CELLULAR ACTIONS OF CATECHOLAMINE RECEPTORS

儿茶酚胺受体的细胞作用

• 批准号: 2176393
• 负责人: PAUL A INSEL
• 金额: $ 22.26万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-01-01 至 1995-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2176393
• 关键词: SDS polyacrylamide gel electrophoresis; affinity labeling; alpha adrenergic receptor; animal genetic material tag; animal tissue; autoradiography; biological signal transduction; cardiovascular disorder; catecholamines; cell cell interaction; cell type; clone cells; gel electrophoresis; genetic library; kidney cell; nucleic acid hybridization; phospholipase A2; phospholipase D; pinocytosis; protein kinase C; protein structure function; radiotracer; receptor binding; thin layer chromatography; tissue /cell culture; western blottings

This project continues my laboratory's studies of cellular actions of alpha1-adrenergic receptors using a clonal isolate of Madin Darby kidney (MDCK) cells. In recent studies our laboratory has shown that in these cells alpha1-adrenergic receptors may be of a unique subtype that links to a guanine nucleotide binding (G) protein and in turn to multiple phospholipases, including one or more phospholipases A2, C and D. Regulation of phospholipase A2 by alpha1-adrenergic receptors occurs in part via protein kinase C, in particular, by the alpha isoform of this enzyme. the current studies are designed to test hypotheses related to alpha-adrenergic receptor structure, phospholipase regulation, and protein kinase C isoforms in MDCK-D1 cells. Pharmacological methods will be used to define alpha1-receptor subtypes and subsequent molecular biological methods will be used to characterize the subtype(s) present in MDCK D1 cells. Studies of phospholipases will involve biochemical characterization of phospholipase D and phospholipase A2. The third aim is to assess the role of isoforms of protein kinase C in cell regulation, especially in regulation of phospholipase A2. The methods to be used in these latter studies include treatment of cells with antisense oligonucleotides and transient and stable transfection with antisense cDNA's to generate cells null with respect to individual C-kinase isoforms. Other approaches will involve microscopy with immunological probes for particular isoforms. Taken together, the studies proposed should provide new information regarding alpha1-adrenergic receptor action and the role of protein kinase C isoforms in regulation of cellular function. Given the importance of alpha1-adrenergic receptors and protein kinase C for a wide variety of cellular functions, the results may have importance to disease settings such as cancer and several types of cardiovascular disorders.

这个项目继续我的实验室研究的研究 使用Madin Darby肾脏的克隆分离物α1-肾上腺素能受体 （MDCK）细胞。 在最近的研究中，我们的实验室表明，在这些研究中 细胞α1-肾上腺素受体可能是独特的亚型，与 鸟嘌呤核苷酸结合（G）蛋白，然后又 磷脂酶，包括一个或多个磷脂酶A2，C和D。 通过α1-肾上腺素受体调节磷脂酶A2发生在 部分通过蛋白激酶C（特别是通过α同工型） 酶。 当前的研究旨在检验与 α-肾上腺素受体结构，磷脂酶调节和蛋白质 MDCK-D1细胞中的激酶C同工型。 将使用药理方法 定义α1受体亚型和随后的分子生物学 方法将用于表征MDCK D1中存在的亚型 细胞。 磷脂酶的研究将涉及生化特征 的磷脂酶D和磷脂酶A2。 第三个目的是评估 蛋白激酶C同工型在细胞调节中的作用，尤其是在 调节磷脂酶A2。 这些方法在后者中使用 研究包括用反义寡核苷酸和 用反义cDNA的瞬时和稳定转染产生细胞 相对于单个C-激酶同工型零。 其他方法也会 与特定同工型的免疫学探针有关显微镜检查。 综上所述，提出的研究应提供新信息 关于α1-肾上腺素受体的作用和蛋白激酶的作用 c在调节细胞功能方面的同工型。 考虑到重要性 α1-肾上腺素能受体和蛋白激酶C多种多样 细胞功能，结果可能对疾病环境有重要意义 例如癌症和几种类型的心血管疾病。