UFMylation suppresses Type IFN signaling during Mycobacterium tuberculosis infection of human macrophages

Type I Interferons (IFN-I) promote host defense against a wide range of viral infections, but inhibit control of several bacterial pathogens, including Mycobacterium tuberculosis (Mtb). Given the significance of IFN-I signaling in determining Mtb infection outcomes, we sought to uncover new molecular mechanisms that regulate IFN-I during mycobacterial infection, specifically focusing on how IFN-I signaling is regulated at the earliest stages of mycobacterial infection as modeled by macrophage infection. To comprehensively identify these host genes, we performed a reporter-based, genome- wide CRISPR-interference screen in human macrophages infected with Mycobacterium marinum , a close relative of Mtb. Our screen detected 2035 significantly enriched genes (p<0.05), which included many known regulators of IFN-I, but also many unexpected genes as potentially novel regulators of IFN-I signaling. One of these unexpected genes was UFL1, an E3-like ligase that catalyzes conjugation of the ubiquitin-like protein, UFM1, in a process termed UFMylation. UFL1-deficiency during Mtb macrophage infection resulted in increased expression of IFN-β, interferon-stimulated genes, and other pro-inflammatory genes, such as TNF and IL-6, at both the transcript and protein level. Depletion of other UFMylation components phenocopied UFL1-deficiency, suggesting that UFMylation activity is required for IFN-I repression. Full transcriptional profiling revealed a broad increase in the inflammatory response of UFL1-deficient cells during Mtb infection, including both protective inflammatory cytokines and potentially detrimental interferon-stimulated genes. Our results suggest a role for UFMylation in suppressing IFN-I signaling and inflammatory responses during the earliest stages of Mtb infection.