Summary Activation of the DNA-dependent cytosolic surveillance pathway in response to Mycobacterium tuberculosis infection stimulates ubiquitin-dependent autophagy and inflammatory cytokine production, and plays an important role in host defense against M. tuberculosis. However, the identity of the host sensor for M. tuberculosis DNA is unknown. Here we show that M. tuberculosis activated cyclic guanosine monophosphate–adenosine monophosphate (cGAMP) synthase (cGAS) in macrophages to produce cGAMP, a second messenger that activates the adaptor protein stimulator of interferon genes (STING) to induce type I interferons and other cytokines. cGAS localized with M. tuberculosis in mouse and human cells and in human tuberculosis lesions. Knockdown or knockout of cGAS in human or mouse macrophages blocked cytokine production and induction of autophagy. Mice deficient in cGAS were more susceptible to lethality caused by infection with M. tuberculosis. These results demonstrate that cGAS is a vital innate immune sensor of M. tuberculosis infection.

SUMMARY During antibacterial autophagy, ubiquitination of intracellular bacteria recruits proteins that mediate bacterial delivery to the lysosome for degradation. Smurf1 is an E3 ubiquitin-ligase whose role in selective bacterial autophagy is unknown. We show that Smurf1 facilitates selective autophagy of the human pathogen Mycobacterium tuberculosis (Mtb). Smurf1−/− macrophages are defective in recruiting polyubiquitin, the proteasome, the ubiquitin-binding autophagy adaptor NBR1, the autophagy protein LC3, and the lysosomal marker LAMP1 to Mtb-associated structures, and are more permissive for Mtb growth. This function of Smurf1 requires both its ubiquitin-ligase and C2 phospholipid-binding domains, and involves K48-rather than K63-linked ubiquitination. Chronically infected Smurf1−/− mice have increased bacterial load, increased lung inflammation, and accelerated mortality. SMURF1 controls Mtb replication in human macrophages and associates with bacteria in lungs of patients with pulmonary tuberculosis. Thus, Smurf1 is required for selective autophagy of Mtb and host defense against tuberculosis infection.

Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis, is responsible for 1.5 million deaths annually. We previously showed that Mtb infection in mice induces expression of the carbon monoxide (CO) producing enzyme heme oxygenase (HO1) and that CO is sensed by Mtb to initiate a dormancy program. Further, mice deficient in HO1 succumb to Mtb infection more readily than wild type mice. While mouse macrophages control intracellular Mtb infection through several mechanisms such as nitric oxide synthase, the respiratory burst, acidification and autophagy, how human macrophages control Mtb infection remains less well understood. Here we show that Mtb induces and colocalizes with HO1 in both mouse and human tuberculosis lesions in vivo, and that Mtb induces and colocalizes with HO1 during primary human macrophage infection in vitro. Surprisingly, we find that chemical inhibition of HO1 both reduces inflammatory cytokine production by human macrophages and restricts intracellular growth of mycobacteria. Thus, induction of HO1 by Mtb infection may be a mycobacterial virulence mechanism to enhance inflammation and bacterial growth.

A reader brought errors in Figure 4 to the authors' attention. In Figures 4A and 4D, several immunofluorescence panels were inadvertently reversed during their final arrangement. Panels that should have indicated the merge image were incorrectly placed where the single channel GFP image should be, and vice versa. Figures 4A and 4D have now been corrected online. The original results and conclusions are unaffected by these corrections. The authors accept responsibility for not detecting these errors prior to publication and regret any inconvenience this has caused.

Summary The prevailing paradigm is that tuberculosis infection is initiated when patrolling alveolar macrophages and dendritic cells within the terminal alveolus ingest inhaled M. tuberculosis (Mtb). However, definitive data for this model are lacking. Among the epithelial cells of the upper airway a specialized epithelial cell known as a microfold cell (M-cell) overlies various components of mucosa associated lymphatic tissue. Here we show using multiple mouse models that Mtb invades via M-cells to initiate infection. Intranasal Mtb infection in mice lacking M-cells either genetically or by antibody depletion resulted in reduced invasion and dissemination to draining lymph nodes. M-cell depleted mice infected via aerosol also had delayed dissemination to lymph nodes and reduced mortality. Translocation of Mtb across two M-cell transwell models was rapid and transcellular. Thus, M-cell translocation is a vital entry mechanism that contributes to the pathogenesis of Mtb.