Role of protein kinase C and phosphatases in 12(S)-HETE-induced tumor cell cytoskeletal reorganization.

Adherent B16 amelanotic melanoma (B16a) cells exposed to fatty acid 12(S)-HETE, a lipoxygenase metabolite of arachidonic acid, demonstrated a gradual dissolution of stress fibers and bundling-together of vimentin. The 12(S)-HETE effects on tumor cell cytoskeleton appeared 5 min after treatment, became prominent approximately 15 min following stimulation, and generally disappeared by 30 min. Simultaneous treatment of cells with 12(S)-HETE and okadaic acid (OA) prevented disappearance of the 12(S)-HETE effects by 30 min. Quantitative double immunoblotting of actin and vimentin indicated that actin, but not vimentin, underwent a time-related depolymerization. On the other hand, enhanced phosphorylation of vimentin but not of actin was observed after 12(S)-HETE treatment. 12(S)-HETE-enhanced vimentin phosphorylation was abolished by protein kinase C (PKC) inhibitor calphostin C, thus suggesting the involvement of PKC.