CELL MIGRATION, CHEMOATTRACTION AND THE RET/GDNF PATHWAY

哺乳动物肾脏的发育需要 两个祖细胞组织，输尿管芽上皮和 后肾间充质 输尿管芽发出的信号 诱导间充质细胞增殖分化， 肾单位中的大多数上皮细胞类型。 反过来说， 间充质来源的信号诱导输尿管芽上皮， 增殖并经历分支形态发生， 集气管系统 尽管这些信号及其受体具有 仍然难以捉摸，它们的作用机制对于 了解肾细胞生长的分子基础， 分化 受体酪氨酸激酶RET表达于 输尿管芽的顶端，并调节增殖和分支 形态发生 PI实验室最近的工作表明， 胶质细胞衍生生长因子（GDNF）与RET相互作用， 刺激RET磷酸化。 GDNF和RET是一个共同的 促进输尿管芽增殖的信号通路， 分支 这一通路不仅对每个肾脏至关重要， 发育和生长，但也可能在异常增殖中起作用， 输尿管芽来源的细胞，如在囊性疾病的 集合管 我们已经建立了一个生物测定RET活性的转化 MDCK肾上皮细胞 这些细胞的散射增强， 细胞运动性和响应RET激活的形态学变化。 现在已经确定GDNF/RET途径促进迁移 输尿管芽上皮向后肾间充质，一个强有力的 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，在本发明中， 输尿管芽生长是由于朝向目标的化学吸引 导出的引导提示。 GDNF信号如何被接收和解释 上皮细胞是这个建议的主要焦点。 的 具体的目的是利用酵母双杂交系统来鉴定 与RET的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细胞开发差异筛选 系统来鉴定由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介导的细胞迁移和趋化作用。 除了 促进我们对基本发展过程的理解， 致癌RET源性肿瘤细胞的迁移和侵袭机制 也将被照亮。

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.