CELL MIGRATION, CHEMOATTRACTION AND THE RET/GDNF PATHWAY

哺乳动物肾脏的发育需要诱导性 输尿管芽上皮和输尿管芽上皮两种前体组织的相互作用 后肾间充质。从输尿管芽发出的信号 诱导间充质细胞增殖和分化，从而生成 肾单位内的大多数上皮细胞类型。反过来， 间充质来源的信号诱导输尿管芽上皮 增殖并经历分枝形态发生以产生 收集风管系统。尽管这些信号和它们的受体 仍然难以捉摸，它们的作用机制是 了解肾细胞生长和发育的分子基础 差异化。受体酪氨酸激酶RET在 输尿管芽尖端，调节增殖和分支 形态发生。PI实验室最近的工作表明， 胶质细胞源性生长因子与RET和RET相互作用 刺激RET的磷酸化。GDNF和RET是公共的 促进输尿管芽增殖的信号通路和 分支。这一途径可能不仅对伊利肾脏至关重要 发育和生长，但也可能在异常增殖中发挥作用 输尿管芽细胞，如在囊性疾病中 收集风管。 我们建立了一种RET活性的生物测定方法 MDCK肾上皮细胞。这些细胞表现出更强的散射， 细胞运动性，以及RET激活后的形态变化。 现已证实GDNF/RET途径促进迁移 输尿管芽上皮朝向后肾间充质，一种有效的 GDNF的来源。我们的初步数据显示，RET表达 上皮细胞可以向GDNF的局部来源迁移。因此， 输尿管芽长出是由于对靶点的化学吸引。 派生的指导线索。如何接收和解释GDNF信号 通过上皮细胞是这一提议的主要关注点。这个 特异性AIMS将利用酵母双杂交系统鉴定 与RET的细胞质结构域相互作用的信号分子。 我们还将使用我们的模型MDCK细胞开发一种差异筛查 系统识别由GDNF/RET途径激活的基因。 这项建议将解决潜在的分子机制 GDNF/RET介导细胞迁移和趋化作用。除了……之外 促进我们对基本发展过程的理解， 癌基因RET来源的肿瘤细胞的迁移和侵袭机制 也会被照亮。

The development of the mammalian kidney requires the inductive interactions of two progenitor tissues, the ureteric bud epithelium and the metanephric mesenchyme. Signals emanating from the ureteric bud induce the mesenchyme to proliferate and differentiate, thus generating most of the epithelial cell types in the nephron. Reciprocally, mesenchyme derived signals induce the ureteric bud epithelium to proliferate and undergo branching morphogenesis to generate the collecting duct system. Although these signals and their receptors have remained elusive, their mechanisms of action ar fundamental to understanding the molecular basis of renal cell growth and differentiation. The receptor tyrosine kinase RET is expressed at the tips of the ureteric bud and regulates proliferation and branching morphogenesis. Recent work in the PI's lab has demonstrated that the glial cell derived growth factor (GDNF) interacts with RET and stimulates RET phosphorylation. GDNF and RET are part of a common signaling pathway that promotes ureteric bud proliferation and branching. This pathway may not only be critical for ealry kidney development and growth, but may also function in aberrant proliferation of ureteric bud derived cells, such as in cystic diseases of the collecting duct. We have established a biological assay for RET activity in transformed MDCK renal epithelial cells. These cells show increased scattering, cell motility, and morphological changes in response to RET activation. It is now well established that the GDNF/RET pathway promotes migration of ureteric bud epthelium towards the metanephric mesenchyme, a potent source of GDNF. Our preliminary data shows that RET expressing epithelial cells can migrate towards a localized source of GDNF. Thus, ureteric bud outgrowth is due to chemoattraction towards a target derived guidance cue. How the GDNF signal is received and interpreted by the epithelial cell is the major focus of this proposal. The specific aims will utilize the yeast two hybrid system to identify signaling molecules that interact wih the cytoplasmic domain of RET. We will also develop a differential screeen using our model MDCK cell system to identify genes that are activated by the GDNF/RET pathway. This proposal will address the underlying molecular mechanisms of GDNF/RET mediated cell migration and chemoattaction. In addition to advancing our understanding of basic developmental processes, the mechnisms of migration and invasion by oncogenic RET derived tumor cells will also be illuminated.