title: BH3-only proteins are tail-anchored in the outer mitochondrial membrane and can initiate the activation of Bax.
abstract: During mitochondrial apoptosis, pro-apoptotic BH3-only proteins cause the translocation of cytosolic Bcl-2-associated X protein (Bax) to the outer mitochondrial membrane (OMM) where it is activated to release cytochrome c from the mitochondrial intermembrane space, but the mechanism is under dispute. We show that most BH3-only proteins are mitochondrial proteins that are imported into the OMM via a C-terminal tail-anchor domain in isolated yeast mitochondria, independently of binding to anti-apoptotic Bcl-2 proteins. This C-terminal domain acted as a classical mitochondrial targeting signal and was sufficient to direct green fluorescent protein to mitochondria in human cells. When expressed in mouse fibroblasts, these BH3-only proteins localised to mitochondria and were inserted in the OMM. The BH3-only proteins Bcl-2-interacting mediator of cell death (Bim), tBid and p53-upregulated modulator of apoptosis sensitised isolated mitochondria from Bax/Bcl-2 homologous antagonist/killer-deficient fibroblasts to cytochrome c-release by recombinant, extramitochondrial Bax. For Bim, this activity is shown to require the C-terminal-targeting signal and to be independent of binding capacity to and presence of anti-apoptotic Bcl-2 proteins. Bim further enhanced Bax-dependent killing in yeast. A model is proposed where OMM-tail-anchored BH3-only proteins permit passive 'recruitment' and catalysis-like activation of extra-mitochondrial Bax. The recognition of C-terminal membrane-insertion of BH3-only proteins will permit the development of a more detailed concept of the initiation of mitochondrial apoptosis.
authors.affiliation: Institute of Medical Microbiology, Technische Universität München, Germany.
authors.forename: F
authors.initials: F
authors.lastname: Wilfling
authors.forename: A
authors.initials: A
authors.lastname: Weber
authors.forename: S
authors.initials: S
authors.lastname: Potthoff
authors.forename: F-N
authors.initials: FN
authors.lastname: Vögtle
authors.forename: C
authors.initials: C
authors.lastname: Meisinger
authors.forename: S A
authors.initials: SA
authors.lastname: Paschen
authors.forename: G
authors.initials: G
authors.lastname: Häcker
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000502:physiology; D051023:BH3 Interacting Domain Death Agonist Protein / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D045744:Cell Line, Tumor; D006801:Humans; D051379:Mice; D018613:Microscopy, Confocal; D051336:Mitochondrial Membranes / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D051028:bcl-2-Associated X Protein / Q000378:metabolism*
chemical_list: D051023:BH3 Interacting Domain Death Agonist Protein; D024101:Mitochondrial Proteins; D051028:bcl-2-Associated X Protein