GTP-GAMMA-S-STIMULATED PHOSPHOLIPASE-D ACTIVATION IN HUMAN NEUTROPHILS OCCURS BY PROTEIN-KINASE-C-DEPENDENT AND PROTEIN-KINASE-INDEPENDENT PATHWAYS BUT NOT VIA TYROSINE KINASES

Addition of GTP gamma S to saponin-permeabilised human neutrophils act ivated both the NADPH oxidase and phospholipase D (PLD). This PLD acti vation was hardly affected by staurosporine or Ro31-8220 (at concentra tions which inhibited PMA stimulated PLD activity), indicating that it was largely independent of protein kinase C (PKC). This GTP gamma S s timulated PLD activity was enhanced by 1 mM ATP, but this ATP-enhanced activity was blocked by inhibitors of PKC. Addition of GTP gamma S re sulted in very low levels of phosphorylation on tyrosine residues, but higher levels of phosphorylation on serine/threonine residues. Additi on of pervanadate hydroperoxides stimulated phosphorylation on tyrosin e residues and activated PLD which was blocked by addition of inhibito rs of tyrosine kinases. Thus, GTP gamma S can stimulate PKC-dependent and -independent pathways of PLD activation. Whilst phosphorylation on tyrosine residues can result in activation of PLD, this is regulated independently of activation via G-proteins. (C) 1996 Academic Press, I nc.