Mycobacterium tuberculosis is protected from NADPH oxidase and LC3-associated phagocytosis by the LCP protein CpsA.

Mycobacterium tuberculosis' success as a pathogen comes from its ability to evade degradation by macrophages. Normally macrophages clear microorganisms that activate pathogen-recognition receptors (PRRs) through a lysosomal-trafficking pathway called "LC3-associated phagocytosis" (LAP). Although Mtuberculosis activates numerous PRRs, for reasons that are poorly understood LAP does not substantially contribute to Mtuberculosis control. LAP depends upon reactive oxygen species (ROS) generated by NADPH oxidase, but Mtuberculosis fails to generate a robust oxidative response. Here, we show that CpsA, a LytR-CpsA-Psr (LCP) domain-containing protein, is required for Mtuberculosis to evade killing by NADPH oxidase and LAP. Unlike phagosomes containing wild-type bacilli, phagosomes containing the ΔcpsA mutant recruited NADPH oxidase, produced ROS, associated with LC3, and matured into antibacterial lysosomes. Moreover, CpsA was sufficient to impair NADPH oxidase recruitment to fungal particles that are normally cleared by LAP. Intracellular survival of the ΔcpsA mutant was largely restored in macrophages missing LAP components (Nox2, Rubicon, Beclin, Atg5, Atg7, or Atg16L1) but not in macrophages defective in a related, canonical autophagy pathway (Atg14, Ulk1, or cGAS). The ΔcpsA mutant was highly impaired in vivo, and its growth was partially restored in mice deficient in NADPH oxidase, Atg5, or Atg7, demonstrating that CpsA makes a significant contribution to the resistance of Mtuberculosis to NADPH oxidase and LC3 trafficking in vivo. Overall, our findings reveal an essential role of CpsA in innate immune evasion and suggest that LCP proteins have functions beyond their previously known role in cell-wall metabolism.

Mycobacterium tuberculosis EsxH inhibits ESCRT-dependent CD4+ T-cell activation.

Mycobacterium tuberculosis (Mtb) establishes a persistent infection, despite inducing antigen-specific T-cell responses. Although T cells arrive at the site of infection, they do not provide sterilizing immunity. The molecular basis of how Mtb impairs T-cell function is not clear. Mtb has been reported to block major histocompatibility complex class II (MHC-II) antigen presentation; however, no bacterial effector or host-cell target mediating this effect has been identified. We recently found that Mtb EsxH, which is secreted by the Esx-3 type VII secretion system, directly inhibits the endosomal sorting complex required for transport (ESCRT) machinery. Here, we showed that ESCRT is required for optimal antigen processing; correspondingly, overexpression and loss-of-function studies demonstrated that EsxH inhibited the ability of macrophages and dendritic cells to activate Mtb antigen-specific CD4+ T cells. Compared with the wild-type strain, the esxH-deficient strain induced fivefold more antigen-specific CD4+ T-cell proliferation in the mediastinal lymph nodes of mice. We also found that EsxH undermined the ability of effector CD4+ T cells to recognize infected macrophages and clear Mtb. These results provide a molecular explanation for how Mtb impairs the adaptive immune response.

Ubiquilin 1 Promotes IFN-γ-Induced Xenophagy of Mycobacterium tuberculosis.

The success of Mycobacterium tuberculosis (Mtb) as a pathogen rests upon its ability to grow intracellularly in macrophages. Interferon-gamma (IFN-γ) is critical in host defense against Mtb and stimulates macrophage clearance of Mtb through an autophagy pathway. Here we show that the host protein ubiquilin 1 (UBQLN1) promotes IFN-γ-mediated autophagic clearance of Mtb. Ubiquilin family members have previously been shown to recognize proteins that aggregate in neurodegenerative disorders. We find that UBQLN1 can interact with Mtb surface proteins and associates with the bacilli in vitro. In IFN-γ activated macrophages, UBQLN1 co-localizes with Mtb and promotes the anti-mycobacterial activity of IFN-γ. The association of UBQLN1 with Mtb depends upon the secreted bacterial protein, EsxA, which is involved in permeabilizing host phagosomes. In autophagy-deficient macrophages, UBQLN1 accumulates around Mtb, consistent with the idea that it marks bacilli that traffic through the autophagy pathway. Moreover, UBQLN1 promotes ubiquitin, p62, and LC3 accumulation around Mtb, acting independently of the E3 ligase parkin. In summary, we propose a model in which UBQLN1 recognizes Mtb and in turn recruits the autophagy machinery thereby promoting intracellular control of Mtb. Thus, polymorphisms in ubiquilins, which are known to influence susceptibility to neurodegenerative illnesses, might also play a role in host defense against Mtb.