Project Summary/Abstract Proper nephron endowment and segmentation are essential for building healthy kidneys. Nephron number is highly variable among humans. Low nephron number is associated with increased risk of hypertension and end-stage renal disease. A better understanding of the mechanisms of nephrogenesis will help us devise a way to increase nephron endowment. Along the proximal-distal axis, the renal corpuscle is connected to the collecting duct via the proximal tubule, loop of Henle, and distal tubule. Each nephron segment carries out distinct physiological functions. Understanding how multipotent nephron progenitors make their cell fate decisions to develop into different nephron segments is essential not only for building a functional nephron in vitro but also for promoting regeneration after kidney injury. In this proposed study, we will investigate how retinoic acid signaling regulates nephron endowment and segmentation. We found that, in mice, inhibition of retinoic acid signaling in mesenchymal nephron progenitor cells caused significantly lower nephron endowment. We also found that inhibition of retinoic acid signaling in the newly formed developing nephron interfered with the formation and maturation of proximal tubule cells. Our preliminary data suggest that retinoic acid signaling plays important roles in early and later stages of nephrogenesis, contributing to proper nephron endowment and segmentation. In Aim 1, by manipulating retinoic acid signaling genetically in vivo and pharmacologically in vitro, we will test if retinoic acid signaling is required for mesenchymal-to-epithelial transition of nephron progenitors by coordinating with Wnt/-catenin signaling. To understand how retinoic acid signaling contributes to nephron endowment, we will determine the retinoic acid gene regulatory network in nephron progenitors. In Aim 2, by performing genetic gain-of-function and loss-of-function studies of retinoic acid signaling in the newly formed developing nephron, we will test if retinoic acid signaling is necessary or sufficient for the formation and maturation of proximal tubules. We will also determine the retinoic acid gene regulatory network in the developing nephrons to understand how retinoic acid signaling regulates nephron segmentation. Our proposed studies will fill a longstanding gap of knowledge in the molecular mechanisms underlying the determination of nephron endowment as well as the formation or maturation of proximal tubules in mammals. Our long-term goals are to devise a way to increase nephron endowment during development and to generate nephron tubule cells for potential cell replacement therapy.