The impact of mab_1638 on the M. abscessus transcriptome [ChIP-seq]

Mycobacterium abscessus is a major human pathogen, mostly infecting people with pre-existing pathological lung conditions such as cystic fibrosis. Only little is known about the ways M. abscessus regulates its virulence factors and pathogenesis phenotypes. The production of glycopeptidolipids (GPL) is a major determinant of virulence of this bacterium, with clinical isolates that lack GPL synthesis and export generally exhibiting more aggressive clinical behavior. The current paradigm is that GPL production is abolished in vivo via irreversible, spontaneous mutations taking place as part of in-host evolution. Little is known about the mechanisms or extent to which GPL production may be regulated. We have found an unusual TetR-like transcription factor of M. abscessus, mab_1638, identified via screening of a comprehensive transposon-mutant library, that appears to be a strong positive regulator of the entire GPL biosynthesis and export gene cluster through a combination of direct and indirect mechanisms. The inactivation of mab_1638 by a transposon led to stable and visually obvious rough colony morphology, as well as increased virulence in infection models, characteristic of rough, non-GPL-producers. Transcriptome analysis found the mab_1638:tn mutant to have 118 differentially expressed genes, including the GPL locus. It appears mab_1638 encodes an unusual TetR transcription factor required for the production of GPL in M. abscessus and therefore having a profound effect on virulence traits. This finding raises the important possibility that M. abscessus strains appearing smooth in laboratory growth conditions may nonetheless downregulate GPL-cluster genes in other conditions and thus acquire the phenotypic characteristics of rough strains. Overall design: ChIP-seq was performed with an anti-HA antibody on four strains: (1) WT M. abscessus ATCC19977; (2) M. abscessus ATCC19977 transformed with a construct encoding an HA-tagged M. tuberculosis protein (mmaA2) not expected to bind DNA; (3) M. abscessus ATCC19977 with a transposon disrupting the gene mab_1638 (mab_1638::tn) and transformed with a construct encoding MAB_1638 with an N-terminal HA tag; and (4) mab_1638::tn transformed with a construct encoding MAB_1638 with an C-terminal HA tag.