G PROTEIN-DEPENDENT SIGNALING IN RENAL EPITHELIAL CELLS

这项研究计划的长期目标是了解 细胞内的分子机制， 信号系统调节肾上皮细胞功能。 G蛋白- 偶联受体（例如，对于细胞外Ca、A11和AVP）激活 涉及不同G蛋白，激酶， 和转录因子，以提供整合的控制网络， 调节多种细胞功能。 尽管配体 这些受体及其对第二信使的影响， 运输已经研究得相当详细，其机制 信息在细胞中的传播仍然是不完全的 定义的原因是：1）许多蛋白质在 结构和功能; 2）它们在低水平表达; 3） 化学活化剂或抑制剂并不存在， 蛋白质;和4）许多蛋白质可能仍然有待鉴定。 将检查细胞外Ca受体（Cao-R）的信号传导 使用允许分离信令过程的模型系统 转化成可以精确分析的成分， 假设 其组成部分是：1）G受体的模式 蛋白偶联; 2）细胞内激酶的激活， G蛋白; 3）Na/H交换启动子（NHE-1）的激活 通过单个G蛋白和激酶;和4）鉴定 响应G蛋白的NHE-1启动子元件， 激酶。 我们将检验的假设是：1）Ca o -R激活 多个G蛋白（Gq~ G12'和一个Gi）：2） 通过不同的机制与Cao-R信号偶联， 不同的MAP激酶; 3）G蛋白和激酶激活 NHE-1启动子的不同元件;和4） 调节Na/H交换也调节其他转运蛋白， 协调的方式。 研究将在细胞中进行， 允许同时进行遗传和生物化学分析， 分离的MTAL细胞，以验证遗传和生化研究 并培养MTAL细胞来测试这些信号的作用 运输监管制度。 建立机制， Cao-R的行为将提高我们对遗传学的理解， 与Cao-R和电解质异常有关的疾病， 与高钙血症相关的间质性肾炎。

The long term goals of this research program is to understand the intracellular molecular mechanisms by which G protein-dependent signaling systems regulate renal epithelial cell function. G protein- coupled receptors (eg. for extracellular Ca, A11, and AVP) activate multiple parallel pathways involving distinct G proteins, kinases, and transcription factors to provide an integrated control network to regulate multiple cell functions. Despite the fact that the ligands for these receptors and their effects on second messengers and transport have been studied in considerable detail, the mechanisms by which information travels through the cell remain incompletely defined because: 1) many of the proteins are closely related in structure and function; 2) they are expressed at low levels; 3) chemica activators or inhibitors do not exist for many known proteins; and 4) many proteins probably remain to be identified. Signaling by the extracellular Ca receptor (Cao-R) will be examined using a model system that allows separation of the signaling process into components that can be analyzed with precision to test specific hypotheses. The components are: 1) the pattern of receptor - G protein coupling; 2) activation of intracellular kinases by individual G proteins; 3) activation of the Na/H exchanger promoter (NHE-1) by individual G proteins and kinases; and 4) identification of the NHE-1 promoter elements that respond to the G proteins and kinases. The hypotheses we will test are: 1) the Ca o -R activates of multiple G proteins (Gq~ G12' and a Gi): 2) the G proteins that couple to the Cao-R signal by different mechanisms, and activate different sets of MAP kinases; 3) the G proteins and kinase activate different elements of the NHE-1 promoter; and 4) the signals that regulate Na/H exchange also regulate other transporters in a coordinated manner. The studies will be performed in cells that allow simultaneous genetic and biochemical analyses, freshly isolated MTAL cells to verify the genetic and biochemical studies and cultured MTAL cells to test the effects of these signaling systems on regulation of transport. Establishing the mechanisms by which the Cao-R acts will improve our understanding of the genetic diseases linked to the Cao-R and the electrolyte abnormalities and interstitial nephritis associated with hypercalcemia.