c-Jun NH2-terminal kinase-dependent Fas activation contributes to etoposide-induced apoptosis in p53-mutated prostate cancer cells.
BACKGROUND
The death receptor, Fas, has recently been demonstrated to contribute the chemotherapeutic agents-induced apoptosis, however, the detail mechanisms have yet to be fully understood, especially in prostate cancer cells.


METHODS
PC-3 and DU145 stably transfected with dominant negative form of Fas-associated death domain (FADD) or specific kinase of c-Jun NH2-terminal kinase (JNK) (mitogen-activated protein kinase kinase, MKK7) were selected in the presence of hygromycin B (Hyg B). Cell viability was examined by (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulphonyl)-2H-tetrazolium, inner salt (MTS) assay or flowcytometric analysis using green fluorescent protein (GFP). Apoptosis was examined by DNA ladder, Western blotting analysis of cleaved caspases, or morphological analysis. The expression of Fas and JNK activation were investigated by Western blotting/flowcytometric analysis and in vitro kinase assay, respectively.


RESULTS
Stimulation with etoposide significantly up-regulated Fas, and the death-inducing signaling complex (DISC) was formed in PC-3 and DU145. Stable transfection with dominant-negative FADD inhibited etoposide-induced apoptosis. In addition, stable transfection with dominant-negative MKK7, by which JNK activation was inhibited, canceled both the up-regulation of Fas and the formation of DISC by etoposide. Re-introduction of wild type p53 into PC-3 and DU145 completely suppressed these inhibitory effects.


CONCLUSIONS
These results suggest that, in p53-mutated prostate cancer, JNK-initiated Fas-mediated apoptotic signals may play an important role in chemosensitivity.
Shimada
Keiji
K
Second Department of Pathology, Nara Medical University, Kashihara, Nara, Japan.
Nakamura
Mitsutoshi
M
Ishida
Eiwa
E
Kishi
Munehiro
M
Yonehara
Shin
S
Konishi
Noboru
N
D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D029681:Arabidopsis Proteins; D015153:Blotting, Western; D020169:Caspases / Q000378:metabolism; D053938:DNA Fragmentation / Q000502:physiology; D005047:Etoposide / Q000494:pharmacology*; D044943:Fatty Acid Desaturases / Q000235:genetics / Q000378:metabolism; D005434:Flow Cytometry; D016158:Genes, p53 / Q000235:genetics*; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases; D048688:MAP Kinase Kinase 7; D008297:Male; D020929:Mitogen-Activated Protein Kinase Kinases / Q000235:genetics / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009154:Mutation; D011471:Prostatic Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D011506:Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured; D019014:fas Receptor / Q000235:genetics / Q000378:metabolism*
D000972:Antineoplastic Agents, Phytogenic; D029681:Arabidopsis Proteins; D011506:Proteins; D019014:fas Receptor; D005047:Etoposide; D044943:Fatty Acid Desaturases; C084083:Fad7 protein, Arabidopsis; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048688:MAP Kinase Kinase 7; C482569:MAP2K7 protein, human; D020929:Mitogen-Activated Protein Kinase Kinases; D020169:Caspases