Glenn D E, Thomas G M, O'Sullivan A J, Burgoyne R D
Physiological Laboratory, University of Liverpool, England, UK.
J Neurochem. 1998 Nov;71(5):2023-33. doi: 10.1046/j.1471-4159.1998.71052023.x.
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 Ca2+ 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.
研究了ADP-核糖基化因子（ARF）激活的和组成型磷脂酶D（PLD）活性在肾上腺嗜铬细胞和PC12细胞中预制分泌颗粒的调节性胞吐作用中的可能作用。使用洋地黄皂苷通透细胞，测定了GTP类似物和外源性ARF1对PLD活性的影响。在这些细胞类型中未发现ARF刺激PLD活性的证据。添加重组非肉豆蔻酰化或肉豆蔻酰化的ARF1不会增加来自耗尽胞质溶胶的通透嗜铬细胞的胞吐作用，并且两种细胞类型的胞吐作用均对布雷菲德菌素A（BFA）具有抗性。在通透的嗜铬细胞中添加具有明显高活性的细菌PLD不会增加耗尽胞质溶胶的嗜铬细胞的胞吐作用。由于存在4%乙醇，PLD活性从磷脂酸（PA）的产生转移，这不会抑制通透的嗜铬细胞或PC12细胞中由Ca2+或难水解的GTP类似物触发的胞吐作用。这些结果表明，这些细胞类型中的胞吐作用似乎不需要对BFA敏感的ARF，并且胞吐作用的触发不需要PLD活性和PA的形成。这些发现排除了调节性胞吐过程中对PLD活性的普遍需求。
