PROJECT SUMMARY During chronic infections such as those caused by HIV-1, hepatitis C virus (HCV), and lymphocytic choriomeningitis virus (LCMV), as well as during tumor outgrowth, antigen-specific CD8 T cells often progressively lose function in a step-wise manner. Understanding the factors that impact CD8 T cell responses during chronic viral infections is necessary for the design of effective immunotherapeutic strategies. Recent reports demonstrate that a discrete subset of exhausted CD8 T cells exists that displays stem cell-like properties, including the ability to self-renew and further differentiate in vivo. Importantly, this subset of cells is exquisitely responsive to anti-PD-L1 immunotherapy during chronic LCMV clone 13 infection. Hence, determining the mechanisms that promote the maintenance and survival of stem-like CD8 T cells during chronic viral infection is of central importance. We present preliminary findings that show a distinct fraction of the virus-specific stem-like CD8 T cells express cell surface N-glycans with α-2,6 linked sialic acids, a posttranslational modification mediated by the glycosyltransferase ST6Gal-I. Importantly, published studies, as well as our own data, establish that this enzyme is associated with pluripotency and stemness. Together, these data lead us to hypothesize that ST6Gal-I sialylation positively regulates CD8 T cell stemness and responsiveness to immunotherapy during chronic viral infection. Our objectives of this exploratory R21 application are to: Aim 1. Determine if ST6Gal-I sialylation identifies stem-like CD8 T cells during chronic viral infection. Aim 2: Determine how modulation of ST6Gal-I sialylation impacts CD8 T cell stemness. Collectively, these studies are designed to provide new information regarding the impact of cell surface sialylation of CD8 T cell responses, including CD8 T cell stemness and exhaustion.