RESUMO
12(S)-Hydroxyeicosatetraenoic acid (12(S)-HETE), a 12-lipoxygenase metabolite of arachidonic acid, has multiple effects on tumor and endothelial cells, including stimulation of invasion and angiogenesis. However, the signaling mechanisms controlling these physiological processes are poorly understood. In a human epidermoid carcinoma cell line (i.e. A431), 12(S)-HETE activates extracellular signal-regulated kinases 1/2 (ERK1/2), which is mediated by upstream kinases MEK and Raf. 12(S)-HETE stimulates phosphorylation of phospholipase Cgamma1 and activity of protein kinase Calpha (PKCalpha). In addition, independent of PKC 12(S)-HETE increases tyrosine phosphorylation of Shc, and Grb2, stimulates association between Shc and Src, and increases the activity of Ras, via Src family kinases. Furthermore, at low (10-100 nm) concentrations 12(S)-HETE counteracts epidermal growth factor-stimulated activation of ERK1/2 via stimulating protein tyrosine phosphatases. We also present evidence that 12(S)-HETE stimulates ERK1/2 via G proteins and that A431 cells have multiple binding sites for 12(S)-HETE. Finally, inhibition of 12-lipoxygenase induced apoptosis of A431 cells, which was reversed by addition of exogenous 12(S)-HETE. Collectively we demonstrate that the activation of ERK1/2 by 12(S)-HETE may be regulated by multiple receptors triggering PKC-dependent and PKC-independent pathways in A431 cells.
Assuntos
Ácido 12-Hidroxi-5,8,10,14-Eicosatetraenoico/farmacologia , Carcinoma de Células Escamosas/enzimologia , MAP Quinase Quinase Quinase 1 , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Proteína Quinase C/metabolismo , Transdução de Sinais/fisiologia , Araquidonato 12-Lipoxigenase/metabolismo , Ativação Enzimática , Fator de Crescimento Epidérmico/antagonistas & inibidores , Fator de Crescimento Epidérmico/farmacologia , Humanos , Isoenzimas/metabolismo , Cinética , Inibidores de Lipoxigenase , Proteína Quinase 3 Ativada por Mitógeno , Fosfolipase C gama , Fosforilação , Proteína Quinase C-alfa , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Tirosina Fosfatases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Células Tumorais Cultivadas , Fosfolipases Tipo C/metabolismoRESUMO
The arachidonic acid metabolite of 12 lipoxygenase, 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE) promotes metastatic behavior of tumor cells. In this study we set out to identify 12(S)-HETE signaling pathways, and their contribution to cellular functions in A431 epidermoid carcinoma. (1) 12(S)-HETE stimulated phosphotyrosine associated PI3 kinase activity. (2) 12(S)-HETE stimulated ERK1/2 in a PI3 kinase dependent manner. (3) PI3 kinase affected the 12(S)-HETE stimulated Raf/MEK/ERK cascade at the level of MEK. (4) 12(S)-HETE stimulated ERK1/2 via PKCzeta. (5) 12(S)-HETE stimulated cell migration on laminin, which was eliminated by PI3 kinase and cPKC inhibitors, but it was unaffected by inhibition of ERK1/2.