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Examination of the Role of ADP‐Ribosylation Factor and Phospholipase D Activation in Regulated Exocytosis in Chromaffin and PC12 Cells
Author(s) -
Glenn Daphne E.,
Thomas Geraint M. H.,
O'Sullivan Antony J.,
Burgoyne Robert D.
Publication year - 1998
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.1998.71052023.x
Subject(s) - exocytosis , chromaffin cell , phospholipase d , adp ribosylation factor , phospholipase c , microbiology and biotechnology , chemistry , phospholipase , adp ribosylation , endocrinology , medicine , biology , enzyme , signal transduction , biochemistry , catecholamine , cell , adrenal medulla , nad+ kinase , secretion , golgi apparatus
The possible role of ADP‐ribosylation factor (ARF)‐activated and constitutive phospholipase D (PLD) activity in regulated exocytosis of preformed secretory granules in adrenal chromaffin and PC12 cells was examined. With use of digitonin‐permeabilised cells, the effect of GTP analogues and exogenous ARF1 on PLD activity was determined. No evidence was seen for ARF‐stimulated PLD activity in these cell types. Exocytosis from cytosol‐depleted permeabilised chromaffin cells was not increased by adding recombinant nonmyristoylated or myristoylated ARF1, and exocytosis from both cell types was resistant to brefeldin A (BFA). Addition of bacterial PLD with demonstrably high activity in permeabilised chromaffin cells did not increase exocytosis in cytosol‐depleted chromaffin cells. Diversion of PLD activity from production of phosphatidic acid (PA) due to the presence of 4% ethanol did not inhibit exocytosis triggered by Ca 2+ or poorly hydrolysable GTP analogues in permeabilised chromaffin or PC12 cells. These results indicate that exocytosis in these cell types does not appear to require a BFA‐sensitive ARF and the triggering of exocytosis does not require PLD activity and formation of PA. These findings rule out a general requirement for PLD activity during regulated exocytosis.