PROJECT SUMMARY This proposal describes the framework of an NIAID New Innovators DP2 award for Kiera Clayton, PhD. Dr. Clayton is currently a postdoctoral fellow under the supervision of Dr. Bruce Walker at the Ragon Institute of MGH, MIT and Harvard, whose current research focuses on T cell and NK cell-mediated killing of HIV-infected macrophages, and the implications for HIV reservoir persistence and macrophage-mediated inflammation. In addition to Dr. Clayton’s published work suggesting that macrophages are resistant to T cell-mediated killing, preliminary results included in this proposal suggest that HIV exploits macrophages to evade NK cell-mediated killing, by mechanisms distinct from those used to evade T cell responses. As macrophages also act as the primary cellular reservoir for other chronic pathogens, including Mycobacterium tuberculosis (Mtb), the question remains whether macrophages provide a common “Hide-Out” for immunoevasion. Thus, the overall goal of this proposal is to characterize the mechanisms employed by macrophages infected by different chronic pathogens to evade immune-mediated clearance and explore the possibility of targeting a common immunoevasion pathway to enhance T cell and NK cell killing of infected macrophages. This proposal focuses on three Research Areas. The first Research Area will build on preliminary data to define the mechanisms by which HIV-infected macrophages prevent NK cell-mediated killing. To quantify the resistance of macrophages infected with Mtb to T cell and NK cell-mediated elimination and characterize the accompanying immunoevasion mechanisms, the second Research Area will characterize the interactions between Mtb-infected macrophage and antigen-specific T cells and NK cells. Finally, as Dr. Clayton’s published work suggests that inefficient killing of macrophages is a result of resistance to apoptosis-inducing T cell-derived granzymes, the third Research Area will probe for the identities of granzyme inhibitors expressed by macrophages, and subsequently knockout candidate genes to enhance T and NK cell-mediated killing of infected macrophages. Together, these studies will provide insight into whether targeting natural granzyme inhibitors can provide a common method to enhance immune-mediated clearance of macrophages infected by different pathogens. The goals of this proposal are highly relevant to the mission of NIAID, and will enhance our understanding of distinct pathogen-specific immunoevasion mechanisms and those inherently characteristic of macrophages that allow for pathogen persistence despite strong immune responses. Ultimately, this work will aid the design and development of therapeutics to target HIV and Mtb infection in macrophages, and will potentiate future studies to assess how macrophage-mediated immunoevasion contributes to persistence of other pathogens, including human cytomegalovirus.