BH-3-only BIK functions at the endoplasmic reticulum to stimulate cytochrome c release from mitochondria.
Stimulation of apoptosis by p53 is accompanied by induction of the BH-3-only proapoptotic member of the BCL-2 family, BIK, and ectopic expression of BIK in p53-null cells caused the release of cytochrome c from mitochondria and activation of caspases, dependent on a functional BH-3 domain. A significant fraction of BIK, which contains a predicted transmembrane segment at its COOH terminus, was found inserted in the endoplasmic reticulum (ER) membrane, with the bulk of the protein facing the cytosol. Restriction of BIK to this membrane by replacing its transmembrane segment with the ER-selective membrane anchor of cytochrome b(5) also retained the cytochrome c release and cell death-inducing activity of BIK. Whereas induction of cell death by BIK was strongly inhibited by the caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone, the inhibitor was without effect on the ability of BIK to stimulate egress of cytochrome c from mitochondria. This benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone-insensitive pathway for stimulating cytochrome c release from mitochondria by ER BIK was successfully reconstituted in vitro and identified the requirement for components present in the light membrane (ER) and cytosol as necessary for this activity. Collectively, the results identify BIK as an initiator of cytochrome c release from mitochondria operating from a location at the ER.
Germain
Marc
M
Department of Biochemistry, McGill University, Montréal, Québec H3G 1Y6, Canada.
Mathai
Jaigi P
JP
Shore
Gordon C
GC
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000502:physiology; D051017:Apoptosis Regulatory Proteins; D002084:Butylated Hydroxytoluene / Q000031:analogs & derivatives / Q000494:pharmacology*; D061945:Caspase Inhibitors; D003574:Cytochrome c Group / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008565:Membrane Proteins; D008928:Mitochondria / Q000201:enzymology*; D024101:Mitochondrial Proteins; D011506:Proteins / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2; D014407:Tumor Cells, Cultured; D051028:bcl-2-Associated X Protein
D000590:Amino Acid Chloromethyl Ketones; D051017:Apoptosis Regulatory Proteins; C096820:BIK protein, human; D061945:Caspase Inhibitors; D003574:Cytochrome c Group; D004791:Enzyme Inhibitors; D008565:Membrane Proteins; D024101:Mitochondrial Proteins; D011506:Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; C426824:benzyloxycarbonyl-valyl-alanyl-(methyl)aspartic acid fluoromethyl ketone; D002084:Butylated Hydroxytoluene; C006008:BH 3