that Rv3676 is a direct regulator of rpfA expression, and because rpfA codes for a resuscitation promoting factor this may implicate Rv3676 in reactivation of dormant M. tuberculosis infections.
authors.affiliation: Division of Mycobacterial Research, National Institute for Medical Research, Mill Hill, London NW7 1AA, UK.
authors.forename: Lisa
authors.initials: L
authors.lastname: Rickman
authors.forename: Colin
authors.initials: C
authors.lastname: Scott
authors.forename: Debbie M
authors.initials: DM
authors.lastname: Hunt
authors.forename: Thomas
authors.initials: T
authors.lastname: Hutchinson
authors.forename: M Carmen
authors.initials: MC
authors.lastname: Menéndez
authors.forename: Rachael
authors.initials: R
authors.lastname: Whalan
authors.forename: Jason
authors.initials: J
authors.lastname: Hinds
authors.forename: M Joseph
authors.initials: MJ
authors.lastname: Colston
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Green
authors.forename: Roger S
authors.initials: RS
authors.lastname: Buxton
mesh_terms: D000154:Aconitate Hydratase / Q000096:biosynthesis* / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D019677:Artificial Gene Fusion; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics; D015169:Colony Count, Microbial; D002373:Cyclic AMP Receptor Protein / Q000235:genetics / Q000502:physiology*; D004195:Disease Models, Animal; D005260:Female; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial / Q000502:physiology; D017930:Genes, Reporter / Q000502:physiology; D007763:Lac Operon / Q000502:physiology; D008168:Lung / Q000382:microbiology; D008264:Macrophages / Q000382:microbiology; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000254:growth & development / Q000472:pathogenicity*; D020411:Oligonucleotide Array Sequence Analysis; D012045:Regulatory Sequences, Nucleic Acid; D013154:Spleen / Q000382:microbiology; D014158:Transcription, Genetic; D014376:Tuberculosis / Q000382:microbiology; D014774:Virulence / Q000235:genetics; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D002373:Cyclic AMP Receptor Protein; D001616:beta-Galactosidase; D000154:Aconitate Hydratase; C105962:RpfA protein, bacteria