title: Endogenous surface expression of ΔF508-CFTR mediates cAMP-stimulated Cl(-) current in CFTR(ΔF508/ΔF508) pig thyroid epithelial cells.
abstract: The cystic fibrosis transmembrane conductance regulator (CFTR) is both an anion channel and a regulator of other transport proteins. Mutations in the CFTR gene underlie the human disease, cystic fibrosis. The most common CFTR mutation, ΔF508, produces a misfolded protein which traffics improperly. The availability of transgenic CFTR(ΔF508/ΔF508) pigs allows measurement of the impact of ΔF508 in native tissue. Thyroid epithelia respond to cAMP-elevating agents by increasing anion transport, a process reliant on functional CFTR. To assess whether endogenous levels of ΔF508-CFTR mediate thyroid transport, primary thyroid epithelial cultures (pThECs) were grown from newborn CFTR(+/+) (wild-type) and CFTR(ΔF508/ΔF508) (ΔF) pig thyroids and the stimulated, secretory components of short-circuit current (I(sc)) compared. Surface biotinylation studies assessed the surface presentation of ΔF508-CFTR. Baseline I(sc) levels of both wild-type and ΔF pThECs consisted of an amiloride-sensitive component. In ΔF pThECs, this mirrored previous measurements in CFTR(-/-) (knockout) pThECs. Surprisingly, elevation of cAMP transiently increased I(sc) to peak levels ∼65% of those achieved by wild-type. In contrast, knockout pThECs were indifferent to cAMP activation. In ΔF pThECs, total ΔF508-CFTR expression was ∼9% that of wild-type, consistent with misfolding and enhanced degradation. Surface biotinylation studies indicated that ∼4% of the total ΔF508 resided at the surface and did not increase with cAMP elevation. The present findings show that low endogenous levels of pig ΔF508-CFTR can mediate substantial anion transport by thyroid epithelia. These data suggest that both wild-type and ΔF508-CFTR regulate additional thyroid transporters, and together co-ordinate the overall I(sc) response.
authors.affiliation: Department of Anatomy and Physiology, Kansas State University College of Veterinary Medicine, 1600 Denison Avenue, Manhattan, KS 66506, USA.
authors.forename: Yonghai
authors.initials: Y
authors.lastname: Li
authors.forename: Suhasini
authors.initials: S
authors.lastname: Ganta
authors.forename: Peying
authors.initials: P
authors.lastname: Fong
mesh_terms: D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D030801:Animals, Genetically Modified; D000838:Anions / Q000378:metabolism; D002478:Cells, Cultured; D018118:Chloride Channels / Q000378:metabolism*; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism*; D003550:Cystic Fibrosis / Q000235:genetics / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism; D055786:Gene Knockout Techniques; D017136:Ion Transport; D008564:Membrane Potentials / Q000235:genetics / Q000502:physiology; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D017510:Protein Folding; D021381:Protein Transport; D059748:Proteolysis; D034421:Sus scrofa / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D013552:Swine; D013696:Temperature; D013961:Thyroid Gland / Q000166:cytology / Q000378:metabolism*
chemical_list: D000838:Anions; D018118:Chloride Channels; D008565:Membrane Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D005576:Colforsin; D000584:Amiloride; D000242:Cyclic AMP