title: Functional analysis of the EspR binding sites upstream of espR in Mycobacterium tuberculosis.
abstract: The ESX-1 secretion system exports substrate proteins into host cells and is crucial for the pathogenesis of Mycobacterium tuberculosis. EspR is one of the characterized transcriptional regulators that modulates the ESX-1 system by binding the conserved EspR binding sites in the promoter of espA, the encoding gene of EspA, which is also a substrate protein of the ESX-1 system and is required for the ESX-1 activity. EspR is autoregulatory and conserved EspR binding sites are present upstream of espR. In this study, we showed that these EspR sites had varying affinities for EspR, with site B being the strongest one. Point mutations of the DNA sequence at site B abolished binding of EspR to oligonucleotides containing site B alone or with other sites, further suggesting that site B is a major binding site for EspR. Complementation studies showed that constructs containing espR, and the upstream intergenic region fully restored espR expression in a ΔespR mutant strain. Although recombinant strains with mutations at more than one EspR site showed minimal differences in espR expression, reduced expression of other EspR target genes was observed, suggesting that slight changes in EspR levels can have downstream regulatory effects. These findings contribute to our understanding of the regulation of the ESX-1 system.
authors.affiliation: The State Key Laboratory of Microbial Technology, Shandong University, Jinan, 250100, China.
authors.forename: Guangxiang
authors.initials: G
authors.lastname: Cao
authors.forename: Susan T
authors.initials: ST
authors.lastname: Howard
authors.forename: Peipei
authors.initials: P
authors.lastname: Zhang
authors.forename: Guihua
authors.initials: G
authors.lastname: Hou
authors.forename: Xiuhua
authors.initials: X
authors.lastname: Pang
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D002874:Chromosome Mapping; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000235:genetics* / Q000378:metabolism*; D009876:Operon; D011401:Promoter Regions, Genetic; D024363:Transcription Initiation Site; D014158:Transcription, Genetic
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins