Project Summary/Abstract (Project 2, Cox) Tuberculosis (TB), caused by infection with Mycobacterium tuberculosis, remains a major cause of human morbidity and mortality, particularly in the developing world. Chronic M. tuberculosis infection requires long- term interactions between the bacterium and host immune system, and tissue macrophages play key roles in the outcome of infection. The goal of this project is to identify the interactions between M. tuberculosis and macrophages at the molecular level, and to understand how they influence bacterial killing. Our previous work has identified a macrophage pathway activated by M. tuberculosis infection that induces both antibacterial responses of macrophages and, paradoxically, anti-viral responses that inhibit immunity. In this proposal, we seek to dissect these two antagonistic responses activated by this pathway, which requires the host factor STING, with the long-term goal of developing host-directed therapeutics that push the balance in favor of TB immunity. In Aim 1 we investigate the molecular mechanisms by which the STING pathway targets M. tuberculosis to autophagy, a powerful anti-infection pathway of the host. In Aim 2, we explore the role of Galectin-9 in host defense against M. tuberculosis infection, as this carbohydrate-binding protein is a prime candidate for collaborating with STING to promote autophagy. In Aim 3, we seek to understand the mechanism by which STING-activated interferon responses function to promote M. tuberculosis infection. Importantly, both the autophagy targeting and anti-viral signaling pathways are controlled by important kinases, indicating that post-translational modification of host proteins play important roles in mediating these effects. Thus, our proposed use of cutting-edge proteomics in Aim 4 to identify the proteins modified by these enzymes during infection represents a powerful way to uncover mechanisms by which macrophages control infection. Importantly, our work is highly synergistic with the other projects and cores that make up this Program Project, and the comparisons of host responses elicited by M. tuberculosis with those induced by L. monocytogenes and L. pneumophila will allow us to identify pathogen-specific responses as well as common responses induced by diverse intracellular bacterial pathogens. These studies will not only provide deeper mechanistic understanding of how autophagy and anti-viral signaling control infection, but may also uncover novel host pathways that function to eliminate intracellular bacterial pathogens.