Suicidal differential housekeeping gene activity in apoptosis induced by DCNP.
Previous suggestions of CpG-specific apoptotic commitment implied critical epigenetic modulation of housekeeping genes which have canonical CpG islands at 5' promoter regions. Differential housekeeping gene activity however has not been shown. Using a focussed microarray (genechip) of 22 housekeeping genes we show this in apoptosis induced in human Chang liver cells by DCNP (2,6-dichloro-4-nitrophenol), a non-genotoxic inhibitor of sulfate detoxification. 3-7 folds downregulation of 9 genes in glycolysis, tricarboxylic acid cycle and the respiratory electron transport chain suggested gene-directed energy depletion which was correlated with observed ATP depletion. 4 folds downregulation of the pyruvate dehydrogenease gene suggested gene-directed metabolic acidosis which was correlated with observed cell acidification. Other differential housekeeping gene activity, including 4 folds upregulation of microtubular alpha-tubulin gene, and 2 folds upregulation of ubiquitin, also had a bearing on apoptosis. Broadspectrum zVAD-fmk caspase inhibition abolished 200 bp DNA ladder fragmentations but not the CpG-specific megabase fragmentations and other hallmarks of cell destruction, suggesting a caspase-independent cell death. Death appeared committed at gene-level.
Department of Anatomy, Faculty of Medicine, National University of Singapore, Kent Ridge.
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D000255:Adenosine Triphosphate / Q000378:metabolism; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D053938:DNA Fragmentation; D004305:Dose-Response Relationship, Drug; D004734:Energy Metabolism / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D004789:Enzyme Activation; D005434:Flow Cytometry; D005786:Gene Expression Regulation / Q000187:drug effects*; D022781:Hepatocytes / Q000166:cytology / Q000187:drug effects*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008928:Mitochondria / Q000378:metabolism; D009596:Nitrophenols / Q000494:pharmacology*; D020411:Oligonucleotide Array Sequence Analysis
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D009596:Nitrophenols; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C015802:2,6-dichloro-4-nitrophenol; D000255:Adenosine Triphosphate; D020169:Caspases