p53 activates the mitochondrial death pathway and apoptosis of ventricular myocytes independent of de novo gene transcription.
The tumor suppressor p53 is known to regulate gene transcription and apoptosis in mammalian cells. In the present study we ascertain whether these events are mutually dependent and obligatorily linked for induction of apoptosis of ventricular myocytes. Adenovirus mediated gene delivery of wild p53 (p53WT) or a mutant form of p53 (p53MT) defective for gene transcription to ventricular myocytes was confirmed by Western blot analysis. A significant increase in the p53 dependent genes Bax and MDM2 was observed with p53WT but not p53MT. Nuclear DNA visualized by agarose gel electrophoresis revealed nucleosomal DNA laddering in the presence of either p53 protein. Apoptosis was substantiated by Hoechst 33258 nuclear staining. Perturbations to mitochondria consistent with the mitochondrial death pathway, including loss of mitochondrial transmembrane potential Delta(psi)m and cytochrome c release were observed with p53WT and p53MT. An increase in caspase 3-like activity was noted with either p53WT or p53MT protein that was suppressed by the caspase 3 inhibitor Ac-DEVD-CHO. To our knowledge the experiments described here provide the first indication that p53 activates the mitochondrial death pathway and provokes apoptosis of ventricular myocytes independent of DNA binding and de novo gene activation.
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Institute of Cardiovascular Sciences, St Boniface General Hospital Research Centre, Winnipeg, Manitoba, R2H 2A6, Canada.
Kirshenbaum
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D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003574:Cytochrome c Group / Q000378:metabolism; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D016158:Genes, p53 / Q000502:physiology*; D005822:Genetic Vectors; D006352:Heart Ventricles / Q000166:cytology; D006801:Humans; D066298:In Vitro Techniques; D008929:Mitochondria, Heart / Q000473:pathology*; D009206:Myocardium / Q000473:pathology*; D009842:Oligopeptides / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000235:genetics; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection
D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D003574:Cytochrome c Group; D009842:Oligopeptides; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; C099158:acetyl-aspartyl-glutamyl-valyl-aspartal