STING is an endoplasmic reticulum (ER)-associated transmembrane protein that turns on and quickly turns off downstream signaling as it translocates from the ER to vesicles. How STING signaling is attenuated during trafficking remains poorly understood. Here, we show that trafficking-mediated STING degradation requires ER exit and function of vacuolar ATPase complex. Late-stage STING vesicles are sorted to Rab7-positive endolysosomes for degradation. Based on analysis of existing structures, we also identified the helix amino acid 281 (aa281)-297 as a motif required for trafficking-mediated STING degradation. Immuno-electron microscopy (EM) reveals the size and clustering of STING vesicles and topology of STING on the vesicle. Importantly, blockade of trafficking-mediated STING degradation using bafilomycin A1 specifically enhanced cyclic guanosine monophosphate (GMP)-AMP (cGAMP)-mediated immune response and anti-tumor effect in mice. Together, our findings provide biochemical and imaging evidence for STING degradation by the lysosome and pinpoint trafficking-mediated STING degradation as a previously unanticipated therapeutic target for enhancing STING signaling in cancer therapy.

Mycobacterium tuberculosis is a hard-to-eradicate intracellular pathogen that infects one-third of the global population. It can live within macrophages owning to its ability to arrest phagolysosome biogenesis. Autophagy has recently been identified as an effective way to control the intracellular mycobacteria by enhancing phagosome maturation. In the present study, we demonstrate a novel role of miR-155 in regulating the autophagy-mediated anti-mycobacterial response. Both in vivo and in vitro studies showed that miR-155 expression was significantly enhanced after mycobacterial infection. Forced expression of miR-155 accelerated the autophagic response in macrophages, thus promoting the maturation of mycobacterial phagosomes and decreasing the survival rate of intracellular mycobacteria, while transfection with miR-155 inhibitor increased mycobacterial survival. However, macrophage-mediated mycobacterial phagocytosis was not affected after miR-155 overexpression or inhibition. Furthermore, blocking autophagy with specific inhibitor 3-methyladenine or silencing of autophagy related gene 7 (Atg7) reduced the ability of miR-155 to promote autophagy and mycobacterial elimination. More importantly, our study demonstrated that miR-155 bound to the 3′-untranslated region of Ras homologue enriched in brain (Rheb), a negative regulator of autophagy, accelerated the process of autophagy and sequential killing of intracellular mycobacteria by suppressing Rheb expression. Our results reveal a novel role of miR-155 in regulating autophagy-mediated mycobacterial elimination by targeting Rheb, and provide potential targets for clinical treatment. microRNA-155 (miR-155) plays an essential role in regulating the host immune response by post-transcriptionally repressing the expression of target genes. However, little is known regarding its activity in modulating autophagy, an important host defense mechanism against intracellular bacterial infection. Mycobacterium tuberculosis is a hard-to-eradicate intracellular pathogen that infects approximately one-third of the global population, and causes 1.5 million deaths annually. The present study explores a novel role of miR-155 in the host response against mycobacterial infection. Our data demonstrates that mycobacterial infection triggers the expression of miR-155, and the induction of miR-155 in turn activates autophagy by targeting Rheb, a negative regulator of autophagy. miR-155-promoted autophagy accelerates the maturation of the mycobacterial phagosome, thus decreasing the survival of intracellular mycobacteria in macrophages. These findings contribute to a better understanding of the host defense mechanisms against mycobacterial infection, providing useful information for development of potential therapeutic interventions against tuberculosis.

Mutations in the NGLY1 (N-glycanase 1) gene, encoding an evolutionarily conserved deglycosylation enzyme, are associated with a rare congenital disorder leading to global developmental delay and neurological abnormalities. The molecular mechanism of the NGLY1 disease and its function in tissue and immune homeostasis remain unknown. Here, we find that NGLY1-deficient human and mouse cells chronically activate cytosolic nucleic acid–sensing pathways, leading to elevated interferon gene signature. We also find that cellular clearance of damaged mitochondria by mitophagy is impaired in the absence of NGLY1, resulting in severely fragmented mitochondria and activation of cGAS–STING as well as MDA5–MAVS pathways. Furthermore, we show that NGLY1 regulates mitochondrial homeostasis through transcriptional factor NRF1. Remarkably, pharmacological activation of a homologous but nonglycosylated transcriptional factor NRF2 restores mitochondrial homeostasis and suppresses immune gene activation in NGLY1-deficient cells. Together, our findings reveal novel functions of the NGLY1–NRF1 pathway in mitochondrial homeostasis and inflammation and uncover an unexpected therapeutic strategy using pharmacological activators of NRF2 for treating mitochondrial and immune dysregulation.