Activation of c-Jun NH2-terminal kinase/stress-activated protein kinase (JNK/SAPK) is critical for hypoxia-induced apoptosis of human malignant melanoma

Mitogen-activated protein kinase (MAPK) signaling was examined in malignant melanoma cells exposed to hypoxia, Here we demonstrate that hypoxia induce d a strong activation of the c-Jun NH2-terminal kinase (JNK), also termed s tress-activated protein kinase (SAPK), in the melanoma cell line 530 in vit ro. Other members of the MAPK family, e.g,, extracellular signal-regulated kinase and p38, remained unaffected by the hypoxic stimulus. Activated JNK/ SAPK could also be observed in the vicinity of hypoxic tumor areas in melan oma metastases as detected by immunohistochemistry. Functional analysis of JNK/SAPK activation in the melanoma cell line 530 revealed that activation of JNK/SAPK is involved in hypoxia-mediated tumor cell apoptosis, Both a do minant negative mutant of JNWSAPK (SAPK beta K -->R) and a dominant negativ e mutant of the immediate upstream activator of JNK/SAPK, SEK1 (SEK1 K -->R ), inhibited hypoxia-induced apoptosis in transient transfection studies, I n contrast, overexpression of the wild-type kinases had a slight proapoptot ic effect. Inhibition of extracellular signal-regulated kinase and p38 path ways by the chemical inhibitors PD98058 and SB203580, respectively, had no effect on hypoxia-induced apoptosis, Under normoxic conditions, no influenc e on apoptosis regulation was observed after inhibition of all three MAPK p athways. In contrast to recent findings, JNK/SAPK activation did not correl ate with Fas or Fas ligand (FasL) expression, suggesting that the Fas/FasL system is not involved in hypoxia-induced apoptosis in melanoma cells. Take n together, our data demonstrate that hypoxia-induced JNK/SAPK activation a ppears to play a critical role in apoptosis regulation of melanoma cells in vitro and in vivo, independent of the Fas/FasL system.