Role of afadin signaling in nephron tubulogenesis

项目摘要/摘要极化上皮小管的形成对肾脏的结构和功能至关重要。尽管它很重要，但在理解小管形成的最早步骤方面存在根本性的空白，即上皮如何建立顶基极性并产生中心管腔。这种差距的持续存在代表了一个重要的问题，因为直到它被填补，对肾脏发育异常的理解在很大程度上仍然是不可理解的。本应用程序的目的是阐明如何建立顶基极性和管腔形成的开始，重点是Afadin信号在肾单位形成中的作用。Afadin是黏附受体连接蛋白家族的一种衔接蛋白。我们的初步数据表明，在发育中的小鼠肾单位中，Afadin对于建立根尖表面和启动管腔形成至关重要。为了进一步阐明启动极性和管腔形成的细胞和分子机制，提出的研究有三个具体目的。第一个目标是在体内非常详细地描述管腔形成的分子步骤。这些研究将通过极性、运输和连接复合物的亚细胞定位，以及通过小鼠管腔形成的实时成像，生成管腔形成的分子时间线。一旦确定了特征，其他的研究将开始使用小鼠模型将蛋白质分配到该途径的不同阶段。第二个目的是利用已建立的三维细胞培养模型，确定新的根尖极性和管腔形成的分子机制。具体来说，提出的实验将阐明连接蛋白和Afadin启动管腔形成的详细信号通路。第三个目的是研究Afadin在缺血性损伤后小管再生中的作用，强调其与发育过程的相似性。总之，这些研究将描述极性和管腔起始的基本步骤，并为理解肾小管的发育和获得性疾病提供一个改进的框架。

DESCRIPTION (provided by applicant): Role of Afadin Signaling in Nephron Tubulogenesis Project Summary/Abstract Formation of polarized epithelial tubules is central to the structure and function of the kidney. Despite its importance, there is a fundamental gap in understanding the earliest steps in tubulogenesis, namely how epithelia establish apical-basal polarity and generate a central lumen. Continued existence of this gap represents an important problem because, until it is filled, an understanding of developmental abnormalities of the kidney will remain largely incomprehensible. The objective of this application is to elucidate how apical- basal polarity is established and lumen formation initiated, focusing on the role of Afadin signaling in nephron formation. Afadin is an adaptor protein to the Nectin family of adhesion receptors. Our preliminary data identify Afadin as critical for establishing an apical surface and initiating lumen formation in developing mouse nephrons. To further elucidate the cellular and molecular mechanisms that initiate polarity and lumen formation, the proposed studies have three specific aims. The first aim is to characterize molecular steps of lumen formation in vivo and in great detail. These studies will generate a molecular timeline of lumen initiation through subcellular localization of polarity, trafficking, and junctional complexes, and through live imaging of lumen formation in mice. Once characterized, additional studies will begin to assign proteins to different stages in the pathway using mouse models. The second aim is to identify novel molecular mechanisms of apical polarity and lumen formation using an established 3D cell culture model. Specifically the proposed experiments will elucidate a detailed signaling pathway by which Nectins and Afadin initiate lumen formation. The third aim will examine the role of Afadin in tubule regeneration after ischemic injury, emphasizing similarities to the developmental process. Together these studies will delineate fundamental steps in polarity and lumen initiation and provide an improved framework for understanding developmental and acquired disorders of renal tubules.