A cytotoxin isolated from Agkistrodon acutus snake venom induces apoptosis via Fas pathway in A549 cells.
ACTX-6 is a protein isolated from Agkistrodon acutus snake venom and demonstrated cytotoxic activity to various cancer cells in vitro. In this paper the exact mechanism in ACTX-6-induced cell death was investigated and it was found that ACTX-6 could induce cell apoptosis. The results of Western blot and RT-PCR showed that ACTX-6 could induce Fas and FasL protein expression. When Fas signaling pathway was blocked by neutralizing antibodies to Fas or FasL, ACTX-6-induced apoptosis was inhibited. DISC formation was also detected by immunoprecipitation. These results suggested that Fas pathway was involved in ACTX-6-induced apoptosis. The activities of caspase-3, 8 and 9 were assayed and the activation of caspase-9 demonstrated that mitochondrial pathway was also involved in ACTX-6-induced apoptosis. Bid cleavage and dissipation of mitochondrial membrane potential (delta psi(m)) verified the involvement of mitochondria. ACTX-6 is an L-amino acid oxidase and can oxidize L-amino acid to generate hydrogen peroxide. The production of ROS in ACTX-6-treated cells was detected and the ROS scavenger catalase could inhibit ACTX-6-induced apoptosis. Western blot analysis showed that JNK was phosphorylated in ACTX-6-treated cells and c-Jun was also activated. JNK inhibitor SP600125 could inhibit ACTX-6-induced apoptosis and catalase could inhibit JNK and c-Jun phosphorylation. It could be concluded that JNK pathway was necessary in ACTX-6-induced apoptosis and the oxidative stress generated by ACTX-6 was responsible for the activation of JNK.
Zhang
Liang
L
School of Pharmacy, Soochow University, Renai Road, Soochow, Jiangsu Province 215123, China. cpuzl@163.com
Cui
Li
L
D017836:Agkistrodon; D000818:Animals; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D003435:Crotalid Venoms / Q000633:toxicity*; D003603:Cytotoxins / Q000633:toxicity*; D053222:Fas Ligand Protein / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D048670:MAP Kinase Kinase 4 / Q000378:metabolism; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D012333:RNA, Messenger / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D019014:fas Receptor / Q000235:genetics / Q000378:metabolism*
C524843:ACTX-6 protein, Agkistrodon acutus; D003435:Crotalid Venoms; D003603:Cytotoxins; D053222:Fas Ligand Protein; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D019014:fas Receptor; D048031:JNK Mitogen-Activated Protein Kinases; D048670:MAP Kinase Kinase 4; D020169:Caspases