Increased mitochondrial cytochrome c levels and mitochondrial hyperpolarization precede camptothecin-induced apoptosis in Jurkat cells.
Mitochondria play a central role in apoptosis through release of cytochrome c and activation of caspases. In the present study, we showed that, in Jurkat human T cells, camptothecin-induced apoptosis is preceded by (i) an increase in cytochrome c and subunit IV of cytochrome c oxidase (COX IV) levels in mitochondria; and (ii) an elevation of the mitochondrial membrane potential (Delta(Psi)m). These events are followed by cytochrome c release into the cytosol, cytochrome c and COX IV depletion from mitochondria, externalization of phosphatidylserine (PS), disruption of Delta(Psi)m, caspase activation, poly(ADP-ribose)polymerase cleavage and DNA fragmentation. The pan-caspase inhibitor z-VAD.fmk blocked camptothecin-induced PS externalization, disruption of Delta(Psi)m and DNA fragmentation, suggesting that these events are mediated by caspase activation. In contrast, z-VAD did not prevent cytochrome c release, despite preventing cytochrome c and COX IV depletion from mitochondria. Together, these data suggest that mitochondrial cytochrome c and COX IV enrichment are early events preceding the onset of apoptosis and that cytochrome c release is upstream of caspase activation and loss of Delta(Psi)m. Furthermore, prevention by z-VAD of cytochrome c and COX IV depletion in mitochondria suggests the possibility that a caspase-like activity in mitochondria is involved in the proteolytic depletion of respiratory chain proteins. Activation of this activity may play an important role in drug-induced apoptosis.
Sánchez-Alcázar
J A
JA
Wadsworth Center, New York State Department of Health, Empire State Plaza, Albany, NY 12201, USA.
Ault
J G
JG
Khodjakov
A
A
Schneider
E
E
D000590:Amino Acid Chloromethyl Ketones / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D053477:Apoptotic Protease-Activating Factor 1; D015153:Blotting, Western; D002166:Camptothecin / Q000494:pharmacology*; D020169:Caspases / Q000378:metabolism; D003574:Cytochrome c Group / Q000378:metabolism*; D003576:Electron Transport Complex IV / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D019169:Jurkat Cells; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008928:Mitochondria / Q000201:enzymology* / Q000235:genetics / Q000502:physiology / Q000648:ultrastructure; D011506:Proteins / Q000378:metabolism; D013997:Time Factors
C506592:APAF1 protein, human; D000590:Amino Acid Chloromethyl Ketones; D053477:Apoptotic Protease-Activating Factor 1; D003574:Cytochrome c Group; D004791:Enzyme Inhibitors; D011506:Proteins; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D003576:Electron Transport Complex IV; D020169:Caspases; D002166:Camptothecin