1. ¹Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria 3800, Australia
2. ²ARC Centre of Excellence in Structural and Functional Microbial Genomics, Monash University, Clayton, Victoria 3800, Australia

Correspondence: P Nagley, Department of Biochemistry and Molecular Biology, Building 13D, Monash University, Victoria 3800, Australia. Tel: +61-3-9905-3735; Fax +61-3-9905-3726; E-mail: phillip.nagley@med.monash.edu.au

Received 17 December 2004; Revised 19 August 2005; Accepted 15 September 2005; Published online 21 October 2005.

Abstract

We have characterised the apoptotic defects in cells null for cytochrome c (cyt c-/-). Such cells treated with staurosporine (STS) exhibited translocation to the mitochondria and activation of the proapoptotic signalling molecule Bax but failed to release Smac/DIABLO from these organelles, judged by both confocal microscopy and Western blotting. While reference cells expressing cytochrome c released both it and Smac/DIABLO under a variety of conditions of apoptotic induction, we have never observed release of Smac/DIABLO from cyt c-/- cells. We eliminate the possibility that proteasomal degradation of cytosolically localised Smac/DIABLO is responsible for our failure to visualise the protein outside the mitochondria. Our findings indicate an unanticipated nexus between release of cytochrome c and Smac/DIABLO from mitochondria, previously thought to be a more or less synchronised event early in apoptosis. We suggest that the failure of cyt c-/- cells to release Smac/DIABLO after recruitment of Bax to mitochondria represents an extreme manifestation of some inherent difference in the regulation of release of these two proteins from mitochondria.