MOLECULAR CHARACTERIZATION OF CHLORIDE CHANNEL PROTEINS

该项目的总体目标是从分子水平上了解 氯离子通道在健康和疾病中的作用氯离子通道在细胞内 在许多细胞和组织过程中起作用;氯离子通道功能障碍是 已知在人类疾病中具有主要致病意义 囊性纤维化先天性肌强直和霍乱充分了解 氯离子通道的结构和功能将提供深入了解 这些疾病的发病机制，并可能提出新的治疗方法。 我们克隆了编码64 kD氯离子通道蛋白的cDNA， 牛肾该基因的表达是表达 囊泡介导的cAMP激活的爪蟾卵母细胞氯通道 纤维化跨膜传导调节因子（CFTR）。这样做的目的 研究P64及其在各种氯离子通道中的作用 包括与CFTR相关的cAMP激活的氯离子通道。的 具体目标是：1）表征P64相关氯离子通道 通过免疫沉淀、重建和体内磷酸化，2） 分离编码P64的人同源物的cDNA，和3）衍生一个 使用反义抑制P64表达的P64表达系统， 内源基因和野生型或突变形式的P64的再引入 用来逃避反义抑制

The overall goal of the project is to gain a molecular understanding of chloride channels in health and disease. Chloride channels play essential roles in many cell and tissue processes; chloride channel dysfunction is known to be of primary pathogenic significance in the human diseases cystic fibrosis, myotonia congenita, and cholera. A full understanding of chloride channel structure and function will provide insight into the pathogenesis of these diseases and may suggest new therapeutic approaches. We have cloned the cDNA encoding a 64 kD chloride channel protein from bovine kidney. Expression of this gene is required for expression of the cAMP-activated chloride channel induced in Xenopus oocytes by the cystic fibrosis transmembrane conductance regulator (CFTR). The purpose of this proposal is to study P64 and its role in various chloride channels including the cAMP-activated chloride channel associated with CFTR. The specific aims are to: 1) Characterize the P64 associated chloride channel by immunoprecipitation, reconstitution and in vivo phosphorylation, 2) Isolate cDNAs encoding the human homologue of P64, and 3) Derive an expression system for P64 using antisense suppression of expression of the endogenous gene and reintroduction of wild type or mutated versions of P64 designed to evade the antisense suppression.