Rac is an important regulatory molecule implicated in c-jun N-terminal kinase (JNK) activation in response to stress and cytokines. However, the signaling events that mediate the activation of JNK by Rac are not yet well characterized. To broaden our understanding of downstream mediators that link Rac signals to the JNK pathway, we investigated whether cytosolic phospholipase A(2) (cPLA(2)) is involved in Rac activation of JNK. In this report we demonstrate that either co-transfection with antisense cPLA(2) oligonucleotide or pretreatment with arachidonyltrifluoromethyl ketone (AACOCF3), a potent and specific inhibitor of cPLA(2), inhibits Rac-mediated JNK activation, implying a potential role of cPLA(2) in Rac-signaling to JNK activation. In accordance with this observation, we demonstrate that the addition of exogenous arachidonic acid (AA), a principal product of Rac-activated cPLA(2), or leukotrienes, products of 5-lipoxygenase (5-LO) of AA, caused a specific stimulation of JNK. Together, our findings suggest that cPLA(2) mediates, at least partly, the signaling cascade by which Rac stimulates JNK.