Genetic analysis conducted during the previous funding period defined two stages where Notch signals play an important role in kidney biology. The first essential function of Notch is during nephron segmentation. Notch2 signals block distal identities and promote proximal tubule (PT) and podocyte (POD) identities either cell autonomously or inducing them in neighboring cells. This activity is required only transiently, becoming dispensable after S-shaped body (SB) formation. A second important function of Notch signals is to act as gatekeepers of renal tubular epithelial homeostasis during adulthood; in their absence, cysts and renal dysfunction develop spontaneously in a substantial fraction of outbred animals with some cysts containing cells that continue to divide and develop papillary adenomas, thought to be the precursor for papillary renal cell carcinoma (PRCC). Injury increases the frequency of cyst and papillary adenoma formation. While these findings explain why reduced Notch2 signaling in humans causes Alagille syndromes (AGS), and suggest that elevating Notch1 activity in such patients should improve health, they do not establish if/how Notch signals direct the acquisition of PT and POD fates. We will take advantage of cell lineage tracing tools prepared during the previous funding period to address the issue of whether the cellular organization in the SB, at the time of Notch2 activation, underlies the organization of nephron sub-structures in the adult nephron and map the potential of de-differentiated renal PT recovering from injury in the adult. These fate mapping studies will allow us to ask if SB cells experiencing Notch activity are fated to become PT, POD, neither, or both, and if de-differentiated adult renal epithelial cells are fate-restricted or oligopotential. The results will direct our subsequent search for genes capable of enhancing POD and PT regeneration in injured adults. In Aim 2, we will ask how these cells acquire their fates. In Aim 3, we will determine the mechanistic contribution of Notch loss to cyst and adenoma formation in adults. In Aim 4, we will develop strategies to transiently augment Notch1 activity in AGS and PRCC with the translational goal of developing clinical interventions aimed at compensating for Notch signal reduction reduction in AGS.