A cytosolic factor is required for mitochondrial cytochrome c efflux during apoptosis.
Treatment of HL-60 cells with staurosporine (STS) induced mitochondrial cytochrome c efflux into the cytosol, which was followed by caspase-3 activation and apoptosis. Consistent with these observations, in vitro experiments demonstrated that, except for cytochrome c, the cytosol of HL-60 cells contained sufficient amounts of all factors required for caspase-3 activation. In contrast, treatment of HCW-2 cells (an apoptotic-resistant HL-60 subclone) with STS failed to induce significant amounts of mitochondrial cytochrome c efflux, caspase-3 activation, and apoptosis. In vitro assays strongly suggested that a lack of cytochrome c in the cytosol was the primary limiting factor for caspase-3 activation in HCW-2 cells. To explore the mechanism which regulates mitochondrial cytochrome c efflux, we developed an in vitro assay which showed that cytosolic extracts from STS-treated, but not untreated, HL-60 cells contained an activity, which we designated 'CIF' (cytochrome c-efflux inducing factor), which rapidly induced cytochrome c efflux from HL-60 mitochondria. In contrast, there was no detectable CIF activity in STS-treated HCW-2 cells although the mitochondria from HCW-2 cells were responsive to the CIF activity from STS-treated HL-60 cells. These experiments have identified a novel activity, CIF, which is required for cytochrome c efflux and they indicate that the absence of CIF is the biochemical explanation for the impaired ability of HCW-2 cells to activate caspase-3 and undergo apoptosis.
Han
Z
Z
Box G, Department of Molecular Biology, Cell Biology, and Biochemistry, Brown University, Providence, Rhode Island 02912, USA.
Li
G
G
Bremner
T A
TA
Lange
T S
TS
Zhang
G
G
Jemmerson
R
R
Wyche
J H
JH
Hendrickson
E A
EA
D017209:Apoptosis; D001685:Biological Factors / Q000378:metabolism*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002999:Clone Cells; D003574:Cytochrome c Group / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism*; D003838:Deoxyadenine Nucleotides / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D018922:HL-60 Cells; D006801:Humans; D007211:Indoles / Q000378:metabolism; D007700:Kinetics; D008928:Mitochondria / Q000378:metabolism*; D019311:Staurosporine / Q000494:pharmacology
D001685:Biological Factors; D003574:Cytochrome c Group; D003838:Deoxyadenine Nucleotides; D007211:Indoles; C007293:DAPI; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D019311:Staurosporine; C026600:2'-deoxyadenosine triphosphate