Summary/Abstract Transplantation is a curative strategy for end stage organ failure. In order to avoid immune-mediated rejection of the grafted tissue, recipients receive life-long immunosuppression. Calcineurin and mTOR inhibitors for immunosuppression are successful in preventing graft rejection but are associated with significant adverse off- target toxicities. In order to better preserve the health of both the graft and the transplant recipient, alternative immunosuppressive therapies have been sought. Belatacept, a CTLA-4Ig fusion protein, suppresses the immune system by binding to CD80 and CD86 on antigen presenting cells (APCs) to block CD28 (costimulatory) and CTLA-4 (coinhibitory) signaling on T cells. By blocking interactions that are specific for immune cells, the use of CTLA-4Ig mitigates off-target toxicities associated with calcineurin inhibitors and increases the longevity of the graft and health of the patient. However, CTLA-4Ig is associated with increased risk of acute rejection episodes compared to calcineurin inhibitors, which has prohibited its widespread clinical adoption despite the improved long-term outcomes. Regulatory T cells are essential to maintaining immune homeostasis and promoting tolerance in transplant, but the blockade of signaling through CD28 and CTLA-4 is detrimental to their survival and function. Therefore, CTLA-4Ig therapy inhibits Tregs from controlling CD8+ T cells as they execute acute rejection. Identifying costimulatory and coinhibitory pathways that can be targeted to enhance the suppressive capacity of Tregs in the setting of CTLA-4Ig therapy is essential to improving targeted immunosuppressive therapies. TIGIT is a T-cell inhibitory receptor with Ig and ITIM domains expressed on Tregs that can be targeted with antibodies to fine-tune Treg function. Agonistic anti-TIGIT antibodies have been shown to induce apoptosis of costimulation blockade-resistant memory T cells in a Treg- dependent manner. The experiments outlined in this proposal study the mechanisms by which TIGIT signaling on Tregs enhances Treg function in the face of CTLA-4Ig therapy to reduce CD8 + T cell responses to allograft challenge. In Aim 1 we will determine if cell-autonomous TIGIT signaling increases the abundance of Tregs within grafted tissue and increases the secretion of suppressive molecules to enhance Treg function. In Aim 2 we will determine the impact of Treg-specific TIGIT signaling on effector CD8+ T cell responses to allograft. These studies will be performed using novel Treg-specific TIGIT knockout animals (Foxp3-Cre x TIGITfl/fl). Together, these data will illuminate the therapeutic potential of TIGIT agonism in combination with costimulation blockade therapeutics to mitigate acute allograft rejection.