Cytosolic phospholipase A2 regulation of human monocyte NADPH oxidase activity: cPLA2 affects translocation but not phosphorylation of p67phox and p47phox*

The NADPH oxidase of human monocytes is activated, upon exposure to opsonized zymosan and a variety of other stimuli, to catalyze the formation of superoxide anion. Assembly of the NADPH oxidase complex is believed to be a highly regulated process and molecular mechanisms responsible for this regulation remain to be fully elucidated. We have previously reported that cytosolic phospholipase A2 (cPLA2) expression and activity are essential for superoxide anion production in activated human monocytes. In this study, we investigated the mechanisms involved in cPLA2 regulation of NADPH oxidase activation by evaluating the effects of cPLA2 on translocation and phosphorylation of p67phox and p47phox. We report that translocation and phosphorylation of p67phox, as well as p47phox, occur upon activation of human monocytes and that decreased cPLA2 protein expression, mediated by antisense oligodeoxyribonucleotides (AS-ODN) specific for cPLA2 mRNA, blocked the stimulation-induced translocation of p47phox and p67phox from the cytosol to the membrane fraction. Inhibition of translocation of both p47phox and p67phox by cPLA2 AS-ODN was above 85%. Arachidonic acid, a product of cPLA2 enzymatic activity, completely restored translocation of both of these oxidase components in the AS-ODNtreated, cPLA2-deficient human monocytes. These results represent the first report that cPLA2 activity or AA is required for p67phox and p47phox translocation in human monocytes. Although cPLA2 was required for translocation of p47phox and p67phox, it did not influence phosphorylation of these components. These results suggest that one mechanism of cPLA2 regulation of NADPH oxidase activity is to control the arachidonate-sensitive assembly of the complete oxidase complex through modulating the translocation of both p47phox and p67phox. These studies provide insight into the mechanisms by which activation signals are transduced to allow the induction of superoxide anion production in human monocytes. by gest on M arch 4, 2020 hp://w w w .jb.org/ D ow nladed from