MOLECULAR CHARACTERIZATION OF CHLORIDE CHANNEL PROTEINS

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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.