Transcriptional Programming in the Maintenance and Commitment of Nephron Progenitors

项目摘要 美国约有490万成年人患有肾脏疾病。目前的疗法不能提供治愈性的 治疗终末期肾病。再生肾学有望开发新的治疗方法 战略布局肾的功能单位是肾单位，它是从一个独特的祖细胞 胚胎发育期间的人口。肾单位祖细胞（NPC）自我更新和 在肾脏的过程中，需要分化以产生物种适当数量的肾单位。 发展转录调节因子Six 2和Wnt/β-catenin信号传导的转录介质 途径，Lef/Tcf因子，在平衡NPC的维持和承诺方面发挥着关键作用。在这方面，Six 2是 对NPC自我更新至关重要，并与Lef/Tcf因子相互作用。β-连环蛋白的积累起到了转换密码的作用 Lef/Tcf因子参与从阻遏物到激活物转录特征的分化靶点。的 在该提议中测试的总体假设是β-连环蛋白积累转换了Lef/Tcf调节性细胞因子。 从OFF到ON状态的签名。在AIM 1中，我将进行功能丧失实验来揭示其机制 β-catenin浓度依赖性Tcf/Lef因子参与导致转录变化的作用。 在AIM 2中，我将进行多蛋白质组学分析，以确定β-连环蛋白周围的多蛋白质复合物 与免疫沉淀和基于质谱的亲和蛋白质组学 Wnt/β-catenin信号与已鉴定的结合伴侣协同作用将促进肾单位样细胞的衍生 用于肾脏疾病的体外和最终再生疗法。

PROJECT ABSTRACT Approximately 4.9 million adults in the US suffer from kidney disease. Current therapies do not provide curative treatments for end state renal disease. Regenerative nephrology holds promise for developing novel therapeutic strategies. The functional unit of the kidney is the nephron, which is derived from a distinct progenitor cell population during embryonic development. A delicate balance of nephron progenitor cell (NPC) self-renewal and differentiation is required to generate a species-appropriate number of nephrons during the course of kidney development. The transcriptional regulator Six2, and transcriptional mediators of the Wnt/β-catenin signaling pathway, Lef/Tcf factors, play critical roles balancing the maintenance and commitment of NPCs. In this, Six2 is essential for NPC self-renewal and interacts with Lef/Tcf factors. β-catenin accumulation acts to switch the code of Lef/Tcf factor engagement at differentiation targets from a repressor to activator transcriptional signature. The overarching hypothesis tested in this proposal is that β-catenin accumulation switches the Lef/Tcf regulatory signature from an OFF to ON state. In AIM 1, I will perform loss of function experiments to unravel the mechanism of action of β-catenin concentration dependent Tcf/Lef factor engagement resulting in transcriptional changes. In AIM 2, I will perform multi-protein proteomic analysis to define multiprotein complexes surrounding β-catenin with immunoprecipitation and mass spectrometry-based affinity proteomics Mechanistic understanding of Wnt/β-catenin signaling in concert with identified binding partners will facilitate the derivation of nephron-like cell types in vitro and ultimately regenerative therapies for renal diseases.