Hierarchy and interconnected networks in the WhiB7 mediated transcriptional response to antibiotic stress in...

Mycobacterium abscessus is intrinsically resistant to antibiotics effective against other pathogenic mycobacteria largely due to the drug-induced expression of genes that confer resistance. WhiB7 is a major hub controlling the induction of resistance to ribosome-targeting antibiotics. It activates the expression of >100 genes, 7 of which are known determinants of drug resistance; the function of most genes within the regulon is however unknown, but some conceivably encode additional mechanisms of resistance. Furthermore, the hierarchy of gene expression within the regulon, if any, is poorly understood. In the present work we have identified 56 WhiB7 binding sites using chromatin immunoprecipitation sequencing (CHIP-Seq) which accounts for the WhiB7-dependent upregulation of 72 genes, and find that M. abscessus WhiB7 functions exclusively as a transcriptional activator at promoters recognized by σA/σB. We have investigated the role of 18 WhiB7 regulated genes in drug resistance. Our results suggest that while some genes within the regulon (eg. erm41, hflX, eis2 and the ABCFs) play a major role in resistance, others make smaller contributions (eg. MAB_4324c and MAB_1409c) and the observed hypersensitivity ΔMabwhiB7 is a cumulative effect of these individual contributions. Moreover, our CHIP-Seq data implicate additional roles of WhiB7 induced genes beyond antibiotic resistance. Finally, we identify a σH-dependent network in aminoglycoside and tigecycline resistance which is induced upon drug exposure and is further activated by WhiB7 demonstrating the existence of a crosstalk between components of the WhiB7-dependent and -independent circuits.