Cytochrome c is released in a single step during apoptosis

J. C. Goldstein, C. Muñoz-Pinedo, J. E. Ricci, S. R. Adams, A. Kelekar, M. Schuler, R. Y. Tsien, Doug R. Green

Research output: Contribution to journalArticlepeer-review

165 Scopus citations


Release of cytochrome c from mitochondria is a central event in apoptotic signaling. In this study, we utilized a cytochrome c fusion that binds fluorescent biarsenical ligands (cytochrome c-4CYS (cyt. c-4CYS)) as well as cytochrome c-green fluorescent protein (cyt. c-GFP) to measure its release from mitochondria in different cell types during apoptosis. In single cells, the kinetics of cyt. c-4CYS release was indistinguishable from that of cyt. c-GFP in apoptotic cells expressing both molecules. Lowering the temperature by 7°C did not affect this corelease, but further separated cytochrome c release from the subsequent decrease in mitochondrial membrane potential (ΔΨm). Cyt. c-GFP rescued respiration in cells lacking endogenous cytochrome c, and the duration of cytochrome c release was approximately 5 min in a variety of cell types induced to die by various forms of cellular stress. In addition, we could observe no evidence of caspase-dependent amplification of cytochrome c release or changes in ΔΨm preceding the release of cyt. c-GFP. We conclude that there is a general mechanism responsible for cytochrome c release that proceeds in a single step that is independent of changes in ΔΨm.

Original languageEnglish (US)
Pages (from-to)453-462
Number of pages10
JournalCell Death and Differentiation
Issue number5
StatePublished - May 2005

Bibliographical note

Funding Information:
This work was supported by NIH Grants AI40646, AI47891, GM52735, and CA69381. C Muñoz-Pinedo is a recipient of a fellowship from the Secretaría de Estado de Educación y Universidades of Spain. We thank G Gaietta, TJ Durnick, DL Price, and MH Ellisman for helpful imaging discussions, AL Nieminen for the gift of the rat cyt. c-GFP construct, T Kanda and GM Wahl for the gift of the H2B-GFP construct, P Fitzgerald and E Ehlert for cloning advice, and CJ Lena for cell sorting.

Copyright 2008 Elsevier B.V., All rights reserved.


  • Apoptosis
  • Cytochrome c
  • Mitochondria
  • Mitochondrial membrane potential
  • Video microscopy


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