Cytochrome c translocation does not lead to caspase activation in maitotoxin-treated SH-SY5Y neuroblastoma cells.
Cytosolic cytochrome c elevation has been associated with activation of caspase-3-like proteases. In this study, we demonstrate that treatment with the neurotoxin and potent calcium channel opener maitotoxin (MTX) induces cytochrome c release from the mitochondria that is not accompanied by caspase activation. Cytochrome c translocation in MTX-treated SH-SY5Y cells was readily apparent after 30 min and peaked at 2.5h. We assayed caspase activity by acetyl-Asp-Glu-Val-Asp-7-amido-4-methylcoumarin (Ac-DEVD-AMC) hydrolysis and by immunoblotting for caspase-3 processing and proteolysis of alphaII-spectrin and PARP. In contrast, treatment with pro-apoptosis agent staurosporine (STS) induced both cytochrome c release and caspase-3 activation after 2h. In addition, with MTX treatment, we found no evidence of caspase activation at any time point or MTX concentration used. Instead, we observed that caspase-9, Apaf-1 and caspase-3 were all partially truncated by calpain under these conditions. These combined effects likely contribute to the lack of caspase activation cascade in MTX-treated cells, despite the presence of cytochrome c in the cytosol.
McGinnis
Kim M
KM
Department of Pharmacology, The University of Michigan Medical School, Ann Arbor, MI 48109, USA.
Gnegy
Margaret E
ME
Falk
Nicole
N
Nath
Rathna
R
Wang
Kevin K W
KK
D020169:Caspases / Q000378:metabolism*; D003574:Cytochrome c Group / Q000378:metabolism*; D004789:Enzyme Activation; D006801:Humans; D008387:Marine Toxins / Q000494:pharmacology*; D009447:Neuroblastoma / Q000201:enzymology* / Q000473:pathology; D035964:Oxocins / Q000494:pharmacology*; D021381:Protein Transport; D014407:Tumor Cells, Cultured; D000095702:Polyether Toxins
D020169:Caspases; D003574:Cytochrome c Group; D008387:Marine Toxins; D035964:Oxocins; C020394:maitotoxin; D000095702:Polyether Toxins