Evidence that Ca2(+)-dependent activation of airway epithelia Cl- channels is not dependent on phosphorylation.
Previous studies have shown that an increase in the cytosolic Ca2+ concentration [( Ca2+]c) activates Cl- channels in airway epithelia but that the effect is indirect. Because adenosine 3',5'-cyclic monophosphate (cAMP) and phorbol myristate acetate (PMA) activate Cl- channels via phosphorylation by cAMP-dependent protein kinase and protein kinase C, respectively, we asked whether Ca2(+)-dependent Cl- channel activation is phosphorylation dependent. We measured 125I- efflux as an assay of Cl- channel activation in the intact cell. We found that depletion of cellular ATP prevented cAMP- and PMA-induced activation but did not alter activation produced by the Ca2+ ionophore A23187. Moreover, addition of high concentrations of staurosporine (5 microM), to nonspecifically inhibit kinase activity, blocked cAMP- and PMA-stimulated 125I- efflux but had no effect on A23187-induced efflux. These results suggest that elevation of [Ca2+]c does not activate Cl- channels via phosphorylation.
Howard Hughes Medical Institute, Department of Internal Medicine, University of Iowa College of Medicine, Iowa City 52242.
J P
JP
Clancy
J D
JD
McCann
M J
MJ
Welsh
D000255:Adenosine Triphosphate / Q000378:metabolism; D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000502:physiology*; D002478:Cells, Cultured; D018118:Chloride Channels; D000242:Cyclic AMP / Q000378:metabolism; D003600:Cytosol / Q000502:physiology; D004285:Dogs; D004848:Epithelium / Q000502:physiology; D007454:Iodides / Q000378:metabolism; D007473:Ion Channels / Q000187:drug effects / Q000502:physiology*; D007700:Kinetics; D008565:Membrane Proteins / Q000502:physiology*; D009130:Muscle, Smooth / Q000502:physiology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014132:Trachea / Q000502:physiology*
D000470:Alkaloids; D018118:Chloride Channels; D007454:Iodides; D007473:Ion Channels; D008565:Membrane Proteins; D000001:Calcimycin; D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D011493:Protein Kinase C; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium