The earliest described progenitors in the adult mouse thymus are termed early thymic progenitors (ETPs). They derive from rare circulating hematopoietic progenitors, whose identity was previously unknown. Our work in the previous funding period established that a population of progenitors termed early lymphoid progenitors (ELPs) circulates, and physiologically settles the thymus. In our current proposal, we wish to understand how rare ELPs settling the thymus give rise to ETPs and other downstream populations from which mature, immunocompetent T cells are ultimately derived. We hypothesize that tremendous proliferation occurs right after thymic settling, by which rare ELPs that settle the thymus generate large numbers of early thymic progenitors (ETPs). Indeed, we think that more proliferation may occur immediately after thymic settling than at any subsequent stage of T lymphopoiesis. One goal of our current proposal is to quantify this proliferation, and to determine how it is regulated. As ELPs settling the thymus possess lineage potentials for multiple cell types, another goal is to address the question of how alternative lineage fates and in particular the myeloid fate are constrained after thymic settling. Notch is a cell surface receptor involved in multiple cell fate decisions, and we wish to probe the role of Notch signaling in these critical early events. Further, we wish to determine the molecular mechanisms by which Notch co-ordinates T lineage commitment and lineage progression. Together, these studies will allow us to piece together a lineage of cells that extends from hematopoietic stem cells in the bone marrow to ETPs within the thymus, and to understand the signals that guide cells down the T cell developmental pathway. Our specific Aims are: Specific Aim 1: To measure proliferation between thymic settling and the generation of ETPs. Specific Aim 2: To determine the function of Notch signaling in ETPs. Specific Aim 3: To determine the mechanism by which Notch signals result in T lineage progression and commitment. PUBLIC HEALTH RELEVANCE: We propose to characterize proliferative and differentiative events at the earliest stages of intrathymic T cell development. Notch plays a critical role at this earliest stage, and we propose to elucidate the function of Notch signaling in early T cell development, and to determine the mechanisms by which it co-ordinates early T lineage progression and commitment.