Mutations in the MAB_2299c TetR Regulator Confer Cross-Resistance to Clofazimine and Bedaquiline in <i>Mycobacterium abscessus</i>

Abstract

New therapeutic approaches are needed against <i>Mycobacterium abscessus</i>, a respiratory mycobacterial pathogen that evades efforts to successfully treat infected patients. Clofazimine and bedaquiline, two drugs used for the treatment of multidrug-resistant tuberculosis, are being considered as alternatives for the treatment of lung diseases caused by <i>M. abscessus</i> With the aim to understand the mechanism of action of these agents in <i>M. abscessus</i>, we sought herein to determine the means by which <i>M. abscessus</i> can develop resistance. Spontaneous resistant strains selected on clofazimine, followed by whole-genome sequencing, identified mutations in <i>MAB_2299c</i>, encoding a putative TetR transcriptional regulator. Unexpectedly, mutants with these mutations were also cross-resistant to bedaquiline. MAB_2299c was found to bind to its target DNA, located upstream of the divergently oriented <i>MAB_2300-MAB_2301</i> gene cluster, encoding MmpS/MmpL membrane proteins. Point mutations or deletion of <i>MAB_2299c</i> was associated with the concomitant upregulation of the <i>mmpS</i> and <i>mmpL</i> transcripts and accounted for this cross-resistance. Strikingly, deletion of <i>MAB_2300</i> and <i>MAB_2301</i> in the <i>MAB_2299c</i> mutant strain restored susceptibility to bedaquiline and clofazimine. Overall, these results expand our knowledge with respect to the regulatory mechanisms of the MmpL family of proteins and a novel mechanism of drug resistance in this difficult-to-treat respiratory mycobacterial pathogen. Therefore, <i>MAB_2299c</i> may represent an important marker of resistance to be considered in the treatment of <i>M. abscessus</i> diseases with clofazimine and bedaquiline in clinical settings.