title: Protein synthesis-dependent but Bcl-2-independent cytochrome C release in zinc depletion-induced neuronal apoptosis.
abstract: Previously, we reported that chelation of intracellular zinc with N, N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN)-induced macromolecule synthesis-dependent apoptosis of cultured cortical neurons. According to the current theory of apoptosis, release of mitochondrial cytochrome C into the cytosol is required for caspase activation. In the present study, we examined whether cytochrome C release is dependent on macromolecule synthesis. Exposure of cortical cultures to 2 microM TPEN for 24 hr induced apoptosis as previously described. Fluorescence immunocytochemical staining as well as immunoblots of cell extracts revealed the release of cytochrome C into the cytosol 18-20 hr after the exposure onset. The cytochrome C release was completely blocked by the addition of cycloheximide or actinomycin D. Addition of the caspase inhibitor zVAD-fmk did not attenuate the cytochrome C release, whereas it blocked TPEN-induced apoptosis. Because Bcl-2 has been shown to block cytochrome C release potently, we exposed human neuroblastoma cells (SH-SY5Y) to TPEN. Whereas Bcl-2 overexpression completely blocked both cytochrome C release and apoptosis induced by staurosporine, it attenuated neither induced by TPEN. The present results suggest that, in neurons, macromolecule synthesis inhibitors act upstream of cytochrome C release to block apoptosis and that, in addition to the classical Bcl-2 sensitive pathway, there may exist a Bcl-2-insensitive pathway for cytochrome C release.
authors.affiliation: School of Biological Sciences and Institute for Molecular Biology and Genetics, Seoul National University, Seoul, Korea.
authors.forename: Y H
authors.initials: YH
authors.lastname: Ahn
authors.forename: J Y
authors.initials: JY
authors.lastname: Koh
authors.forename: S H
authors.initials: SH
authors.lastname: Hong
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D002800:Cholinesterase Inhibitors / Q000494:pharmacology; D003574:Cytochrome c Group / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005029:Ethylenediamines / Q000494:pharmacology; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D006801:Humans; D051379:Mice; D008928:Mitochondria / Q000378:metabolism; D016202:N-Methylaspartate / Q000494:pharmacology; D009447:Neuroblastoma / Q000378:metabolism / Q000473:pathology; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D014176:Protein Biosynthesis; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D015032:Zinc / Q000172:deficiency / Q000378:metabolism*
chemical_list: D002800:Cholinesterase Inhibitors; D003574:Cytochrome c Group; D004791:Enzyme Inhibitors; D005029:Ethylenediamines; D018690:Excitatory Amino Acid Agonists; D011500:Protein Synthesis Inhibitors; D019253:Proto-Oncogene Proteins c-bcl-2; D016202:N-Methylaspartate; D015032:Zinc; C044387:N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine