Ca2+-induced reactive oxygen species production promotes cytochrome c release from rat liver mitochondria via mitochondrial permeability transition (MPT)-dependent and MPT-independent mechanisms: role of cardiolipin.
Release of cytochrome c from mitochondria is considered a critical, early event in the induction of an apoptosis cascade that ultimately leads to programmed cell death. Mitochondrial Ca(2+) loading is a trigger for the release of cytochrome c, although the molecular mechanism underlying this effect is not fully clarified. This study tested the hypothesis that distinct Ca(2+) thresholds may induce cytochrome c release from rat liver mitochondria by membrane permeability transition (MPT)-dependent and independent mechanisms. The involvement of reactive oxygen species (ROS) and cardiolipin in the Ca(2+)-induced cytochrome c release was also investigated. Cytochrome c was quantitated by a new, very sensitive, and rapid reverse-phase high performance liquid chromatography method with a detection limit of 0.1 pmol/sample. We found that a low extramitochondrial Ca(2+) level (2 microM) promoted the release of approximately 13% of the total alamethicin releasable pool of cytochrome c from mitochondria. This release was not depending of MPT; it was mediated by Ca(2+)-induced ROS production and cardiolipin peroxidation and appears to involve the voltage-dependent anion channel. High extramitochondrial Ca(2+) level (20 microM) promoted approximately 45% of the total releasable pool of cytochrome c. This process was MPT-dependent and was also mediated by ROS and cardiolipin. It is suggested that distinct Ca(2+) levels may determine the mode and the amount of cytochrome c release from rat liver mitochondria. The data may help to clarify the molecular mechanism underlying the Ca(2+)-induced release of cytochrome c from rat liver mitochondria and the role played by ROS and cardiolipin in this process.
Department of Biochemistry and Molecular Biology and Consiglio Nazionale delle Ricerche Institute of Biomembranes and Bioenergetics, University of Bari, Via E. Orabona, 4, 70126 Bari, Italy.
Giuseppe
G
Petrosillo
Francesca M
FM
Ruggiero
Marilva
M
Pistolese
Giuseppe
G
Paradies
D000408:Alamethicin / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D002118:Calcium / Q000378:metabolism*; D002308:Cardiolipins / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D016572:Cyclosporine / Q000494:pharmacology; D045304:Cytochromes c / Q000378:metabolism*; D007473:Ion Channels / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D008930:Mitochondria, Liver / Q000378:metabolism*; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D051381:Rats; D017382:Reactive Oxygen Species; D012430:Ruthenium Red / Q000494:pharmacology; D013050:Spectrometry, Fluorescence; D013997:Time Factors
D002308:Cardiolipins; D007473:Ion Channels; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D017382:Reactive Oxygen Species; D012430:Ruthenium Red; D000408:Alamethicin; D016572:Cyclosporine; D045304:Cytochromes c; D002118:Calcium