Differentiation-induced HL-60 cell apoptosis: a mechanism independent of mitochondrial disruption?
HL-60 cell differentiation into neutrophil like cells is associated with their induction of apoptosis. We investigated the cellular events that occur pre and post mitochondrial permeability transition to determine the role of the mitochondria in the induction of differentiation induced apoptosis. Pro-apoptotic Bax was translocated to and cleaved at the mitochondrial membrane in addition to t-Bid activation. These processes contributed to mitochondrial membrane disruption and the release of cytochrome c and Smac/DIABLO. The release of cytochrome c was caspase independent, as the caspase inhibitor Z-VAD.fmk, which inhibited apoptosis, did not block the release of cytochrome c. In contrast, the release of Smac/DIABLO was partially inhibited by caspase inhibition indicating differential release pathways for these mitochondrial pro-apoptotic factors. In addition to caspase inhibition we assessed the effects of the Bcl-2 anti-apoptotic family on differentiation induced apoptosis. BH4-Bcl-xl-TAT recombinant protein did not delay apoptosis, but did block the release of cytochrome c and Smac/DIABLO. Bcl-2 over-expression also inhibited differentiation induced apoptosis but was associated with the inhibition of the differentiation process. Differentiation mediated mitochondrial release of cytochrome c and Smac/DIABLO, may not trigger the induction of apoptosis, as BH4-Bclxl-TAT blocks the release of pro-apoptotic factors from the mitochondria, but does not prevent apoptosis.
Doyle
B T
BT
Department of Surgery, Mater Misericordiae University Hospital, Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Ireland.
O'Neill
A J
AJ
Fitzpatrick
J M
JM
Watson
R W G
RW
D015703:Antigens, CD / Q000032:analysis; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D051023:BH3 Interacting Domain Death Agonist Protein; D015153:Blotting, Western; D039481:CD11b Antigen / Q000032:analysis; D002352:Carrier Proteins / Q000378:metabolism; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002454:Cell Differentiation; D045304:Cytochromes c / Q000378:metabolism; D018922:HL-60 Cells; D006801:Humans; D007425:Intracellular Membranes / Q000502:physiology; D047908:Intracellular Signaling Peptides and Proteins; D008928:Mitochondria / Q000473:pathology* / Q000502:physiology*; D024101:Mitochondrial Proteins / Q000378:metabolism; D010539:Permeability; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D051028:bcl-2-Associated X Protein
D015703:Antigens, CD; D051017:Apoptosis Regulatory Proteins; C494826:BAX protein, human; D051023:BH3 Interacting Domain Death Agonist Protein; C494814:BID protein, human; D039481:CD11b Antigen; D002352:Carrier Proteins; D061945:Caspase Inhibitors; C412403:DIABLO protein, human; D047908:Intracellular Signaling Peptides and Proteins; D024101:Mitochondrial Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D011994:Recombinant Proteins; D051028:bcl-2-Associated X Protein; D045304:Cytochromes c; D020169:Caspases