Signal transduction pathways for activation of extracellular signal-regulated kinase by arachidonic acid in rat neutrophils

Phosphorylation of extracellular signal-regulated kinase (ERK) in response to arachidonic acid (AA) was rapid and transient, peaking at 1 min and disa ppearing after 3 min, and it was accompanied by an increase in ERK activity in rat neutrophils, We examined the upstream regulation of AA-stimulated E RK activation using one of the following signaling pathway inhibitors to pr etreat rat cells: the ERK kinase inhibitor U0126 or PD98059, the G(i/o) inh ibitor pertussis toxin (PTX), the tyrosine kinase inhibitor genistein, the phosphatidylinositol 3-kinase (PI3K) inhibitor wort-mannin or LY294002, the Ca2+ chelator 1,2-bis(O-aminophenoxy) ethane-N,N,N',N'-tetraacetic acid, o r the phospholipase C (PLC) inhibitor U73122. All of these inhibitors atten uated AA-induced ERK activation. Activation of ERK was also effectively att enuated by the cyclooxygenase and lipoxygenase inhibitor BW755C and by the leukotriene biosynthesis inhibitor MK886, but the cyclooxygenase inhibitor indomethacin did not attenuate ERK activation. After exposing cells to thre e distinct protein kinase C (PKC) inhibitors, we found that Go6976 signific antly attenuated ERK phosphorylation but potentiated ERK activity. Neither Go6983 nor GF109203X affected AA-induced responses. These data suggest that the li-poxygenase metabolite(s) produced mediates AA-stimulated ERK activa tion and that this effect is upstream regulated by PT-sensitive G protein, nonreceptor tyrosine kinase, PI3K, and PLC/Ca2+ signaling pathways in rat n eutrophils.