Common tissue residency associated genes. Table S3. key question in transplantation as T cells are detected long-term in allografts, but it is not known whether they are exhausted or are functional memory T cells. Using a mouse model of kidney transplantation, we showed that antigen-specific and polyclonal effector T cells differentiated in the graft into TRM and subsequently caused allograft rejection. TRM identity was established by surface phenotype, transcriptional profile, and Histone Acetyltransferase Inhibitor II inability to recirculate in parabiosis and re-transplantation experiments. Graft TRM proliferated locally, produced IFN upon re-stimulation, and their depletion attenuated rejection. Importantly, the vast DLL4 majority of antigen-specific and polyclonal TRM lacked phenotypic and transcriptional exhaustion markers. Single cell analysis of graft T cells early and late after transplantation identified a transcriptional program associated with transition to the tissue resident state that could serve as a platform for the discovery of therapeutic targets. Thus, recipient effector T cells differentiate into functional graft TRM that maintain rejection Histone Acetyltransferase Inhibitor II locally. Targeting these TRM could improve renal transplant outcomes. One Sentence Summary: Recipient effector T cells differentiate into functional Histone Acetyltransferase Inhibitor II tissue resident memory T cells, causing graft rejection after kidney transplantation. Introduction Recent studies have identified noncirculating, memory T cells in non-lymphoid tissues in mice and humans (1, 2). Aptly named tissue-resident memory T cells (TRM), TRM are phenotypically and transcriptionally distinct from circulating effector and Histone Acetyltransferase Inhibitor II central memory T cells and are numerically the largest T cell memory subset in the body (3C5). TRM form at sites of previous infection outside lymphoid tissues – particularly in barrier organs such as the skin, gut, lungs, and female reproductive tract – and remain there long-term, providing local protection against re-infection (6C11). They have also been observed in cancer (12C14) and in organs affected by chronic allergy or autoimmunity (15C17), where they contribute to tumor immunity and tissue pathology. Despite increasing appreciation of TRM in immunity, their role in organ transplantation is not well understood. Alloreactive effector and memory T cells generated in secondary lymphoid tissues migrate to transplanted organs and are activated and propagate locally to cause rejection (18, 19). In mice and humans, grafts that do Histone Acetyltransferase Inhibitor II not succumb to acute rejection acquire a persistent T cell infiltrate and eventually fail due to chronic rejection (20, 21). Whether TRM form in this setting and contribute to the rejection process is unclear. It has been argued instead that graft T cells become exhausted after repeated exposure to alloantigens (22, 23), and that exhaustion is possibly a mechanism by which grafts are protected from rejection (24C26). Recent human studies have described donor- and host-derived TRM-like cells in small intestine, lung, and kidney allograft samples based on phenotypic and transcriptional characteristics (27C29). These studies support the hypothesis that TRM form in organ transplants but do not establish TRM identity based on nonmigratory behavior and do not ascertain their function. Here, we investigated whether recipient TRM form and function in organ transplants by studying a mouse model in which kidney allografts undergo delayed rejection with chronic features (tubular atrophy, interstitial fibrosis and vasculopathy) (21, 30, 31), and monoclonal and polyclonal T cells.