Transcriptional and proteomic analyses of two-component response regulators in multidrug-resistant Mycobacterium tuberculosis

• We comprehensively determined the expression of 13 response regulators of two-component systems (TCSs) in Mycobacterium tuberculosis clinical isolates.
• Four regulator genes (devR, mtrA, regX3 and Rv3143) are significantly upregulated in multidrug-resistant tuberculosis (MDR-TB) strains; no mutation was found in the promoter regions of these four genes.
• Upregulation of these genes in MDR strains is not derived from different genotypes of clinical strains.
• The four genes can be induced by first-line antitubercular agents.
• Drug susceptibility testing of devR deletion and devR overexpression strains showed that upregulation of devR is not one of the reasons for multidrug resistance, but might be induced by drug stress and hypoxic adaptation after acquisition of multidrug resistance.

Two-component systems (TCSs) have been reported to exhibit a sensing and responding role under drug stress that induces drug resistance in several bacterial species. However, the relationship between TCSs and multidrug resistance in Mycobacterium tuberculosis has not been comprehensively analysed to date. In this study, 90 M. tuberculosis clinical isolates were analysed using 15-loci mycobacterial interspersed repetitive unit (MIRU)-variable number tandem repeat (VNTR) typing and repetitive extragenic palindromic (rep)-PCR-based DNA fingerprinting. The results showed that all of the isolates were of the Beijing lineage, and strains with a drug-susceptible phenotype had not diverged into similar genotype clusters. Expression analysis of 13 response regulators of TCSs using real-time PCR and tandem mass spectrometry (MS/MS) proteomic analysis demonstrated that four response regulator genes (devR, mtrA, regX3 and Rv3143) were significantly upregulated in multidrug-resistant (MDR) strains compared with the laboratory strain H37Rv as well as drug-susceptible and isoniazid-monoresistant strains (P < 0.05). DNA sequencing revealed that the promoter regions of devR, mtrA, regX3 and Rv3143 did not contain any mutations. Moreover, expression of the four genes could be induced by most of the four first-line antitubercular agents. In addition, either deletion or overexpression of devR in Mycobacterium bovis BCG did not alter its sensitivity to the four antitubercular drugs. This suggests that upregulation of devR, which is common in MDR-TB strains, might be induced by drug stress and hypoxic adaptation following the acquisition of multidrug resistance.