During the interaction of T cells with antigen presenting cells (APCs), receptors and intracellular proteins translocate into the contact region known as the Immunological Synapse (IS). In the first Aim we will examine whether CD8 behaves as a coreceptor within the immunological synapse, for the activation of peripheral T cells. In the other component of this proposal we shall follow up on our recent observation that cross-linking of CD8, but not CD4, with antibodies or MHC class I molecules, results in rapid apoptosis of CD4+CD8+ (DP) thymocytes. We have identified the thymocytes that are susceptible to this CD8 mediated apoptosis as an early DP population that expresses low levels of CD2 and CD5 and is CD69-. Treatment with the phorbol ester, PMA, or antibodies to CD3, prevents this induction of apoptosis. We hypothesize that ligation of CD8 by MHC class I molecules in the absence of TCR engagement leads to the death of a subpopulation of early DP thymocytes. Furthermore, our system may provide a model for thymocytes that fail positive selection and are said to undergo "death by neglect". In this proposal we shall further characterize the induction of apoptosis in DP thymocytes by CD8 antibodies and MHC class I molecules according to 3 specific aims: (i) to determine the biochemical events leading to the CD8 mediated death of DP thymocytes;(ii) to determine the mechanism by which treatment with PMA, or engagement of the TCR, prevents CD8 mediated death in DP thymocytes;(iii) to determine whether CD8 mediated death functions under physiological conditions.