Myocrdial biology has been transformed by recognition of the role played by cardiac progenitor cells (CPC) in repair and regeneration following pathological injury as well as their participation in normal cardiac homoestsis, remodeling, and aging. The initial period of this MERIT award focused upon two key regulatory proteins that our group identified as key players in CPC biology: Notch and nucleostemin. Since our initial descriptions of these signaling proteins in the myocardial context both have been studied and integrated into the fabric of cardiovascular literature. This MERIT award extension period will take our understanding of Notch and nucleostemin to a higher level by defining connections between these molecules, canonical pathways in myocardial regulatory signaling, and involvement in the biology of CPC. The significance of this proposal is novel information that can be applied directly toward the manipulation of CPC to enhance myocardial repair and regeneration as well as understanding pathological mechanisms resulting from CPC dysfunction. Innovative aspects of this proposal include the use of unique reagents to manipulate Notch and nucleostemin activity through use of inducible constructs, silencing vectors, and observation of activity through genetically engineered reporters both in cultured cells and transgenic mice. Specific aims are 1) Notch signaling potentiates cardiac progenitor differentiation into cardiac lineages in vitro and in vivo, 2) Notch signaling facilitates repair and regeneration of damaged mouse myocardium in vivo, S) Pim-1 mediated c-Myc stabilization induces nucleostemin that enhances regenerative potential and survival of cardiac progenitor cells, 4) nucleostemin regulates stem and progenitor cell pluripotency, and 5) elevated nucleostemin expression triggers hypertrophic signaling and enhances regenerative potential in the heart. The impact of the proposal will be advancing the use, manipulation, and control of CPC biology to facilitate myocardial reparative processes as well as explaining the underlying biology of myocardial responses to injury, adaptive remodeling, and aging. As cardiovascular stem cell biology enters the clinical arena it is essential to acquire mechanistic understanding to achieve optimal results.