Transcriptional Regulation of the Mycobacterial Weapons System

Tuberculosis, and its causative agent Mycobacterium tuberculosis (M. tb), pose a significant health burden, killing over 1.5 million people every year. M. tb is successful as a pathogen due to its ability to overcome host immune cells by permeabilizing phagosomal membranes. Pathogenic mycobacteria permeabilize phagosomal membranes using the ESX-1 protein secretion system, a complex bacterial weapons system. The ESX-1 secretory apparatus secretes protein substrates across the mycobacterial cytoplasmic membrane. We discovered a role for the ESX-1 system in regulating gene expression. We discovered a mechanism allowing mycobacteria to tune the levels of ESX-1 substrates. We demonstrated that ESX-1 modulates the levels of ESX-1 substrates through a negative feedback loop that represses expression of the WhiB6 transcription factor in the absence of the secretory apparatus. Characterization of this feedback loop led to the identification of the novel transcription factor, EspM. We demonstrated that EspM represses whiB6 expression in the absence of the ESX-1 system by specifically binding to the whiB6 promoter. Based on structural predictions, we defined the function of several domains of EspM. We demonstrated that EspM binds DNA through its C-terminal domain. We identified that EspM multimerizes, and that both the N- and C-terminal domains can multimerize. We demonstrated that EspM’s multimeric capacity plays a role in repression. Finally, we identified several specific residues necessary for EspM multimerization and EspM function. Taken together, this work significantly pushes forward our understanding of mycobacterial pathogenesis. Our work not only elucidates mechanisms of regulation by the key mediator of mycobacterial virulence, but also provides drug targets for future therapeutics.