Protein tyrosine kinases, p56^lck and p59^fyn, and MAP kinase JNK1 provide and early signal required for upregulation of Fas ligand expression in aburatubolactam C-induced apoptosis of human Jurkat T cells

The signaling mechanism underlying aburatubolactam C-induced FasL upregulation was investigated in human Jurkat T cells. After treatment with aburatubolactam C, the src-family PTKs p56^lck and p59^fyn and MAP kinases ERK2 and JNK1, were activated prior to FasL upregulation. Both p56^lck and p59^fyn were directly activated 2.4- and 2.2-fold, respectively, in vitro by aburatabolactam C. The aburatubolactam C-induced cellular changes, including the activation of ERK2 and JNK1, and FasL upregulation, were completely prevented by the PTK inhibitor genistein. The activation of protein kinase C (PKCδ, ε, and μ) was also induced following aburatubolactam C treatment. Although the activation of p56^lck and tyrosine phosphorylation of the cellular proteins were not blocked by the PKC inhibitor GF109203X, the activation of ERK2 was completely abrogated, along with a detectably enhanced JNK1 activation, FasL upregulation, and apoptosis. However, the FasL upregulation and apoptosis were significantly inhibited by the PKC activator PMA, with a remarkable increase in the ERK2 activation. The cytotoxic effect of aburatubolactam C was reduced in the presence of the anti-Fas neutralizing antibody ZB-4. Although ectopic expression of Bcl-2 failed to completely block the cytotoxicity of aburatubolactam C, it was clearly suppressed. The c-Fos mRNA expression was upregulated in a biphasic manner, where the second phasic expression overlapped with the FasL upregulation. Accordingly, these results demonstrate that aburatubolactam C-induced apoptosis is exerted, at least in part, by FasL upregulation dictated by activation of the PTK (p56^lck and p59^fyn)/JNK1 pathway, which is negatively affected by the concurrent activation of the PKC/ERK2 pathway proximal to PTK activation.