Mitochondrial nitric-oxide synthase stimulation causes cytochrome c release from isolated mitochondria. Evidence for intramitochondrial peroxynitrite formation.
Nitric oxide (NO) is synthesized by members of the NO synthase (NOS) family. Recently the existence of a mitochondrial NOS (mtNOS), its Ca(2+) dependence, and its relevance for mitochondrial bioenergetics was reported (Ghafourifar, P., and Richter, C. (1997) FEBS Lett. 418, 291-296; Giulivi, C., Poderoso, J. J., and Boveris, A. (1998) J. Biol. Chem. 273, 11038-11043). Here we report on the possible involvement of mtNOS in apoptosis. We show that uptake of Ca(2+) by mitochondria triggers mtNOS activity and causes the release of cytochrome c from isolated mitochondria in a Bcl-2-sensitive manner. mtNOS-induced cytochrome c release was paralleled by increased lipid peroxidation. The release of cytochrome c as well as increase in lipid peroxidation were prevented by NOS inhibitors, a superoxide dismutase mimic, and a peroxynitrite scavenger. We show that mtNOS-induced cytochrome c release is not mediated via the mitochondrial permeability transition pore because the release was aggravated by cyclosporin A and abolished by blockade of mitochondrial calcium uptake by ruthenium red. We conclude that, upon Ca(2+)-induced mtNOS activation, peroxynitrite is formed within mitochondria, which causes the release of cytochrome c from isolated mitochondria, and we propose a mechanism by which elevated Ca(2+) levels induce apoptosis.
Ghafourifar
P
P
Institute of Biochemistry, Swiss Federal Institute of Technology (ETH), CH-8092 Zurich, Switzerland. pedramgf@bc.biol.ethz.ch
Schenk
U
U
Klein
S D
SD
Richter
C
C
D000818:Animals; D017209:Apoptosis; D002118:Calcium / Q000494:pharmacology; D016572:Cyclosporine / Q000494:pharmacology; D003574:Cytochrome c Group / Q000378:metabolism*; D004789:Enzyme Activation; D007473:Ion Channels; D015227:Lipid Peroxidation; D008564:Membrane Potentials; D008565:Membrane Proteins / Q000378:metabolism; D008930:Mitochondria, Liver / Q000201:enzymology*; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D009566:Nitrates / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D016877:Oxidants / Q000378:metabolism*; D010101:Oxygen Consumption; D019253:Proto-Oncogene Proteins c-bcl-2; D051381:Rats; D012430:Ruthenium Red / Q000494:pharmacology
D003574:Cytochrome c Group; D007473:Ion Channels; D008565:Membrane Proteins; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D009566:Nitrates; D016877:Oxidants; D019253:Proto-Oncogene Proteins c-bcl-2; D012430:Ruthenium Red; C039642:peroxynitric acid; D016572:Cyclosporine; D019001:Nitric Oxide Synthase; D002118:Calcium