title: Ca2+ and pH determine the interaction of chromaffin cell scinderin with phosphatidylserine and phosphatidylinositol 4,5,-biphosphate and its cellular distribution during nicotinic-receptor stimulation and protein kinase C activation.
abstract: Nicotinic stimulation and high K(+)-depolarization of chromaffin cells cause disassembly of cortical filamentous actin networks and redistribution of scinderin, a Ca(2+)-dependent actin filament-severing protein. These events which are Ca(2+)-dependent precede exocytosis. Activation of scinderin by Ca2+ may cause disassembly of actin filaments leaving cortical areas of low cytoplasmic viscosity which are the sites of exocytosis (Vitale, M. L., A. Rodríguez Del Castillo, L. Tchakarov, and J.-M. Trifaró. 1991. J. Cell. Biol. 113:1057-1067). It has been suggested that protein kinase C (PKC) regulates secretion. Therefore, the possibility that PKC activation might modulate scinderin redistribution was investigated. Here we report that PMA, a PKC activator, caused scinderin redistribution, although with a slower onset than that induced by nicotine. PMA effects were independent of either extra or intracellular Ca2+ as indicated by measurements of Ca2+ transients, and they were likely to be mediated through direct activation of PKC because inhibitors of the enzyme completely blocked the response to PMA. Scinderin was not phosphorylated by the kinase and further experiments using the Na+/H+ antiport inhibitors and intracellular pH determinations, demonstrated that PKC-mediated scinderin redistribution was a consequence of an increase in intracellular pH. Moreover, it was shown that scinderin binds to phosphatidylserine and phosphatidylinositol 4,5-biphosphate liposomes in a Ca(2+)-dependent manner, an effect which was modulated by the pH. The results suggest that under resting conditions, cortical scinderin is bound to plasma membrane phospholipids. The results also show that during nicotinic receptor stimulation both a rise in intracellular Ca2+ and pH are observed. The rise in intracellular pH might be the result of the translocation and activation of PKC produced by Ca2+ entry. This also would explain why scinderin redistribution induced by nicotine is partially (26-40%) inhibited by inhibitors of either PKC or the Na+/H+ antiport. In view of these findings, a model which can explain how scinderin redistribution and activity may be regulated by pH and Ca2+ in resting and stimulated conditions is proposed.
authors.affiliation: Department of Pharmacology, Faculty of Medicine, University of Ottawa, Ontario, Canada.
authors.forename: A
authors.initials: A
authors.lastname: Rodríguez Del Castillo
authors.forename: M L
authors.initials: ML
authors.lastname: Vitale
authors.forename: J M
authors.initials: JM
authors.lastname: Trifaró
mesh_terms: D000313:Adrenal Medulla / Q000166:cytology / Q000378:metabolism*; D000584:Amiloride / Q000494:pharmacology; D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D002417:Cattle; D002478:Cells, Cultured; D004789:Enzyme Activation; D018260:Gelsolin; D006863:Hydrogen-Ion Concentration; D007476:Ionophores / Q000494:pharmacology; D007700:Kinetics; D008840:Microfilament Proteins / Q000378:metabolism*; D008954:Models, Biological; D009538:Nicotine / Q000494:pharmacology; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D010718:Phosphatidylserines / Q000378:metabolism; D010743:Phospholipids / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D011978:Receptors, Nicotinic / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D018260:Gelsolin; D007476:Ionophores; D008840:Microfilament Proteins; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates; D010718:Phosphatidylserines; D010743:Phospholipids; D011978:Receptors, Nicotinic; C065395:scinderin; D000643:Ammonium Chloride; D009538:Nicotine; D000584:Amiloride; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium