Transcriptional Regulation Mechanism Of MarR Family Transcriptional Regulator Ms6508 In Mycobacterium Smegmatis

Mycobacterium tuberculosis, the cause agent of Tuberculosis (TB), is a serious threat to human health. The emergence of multi-drug resistant (MDR) and extensively drug-resistant (XDR) M. tuberculosis strains in recent years have made more challenges in TB treatment. Exploring new drug resistance regulators and elucidating the underline regulational pathways can promote our understanding of molecular mechinisms involved in drug resistance and development of new anti-TB drugs.Mycobacterium smegmatis is a widely used model organism for study of M. tuberculosis. Our previous work found that overexpression of transcription factor Ms6508 could increase resistance of M. smegmatis to anti-TB drugs. In this study, we further investigate the potential regulational pathways of the Ms6508 that induce bacteria drug resistance. The main results include:(1) Ms6508 is a MarR family transcriptional regulator and locates in operon Ms6508-Ms6509-Ms6510 which contains drug-transported genes; (2) Ms6508 was found to bind to its own promoter in vitro by bacterial one-hybrid and electrophoretic mobility shift assay and in vivo by chromatin immunoprecipitation assay. The bound of Ms6508 with its promoter is inhibited by salicylic acid; (3) DNaseI footprinting assay indicated that Ms6508 binds to 21bp palindromic sequence (5’-ATACCTCTGTGACAGAGGTAT-3’):Bioinformatic analysis identified 454 potential target genes using this palindromic sequence; (4) p-galactosidase activity assay and qRT-PCR assay showed that Ms6508 negatively regulates the expression of its operon; (5) Overexpression of Ms6508 increased resistance of M. smegmatis to rifampicin and sensitivity to isoniazid; (6) Overexpression of Ms6509-Ms6510 promoted the rifampicin to enter into bacteria cells and increased the sensitivity of strains to rifampicin.In summary, this study identified a new drug-resistant transcriptional regulator Ms6508 and investigated its regulational pathways in M. smegmatis. These results promote our understanding on drug-resistant mechanisms in Mycobacterium.