The role of endogenous and exogenous RasGAP-derived fragment N in protecting cardiomyocytes from peroxynitrite-induced apoptosis.
Peroxynitrite (PN) is a potent nitrating and oxidizing agent generated during various pathological situations affecting the heart. The negative effects of PN result, at least in part, from its ability to activate caspases and apoptosis. RasGAP is a ubiquitously expressed protein that is cleaved sequentially by caspase-3. At low caspase-3 activity, RasGAP is cleaved into an N-terminal fragment, called fragment N, that protects cells by activating the Ras/PI3K/Akt pathway. At high caspase-3 activity, fragment N is further cleaved and this abrogates its capacity to stimulate the antiapoptotic Akt kinase. Fragment N formation is crucial for the survival of cells exposed to a variety of stresses. Here we investigate the pattern of RasGAP cleavage upon PN stimulation and the capacity of fragment N to protect cardiomyocytes. PN did not lead to sequential cleavage of RasGAP. Indeed, PN did not allow accumulation of fragment N because it induced its rapid cleavage into smaller fragments. No situations were found in cells treated with PN in which the presence of fragment N was associated with survival. However, expression of a caspase-resistant form of fragment N in cardiomyocytes protected them from PN-induced apoptosis. Our results indicate that the antiapoptotic pathway activated by fragment N is effective at inhibiting PN-induced apoptosis (as seen when cardiomyocytes express a capase-3-resistant form of fragment N) but because fragment N is too transiently generated in response to PN, no survival response is effectively produced. This may explain the marked deleterious consequences of PN generation in various organs, including the heart.
Department of Physiology, Biology and Medicine Faculty, University of Lausanne, 1005 Lausanne, Switzerland.
Hadi
H
Khalil
Nathalie
N
Rosenblatt
Lucas
L
Liaudet
Christian
C
Widmann
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3 / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival; D002945:Cisplatin / Q000494:pharmacology; D015870:Gene Expression; D006801:Humans; D051379:Mice; D009153:Mutagens / Q000494:pharmacology; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D016877:Oxidants / Q000494:pharmacology*; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D030421:Peroxynitrous Acid / Q000494:pharmacology*; D059748:Proteolysis; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D051381:Rats; D020703:ras GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology
D009153:Mutagens; D016877:Oxidants; D010446:Peptide Fragments; D020703:ras GTPase-Activating Proteins; D030421:Peroxynitrous Acid; D051057:Proto-Oncogene Proteins c-akt; D053148:Caspase 3; D002945:Cisplatin