Role of protein phosphorylation in activation of phospholipase A(2) by thepolychlorinated biphenyl mixture Aroclor 1242

Polychlorinated biphenyls (PCBs) activate neutrophils to induce degranulati on and undergo superoxide production through a mechanism that involves stim ulation of phospholipase A(2) (PLA(2)). Since the biochemical processes lea ding to the PCE-induced activation of this enzyme are unknown, the objectiv e of this study was to determine whether protein phosphorylation has a role in this mechanism. Isolated rat neutrophils were labeled with [H-3]-arachi donic acid ([H-3]-AA), and activation of PLA(2) was determined from release of radioactivity into the medium. Exposure to the PCB mixture Aroclor 1242 induced release of [H-3]-AA, and pretreatment with bromoenol lactone (BEL) , an inhibitor of calcium-independent PLA(2), diminished release by 80%. Ge nistein, an inhibitor of tyrosine kinases, caused a small but significant d ecrease in Aroclor 1242-stimulated release of [H-3]-AA. Daidzein, a geniste in analog with no activity to inhibit tyrosine kinases, had no effect on [H -3]-AA release. An inhibitor of p38 mitogen-activated protein kinase (MAPK) , SB203580, did not affect Aroclor 1242-induced PLA(2) activity at concentr ations selective for p38 MAPK; however, PD 98059, which inhibits MAPK kinas e (MEK), decreased [H-3]-AA release to about the same extent as genistein. Treatment of neutrophils with Aroclor 1242 induced phosphorylation of p44 M APK, and this phosphorylation was unaffected by EEL but was inhibited by PD 98059. Staurosporine, a nonselective: inhibitor of protein kinase C (PKC), inhibited PCB-induced ren lease of [H-3]-AA. Ro 32-0432, a selective inhib itor of PKCalpha and PKCbeta 1, produced the greatest degree of inhibition (40%) among the tested protein kinase inhibitors. These results suggest tha t tyrosine kinases, PKC, and the MEK/MAPK pathway are involved in a fractio n of Aroclor 1242-induced activation of PLA(2). (C) 2000 Academic Press.