New information obtained during the past five years has pointed to an abnormality in secretory chloride channel regulation as the basic defect in cystic fibrosis (CF). The apical membrane-located chloride channel represents the rate-limiting step for Cl- secretion and, thus, influences the composition of the secretory products of the airways, pancreas, and sweat glands, the major target organs in CF. Although much important knowledge has been learned about the phenomenology of epithelial secretory Cl- channel kinetics and regulation by cAMP-and Ca++-dependent pathways, virtually nothing is known about the structural details of the protein(s) comprising the anion channel complex. The biochemical identification of secretory Cl- channels has been hampered by the lack of suitable probe molecules. The central goal of this Cystic Fibrosis Research Center application is to isolate and characterize this secretory C1- channel protein complex and any associated regulatory components so that a molecular basis for CF can be specified. The development of this proposal revolves around the generation of antibodies that appear to interact specifically with the secretory Cl- channel-containing vesicles. Four interdisciplinary, interrelated projects are proposed: 1) isolation of secretory Cl- channels using antibody probes: 2) cloning and expression of the secretory Cl- channel; 3) immunocytochemical localization and intracellular trafficking of Cl- channels; and 4) disulfonic stilbenes as probes of reconstituted Cl- channels. Thus, the secretory Cl- channel will be studied at both the molecular and physiological levels to [provide a comprehensive framework for understanding the mechanisms of anion conduction and regulation. By concentrating our efforts on the molecular and cell biological aspects of C1- channel structure and movement, new information will be generated that will undoubtedly lead to novel strategies for pharmacological intervention to overcome the regulatory deficiencies occurring in CF.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Specialized Center (P50)
Project #
5P50DK042017-03
Application #
3105902
Study Section
Diabetes, Endocrinology and Metabolic Diseases B Subcommittee (DDK)
Project Start
1989-09-01
Project End
1994-08-31
Budget Start
1991-09-01
Budget End
1992-08-31
Support Year
3
Fiscal Year
1991
Total Cost
Indirect Cost
Name
University of Alabama Birmingham
Department
Type
Schools of Dentistry
DUNS #
004514360
City
Birmingham
State
AL
Country
United States
Zip Code
35294
Schwiebert, E M; Benos, D J; Fuller, C M (1998) Cystic fibrosis: a multiple exocrinopathy caused by dysfunctions in a multifunctional transport protein. Am J Med 104:576-90
Ji, H L; DuVall, M D; Patton, H K et al. (1998) Functional expression of a truncated Ca(2+)-activated Cl- channel and activation by phorbol ester. Am J Physiol 274:C455-64
Tousson, A; Fuller, C M; Benos, D J (1996) Apical recruitment of CFTR in T-84 cells is dependent on cAMP and microtubules but not Ca2+ or microfilaments. J Cell Sci 109 ( Pt 6):1325-34
Jovov, B; Ismailov, I I; Benos, D J (1995) Cystic fibrosis transmembrane conductance regulator is required for protein kinase A activation of an outwardly rectified anion channel purified from bovine tracheal epithelia. J Biol Chem 270:1521-8
Cunningham, S A; Awayda, M S; Bubien, J K et al. (1995) Cloning of an epithelial chloride channel from bovine trachea. J Biol Chem 270:31016-26
Sanchez-Olea, R; Fuller, C; Benos, D et al. (1995) Volume-associated osmolyte fluxes in cell lines with or without the anion exchanger. Am J Physiol 269:C1280-6
Singh, A K; Venglarik, C J; Bridges, R J (1995) Development of chloride channel modulators. Kidney Int 48:985-93
Fuller, C M; Bridges, R J; Benos, D J (1994) Forskolin- but not ionomycin-evoked Cl- secretion in colonic epithelia depends on intact microtubules. Am J Physiol 266:C661-8
Fuller, C M; Ismailov, I I; Keeton, D A et al. (1994) Phosphorylation and activation of a bovine tracheal anion channel by Ca2+/calmodulin-dependent protein kinase II. J Biol Chem 269:26642-50
Venglarik, C J; Singh, A K; Bridges, R J (1994) Comparison of -nitro versus -amino 4,4'-substituents of disulfonic stilbenes as chloride channel blockers. Mol Cell Biochem 140:137-46

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