Mixed lineage kinase 3 connects reactive oxygen species to c-Jun NH2-terminal kinase-induced mitochondrial apoptosis in genipin-treated PC3 human prostate cancer cells.
It has been reported that genipin, the aglycone of geniposide, induces apoptotic cell death in human hepatoma cells via a NADPH oxidase-reactive oxygen species (ROS)-c-Jun NH(2)-terminal kinase (JNK)-dependent activation of mitochondrial pathway. This continuing work aimed to define that mixed lineage kinase 3 (MLK3) is a key mediator, which connect between ROS and JNK in genipin-induced cell death signaling. In PC3 human prostate cancer cells, genipin stimulated MLK3 activity in concentration- and time-dependent manner. The PC3 cells stably transfected with dominant-negative form of MLK3 was less susceptible to population of the sub-G1 apoptotic cells, activation of caspase, collapse of mitochondrial membrane potential, and release of cytochrome c triggered by genipin, suggesting a crucial role of MLK3 in genipin signaling to apoptotic cell death. Diphenyleneiodonium (DPI), a specific inhibitor of NADPH oxidase, markedly inhibited ROS generation and MLK3 phosphorylation in the genipin-treated cells. Pretreatment with SP0600125, a specific inhibitor of JNK but neither U0126, a specific inhibitor of MEK1/2 nor PD169316, a specific inhibitor of p38 suppressed genipin-induced apoptotic cell death. Notably, both the phosphorylation of JNK and induction of c-Jun induced by genipin were markedly inhibited in PC3-EGFP-MLK3 (K144R) cells expressing a dominant-negative MLK3 mutant. Taken together, our observations suggest genipin signaling to apoptosis of PC3 cells is mediated via activation of ROS-dependent MLK3, which leads to downstream activation of JNK.
Hong
Hye-Young
HY
Division of Life Sciences, Kangwon National University, Chuncheon 200-701, Republic of Korea.
Kim
Byung-Chul
BC
D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002070:Butadienes / Q000494:pharmacology; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005434:Flow Cytometry; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D057889:Iridoid Glycosides; D039823:Iridoids / Q000494:pharmacology*; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism*; D020930:MAP Kinase Kinase Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D008297:Male; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D009570:Nitriles / Q000494:pharmacology; D011471:Prostatic Neoplasms / Q000378:metabolism / Q000473:pathology; D017382:Reactive Oxygen Species / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D014162:Transfection; D000097767:Mitogen-Activated Protein Kinase Kinase Kinase 11
D002070:Butadienes; D004791:Enzyme Inhibitors; D049452:Green Fluorescent Proteins; D007093:Imidazoles; D057889:Iridoid Glycosides; D039823:Iridoids; D048031:JNK Mitogen-Activated Protein Kinases; D020930:MAP Kinase Kinase Kinases; D009570:Nitriles; D017382:Reactive Oxygen Species; D011993:Recombinant Fusion Proteins; D000097767:Mitogen-Activated Protein Kinase Kinase Kinase 11; C113580:U 0126; C007834:genipin; C408604:2-(4-nitrophenyl)-4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-imidazole