The coordinate release of cytochrome c during apoptosis is rapid, complete and kinetically invariant.
Release of cytochrome c from mitochondria triggers activation of caspase proteases and death of a cell by apoptosis. However, the mechanism and kinetics of cytochrome c release remain unknown. Here we study this event by using green fluorescent protein (GFP)-tagged cytochrome c, and find that the release of cytochrome-c-GFP always precedes exposure of phosphatidylserine and 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 about 5 minutes, regardless of the type or strength of stimulus or the time elapsed 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 addition of caspase inhibitors. Further, we find that the electron-transport chain can maintain the mitochondrial transmembrane potential even after cytochrome c has been released.
Goldstein
J C
JC
Division of Cellular Immunology, La Jolla Institute for Allergy and Immunology, 10355 Science Center Drive, San Diego, California 92121, USA.
Waterhouse
N J
NJ
Juin
P
P
Evan
G I
GI
Green
D R
DR
D000255:Adenosine Triphosphate / Q000378:metabolism; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D017209:Apoptosis; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003574:Cytochrome c Group / Q000235:genetics / Q000378:metabolism*; D004072:Digitonin / Q000494:pharmacology; D004579:Electron Transport / Q000187:drug effects / Q000528:radiation effects; D004789:Enzyme Activation / Q000187:drug effects / Q000528:radiation effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D007091:Image Processing, Computer-Assisted; D007425:Intracellular Membranes / Q000187:drug effects / Q000528:radiation effects; D008164:Luminescent Proteins / Q000235:genetics; D008564:Membrane Potentials / Q000187:drug effects; D008928:Mitochondria / Q000378:metabolism; D009840:Oligomycins / Q000494:pharmacology; D010718:Phosphatidylserines / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D019810:Sodium Azide / Q000494:pharmacology; D013696:Temperature; D014466:Ultraviolet Rays
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D003574:Cytochrome c Group; D004791:Enzyme Inhibitors; D008164:Luminescent Proteins; D009840:Oligomycins; D010718:Phosphatidylserines; D011993:Recombinant Fusion Proteins; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D049452:Green Fluorescent Proteins; D000255:Adenosine Triphosphate; D019810:Sodium Azide; D020169:Caspases; D004072:Digitonin