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Mol. Cell. Biol. doi:10.1128/MCB.00287-06
Copyright (c) 2006, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

The Role of Cytochrome c in Apoptosis: Increased Sensitivity to TNF- is Associated with Respiratory Defects But Not with Lack of Cytochrome c Release

Departments of Neurology, Microbiology and Immunology, Medical Oncology and Cell Biology and Anatomy, University of Miami Miller School of Medicine, Miami, FL; The Burnham Institute, La Jolla Cancer Research Center, La Jolla, CA

^* To whom correspondence should be addressed. Email: cmoraes{at}med.miami.edu.

	Abstract

Although the role of cytochrome c in apoptosis is well established, details of its participation in signaling pathways in vivo are not completely understood. The knockout for the somatic isoform of cytochrome c caused embryonic lethality in mice, but derived embryonic fibroblasts were shown to be resistant to apoptosis induced by agents known to trigger the intrinsic apoptotic pathway. In contrast, these cells were reported to be hypersensitive to TNF- induced apoptosis, which signals through the extrinsic pathway. Surprisingly, we found that this cell line (CRL 2613) respired at close to normal levels because of an aberrant activation of a testis isoform of cytochrome c, which albeit expressed at low levels, was able to replace the somatic isoform for respiration and apoptosis. To produce a bona fide cytochrome c knockout, we developed a mouse knockout for both the testis and somatic isoforms of cytochrome c. The mouse was made viable by the introduction of a ubiquitously expressed cytochrome c transgene flanked by loxP sites. Lung fibroblasts in which the transgene was deleted showed no cytochrome c expression, no respiration and resistance to agents that activate the intrinsic, and to a lesser but significant extent, also the extrinsic pathways. Comparison of these cells with lines with a defective oxidative phosphorylation system, showed that cells with defective respiration have increased sensitivity to TNF- induced apoptosis, but this process was still amplified by cytochrome c. These studies underscore the importance of oxidative phosphorylation and apoptosome function to both the intrinsic and extrinsic apoptotic pathways.