Activation of phospholipase D by calmodulin antagonists and mastoparan in carnation petals

An in vivo assay for phospholipase D (PLD; EC 3.1.4.4) activity, based on its transphosphatidylation property, is described in detail and was used to study putative post-translational regulation mechanisms of PLD activity in carnation (Dianthus caryophyllus L.) petals. A variety of agents was applied to petal discs, The calmodulin (CaM) antagonists propranolol, N-(6-aminohexyl)-5-chloro-1-naphthalenesulphonamide (W7) and N-(6-aminohexyl)-1-naphthalenesulphonamide (W5), stimulated PLD activity in a dose-dependent manner. EGTA partially inhibited the stimulation by the CaM antagonists. Erythrosin B, an inhibitor of CaM-dependent P-type Ca2+-ATPases, slightly stimulated PLD activity. The results suggest that part of the stimulation of PLD activity by CaM antagonists is due to an increased intracellular Ca2+-concentration. PLD activity was stimulated by mastoparan in a dose-and time-dependent manner. The signal-like activation kinetics suggests that mastoparan activates PLD (in)directly via a G protein.