Mycobacterium LacI-type Transcription Regulator Rv3575c Affects Host Innate Immunity by Regulating Bacterial <i>mce4</i> Operon-Mediated Cholesterol Transport

<i>Mycobacterium tuberculosis</i> has evolved a highly specialized system to snatch essential nutrients from its host, among which host-derived cholesterol has been established as one main carbon source for <i>M. tuberculosis</i> to survive within granulomas. The uptake, catabolism, and utilization of cholesterol are important for <i>M. tuberculosis</i> to sustain within the host largely via remodeling of the bacterial cell walls. However, the regulatory mechanism of cholesterol uptake and its impact on bacterium fate within infected hosts remain elusive. Here, we found that <i>M. tuberculosis</i> LacI-type transcription regulator Rv3575c negatively regulates its <i>mce4</i> family gene transcription. Overexpression of <i>Rv3575c</i> impaired the utilization of cholesterol as the sole carbon source by <i>Mycobacterium smegmatis</i>, activating the host's innate immune response and triggering cell pyroptosis. The <i>M. smegmatis</i> homologue of <i>Rv3575c MSMEG6044</i> knockout showed enhanced hydrophobicity and permeability of the cell wall and resistance to ethambutol, suppressed the host innate immune response to <i>M. smegmatis</i>, and promoted the survival of <i>M. smegmatis</i> in macrophages and infected mouse lungs, leading to reduced transcriptional levels of TNFα and IL-6. In summary, these data indicate a role of <i>Rv3575c</i> in the pathogenesis of mycobacteria and reveal the key function of <i>Rv3575c</i> in cholesterol transport in mycobacteria.