SUMMARY Recent findings identified that Programmed Cell Death (PD)-1+ CD8 T cells recognizing tumor or chronic pathogens have a division of labor: T cell factor (TCF)-1+ PD-1+ CD8 T cells function as memory-like resource cells and TCF-1neg PD-1+ CD8 T cells have effector-like function. TCF-1+ PD-1+ CD8 T cells self-renew and differentiate into effector-like and terminally differentiated/more exhausted TCF-1neg PD-1+ CD8 T cells. TCF-1+ memory-like cells have high expression CD28, and we have shown that during PD-1 targeted therapies CD28 costimulation is required for the reinvigoration of CD8 T cell responses. In addition, memory-like cells have high expression of ICOS, and a gene expression program with similarities to follicular helper CD4 T cells. Our preliminary data suggest that during established chronic lymphocytic choriomeningitis virus (LCMV) infection, continuous CD28 signaling is required for differentiation and self-renewal of TCF-1+ memory-like PD-1+ CD8 T cells. In contrast, ICOS signaling diminishes differentiation into effector-like cells. How memory-like cells choose between self-renewal and differentiation, and how to modulate differentiation into effector-like cells are critical questions. In addition to a unique set of costimulatory molecules, TCF-1+ memory-like cells also express a distinct set of chemokine/cytokine receptors. Based on these data and the knowledge gap in the field, we propose to define the role of costimulation in the maintenance and differentiation of PD-1+ CD8 T cells (Aim 1) and uncover the impact of cellular interactions (Aim 2). In Aim 1, we will determine how CD28 and ICOS costimulation affect TCF-1+ memory-like PD-1+ CD8 T cells and identify transcriptional regulators of T cell fate. In Aim 2, we will use an unbiased approach to identify cellular interactions of TCF-1+ memory-like PD-1+ CD8 T cells in vivo and probe the biological consequences of these interactions for T cell fate decisions. XCL-1, a chemoattractant for XCR1+ dendritic cells (DC1) is highly expressed by TCF-1+ memory-like cells and modulated by CD28 signaling. We will address the role of DC1 antigen presentation and XCL-1 production on localization, differentiation and self-renewal of TCF-1+ memory-like PD-1+ CD8 T cells. Understanding the determinants of self-renewal and differentiation of T cells chronically exposed to antigens would have broad implications for immunotherapies in many pathologies (chronic infections, cancer, autoimmunity, transplantation and allergy).