Induction of cytosolic phospholipase A2 activity by phosphatidic acid and diglycerides in permeabilized human neutrophils: interrelationship between phospholipases D and A2.
Relationships between phospholipases are poorly understood, but phosphatidic acid (PA) and diglycerides (DGs), produced by phospholipase D (PLD) and phosphatidate phosphohydrolase actions, might function as second messengers coupling cell stimulation to cellular responses. This study investigates the role of PLD-mediated PA and DG formation in inducing phospholipase A2 (PLA2) activity in intact human neutrophils (PMNs) and in PMNs permeabilized with Staphylococcus aureus alpha-toxin. PMNs were labelled with [3H]arachidonic acid (AA) to assess AA release and metabolism and diacylglycerol formation, or with [3H]1-O-hexadecyl-2-lyso-glycerophosphatidylcholine for the determination of platelet-activating factor (PAF), PA and alkylacylglycerol production. In intact PMNs primed with tumour necrosis factor alpha before stimulation with N-formyl-Met-Leu-Phe, AA release and metabolism and PAF formation increased in parallel with enhanced PA and DG formation, and inhibition of PA and DG production led to a decrease in both AA release and PAF accumulation. In alpha-toxin-permeabilized PMNs, AA release and PAF production result from the specific activation of cytosolic PLA2 (cPLA2). In this system, PA and DG formation were always present when cPLA2 activation occurred; blocking PA and DG production inhibited AA release and PAF accumulation. Adding either PA or DG back to permeabilized cells (with endogenous PA and DG formation blocked) led to a partial restoration of AA release and PAF formation; a combination of PA and DGs reconstituted full cPLA2 activity. These results strongly suggest that products of PLD participate in activating cPLA2 in PMNs.
Bauldry
S A
SA
Pulmonary and Critical Care Medicine, Bowman Gray School of Medicine of Wake Forest University, Winston-Salem, NC 27157, U.S.A.
Wooten
R E
RE
D016718:Arachidonic Acid / Q000378:metabolism; D001427:Bacterial Toxins / Q000494:pharmacology; D002463:Cell Membrane Permeability; D003600:Cytosol / Q000201:enzymology; D004075:Diglycerides / Q000096:biosynthesis* / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004790:Enzyme Induction; D000431:Ethanol / Q000494:pharmacology; D006460:Hemolysin Proteins / Q000494:pharmacology; D006801:Humans; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects / Q000201:enzymology*; D010712:Phosphatidic Acids / Q000096:biosynthesis* / Q000494:pharmacology; D010739:Phospholipase D / Q000378:metabolism*; D010741:Phospholipases A / Q000096:biosynthesis*; D054467:Phospholipases A2; D010972:Platelet Activating Factor / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D001427:Bacterial Toxins; D004075:Diglycerides; D006460:Hemolysin Proteins; D010712:Phosphatidic Acids; D010972:Platelet Activating Factor; D014409:Tumor Necrosis Factor-alpha; C021091:staphylococcal alpha-toxin; D016718:Arachidonic Acid; D000431:Ethanol; D009240:N-Formylmethionine Leucyl-Phenylalanine; D010741:Phospholipases A; D054467:Phospholipases A2; D010739:Phospholipase D