Apoptosis-associated mitochondrial outer membrane permeabilization assays.
Following most cell death signals, pro-apoptotic Bcl-2 members as Bax and Bak are activated and oligomerize into the mitochondria outer membrane, triggering its permeabilization and release into the cytosol of soluble apoptogenic factors such as cytochrome c involved in caspase activation. Thus, in many studies focused on apoptosis, cytochrome c release within cells is frequently examined to assess Bax/Bak activation and mitochondrial outer membrane permeabilization. In addition, cytochrome c release can also be investigated in vitro in functional mitochondria that have been isolated from cultured cells, offering a number of advantages. Here, protocols for measuring cytochrome c release from intact cells as well as from isolated mitochondria is detailed. Finally, assays to investigate Bax/Bak activation and olimerization are also presented.
Arnoult
Damien
D
INSERM U 542, Université Paris-Sud, Hopital Paul Brousse, Batiment Lavoisier, 14 avenue Paul Vaillant Couturier, 94807 Villejuif cedex, France. damarn@vjf.inserm.fr
D000818:Animals; D017209:Apoptosis; D002463:Cell Membrane Permeability; D045304:Cytochromes c / Q000378:metabolism*; D006801:Humans; D051336:Mitochondrial Membranes / Q000378:metabolism*; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000737:chemistry / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000737:chemistry / Q000378:metabolism
D050998:bcl-2 Homologous Antagonist-Killer Protein; D051028:bcl-2-Associated X Protein; D045304:Cytochromes c