Contribution of mitogen-activated protein kinase to stimulation of phospholipase D by the chemotactic peptide fMet-Leu-Phe in human neutrophils

Phospholipase D (PLD) plays an important role in signaling through phosphat idylcholine (PC) and in the production of superoxide (respiratory burst) by polymorphonuclear leukocytes (PMN) stimulated by the chemoattractant fMet- Leu-Phe (fMLP). However, the regulation of PLD activity by protein kinases is not fully understood. In the present study, we have used a mitogen-activ ated protein (MAP) kinase inhibitor (PD 98059) to investigate a possible co nnection between extracellular signal-regulated kinase (ERK) and PLD activi ty and respiratory burst. Using a range of concentrations (3-20 mu M) which inhibit ERK activity, PD 98059 inhibited PLD activity induced by fMLP in c ytochalasin B-primed PMN, as assessed by production-tritiated phosphatidyle thanol (PEt), phosphatidic acid (PA), and hydrolysis of PC. However, the in hibition was partial (approximately 50%), while inhibition of PC hydrolysis was almost complete, suggesting a concomitant inhibition of PLA2 activity. In addition, PD 98059 reduced fMLP-induced respiratory burst by 50%, an ef fect which was correlated with PLD inhibition of PLD (r = 0.981, P < 0.01), and neither did PD 98059 inhibit the PLD activity and respiratory burst in duced by PKC upon its direct activation by phorbol myristate acetate. These data provide the first evidence for implication of the ERK cascade in the stimulation of PLD through Gi signaling. They further indicate that PLD sti mulation by fMLP receptors occurs through two pathways, dependent and indep endent on MAP kinase, the former pathway being linked to superoxide product ion. (C) 1999 Academic Press.