The coordinate release of cytochrome c during apoptosis is rapid, completeand kinetically invariant

Release of cytochrome c from mitochondria triggers activation of caspase pr oteases and death of a cell by apoptosis, However, the mechanism and kineti cs of cytochrome c release remain unknown. Here we study this event by usin g green fluorescent protein (GFP)-tagged cytochrome c, and find that the re lease of cytochrome-c-GFP always precedes exposure of phosphatidylserine an d the loss of plasma-membrane integrity-characteristics of apoptotic cells. Once initiated, the release of cytochrome-c-GFP continues until all of the protein is released from all mitochondria in individual cells, within abou t 5 minutes, regardless of the type or strength of stimulus or the time ela psed since the stimulus was applied, Temperatures ranging from 24 degrees C to 37 degrees C do not change the duration of release, and nor does the ad dition of caspase inhibitors. Further, we find that the electron-transport chain can maintain the mitochondrial transmembrane potential even after cyt ochrome c has been released.