The role of protein kinase C in alpha-adrenergic regulation of NaCl(K) cotransport in human airway epithelial cells.
alpha 1-Adrenergic (alpha 1-AR) agents stimulate NaCl(K) cotransport and phosphatidylinositol-4,5-bisphosphate [PtdIns(4,5)P2]-specific phospholipase C in human trachea and nasal polyp epithelial cells. One second messenger generated by PtdIns(4,5)P2 degradation is inositol trisphosphate. We now show that diglycerides (DG) are also generated during alpha 1-AR stimulation. In cells prelabeled with [3H]arachidonic acid, alpha 1-AR agents produced a biphasic DG generation in normal and cystic fibrosis (CF) cells that is blocked by pertussis toxin. The early DG peak closely paralleled PtdIns(4,5)P2 degradation, stimulation of cotransport by phorbol 12-myristate 13-acetate (PMA), and inhibition of cotransport by the protein kinase C (PKC) inhibitor staurosporine. This suggests that cotransporter activation requires PKC-protein phosphorylation. This possibility was tested using the protein phosphatase inhibitor okadaic acid. Okadaic acid elevated bumetanide-sensitive Cl efflux. Staurosporine also blocked > 63% of okadaic-acid-stimulated Cl transport. The late DG peak did not support hormone-stimulated cotransport. The results demonstrate that DGs are a pivotal link between alpha 1-AR stimulation and NaCl(K) cotransport activation with a role for PKC and protein phosphorylation. alpha 1-AR intracellular signaling mechanisms apparently operate normally in CF cells.
Liedtke
C M
CM
Department of Pediatrics and of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio 44106.
D000470:Alkaloids / Q000494:pharmacology; D016718:Arachidonic Acid / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism; D003550:Cystic Fibrosis / Q000378:metabolism; D004075:Diglycerides / Q000378:metabolism; D004837:Epinephrine / Q000494:pharmacology; D004848:Epithelium / Q000378:metabolism; D006801:Humans; D007545:Isoproterenol / Q000494:pharmacology; D008729:Methoxamine / Q000494:pharmacology; D009298:Nasal Polyps / Q000378:metabolism*; D037342:Pertussis Toxin; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011942:Receptors, Adrenergic, alpha / Q000502:physiology*; D049992:Sodium Chloride Symporters; D019311:Staurosporine; D027981:Symporters; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D000470:Alkaloids; D002352:Carrier Proteins; D002712:Chlorides; D004075:Diglycerides; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates; D010750:Phosphoproteins; D011942:Receptors, Adrenergic, alpha; D049992:Sodium Chloride Symporters; D027981:Symporters; D010566:Virulence Factors, Bordetella; D016718:Arachidonic Acid; D037342:Pertussis Toxin; D011493:Protein Kinase C; D019311:Staurosporine; D008729:Methoxamine; D007545:Isoproterenol; D013755:Tetradecanoylphorbol Acetate; D004837:Epinephrine