Recent advances in the understanding of T-cell activation have led to new therapeutic approaches in the treatment of immunological disorders. One attractive target of intervention has been the blockade of T cell-mediated co-stimulatory pathways, which result in more selective effects on only those T-cells that have encountered specific antigen. In fact, in some instances, CD28/B7 co-stimulatory pathway antagonists can induce antigen-specific tolerance that prevents the progression of autoimmune diseases and organ graft rejection. However, many other cell surface molecules exist on naive T cells that may play an important role in initiating immunity from the quiescent state. Moreover, it is clear that quiescent T cells can also function as regulatory T cells to promote immune tolerance by an as yet unknown mechanism suggesting that some of the cell surface proteins may provide functional activities beyond just determining whether cells effectively go into cell cycle. Notch, Serrate, and Delta are type 1 integral membrane proteins involved in cell-cell signaling crucial to normal cellular differentiation and in organogenesis. In the immune system, Notch signaling is critical for normal T cell development. Recently, a role for Notch signaling in mature T cells has been proposed based on the observation that engagement of Notch by Serrate could induce a profound state of unresponsiveness in peripheral T cells in vivo. Most interesting was the observation that these "unresponsive" T cells possessed regulatory abilities that on several levels appear similar to the CD25+ regulatory cells that are able to regulate autoimmunity. Our preliminary studies suggest that Notch-1 is an important regulator of T cell signaling in resting T cells and may be involved in tolerance. We hypothesize that Notch-1 plays a critical role in the intrinsic regulation of T cell responses by influencing the primary signaling pathways critical for activating naive T cells. This results in T cell inactivation or altered differentiation. Moreover, we hypothesize that manipulation of immune responses through Notch-1 can be used to alter autoimmunity and transplantation tolerance. In this application we propose to explore the mechanisms of Notch signaling in T cells and its application as follows: Specific Aim #1. To Identify the biochemical, cellular and genetic effects of Notch signaling in T cells; and Specific Aim #2. In vivo application of Notch signaling for the establishment of peripheral T cell tolerance.