title: The C-terminal Domains of Apoptotic BH3-only Proteins Mediate Their Insertion into Distinct Biological Membranes.
abstract: Changes in the equilibrium of pro- and anti-apoptotic members of the B-cell lymphoma-2 (Bcl-2) protein family in the mitochondrial outer membrane (MOM) induce structural changes that commit cells to apoptosis. Bcl-2 homology-3 (BH3)-only proteins participate in this process by either activating pro-apoptotic effectors or inhibiting anti-apoptotic components and by promoting MOM permeabilization. The association of BH3-only proteins with MOMs is necessary for the activation and amplification of death signals; however, the nature of this association remains controversial, as these proteins lack a canonical transmembrane sequence. Here we used an in vitro expression system to study the insertion capacity of hydrophobic C-terminal regions of the BH3-only proteins Bik, Bim, Noxa, Bmf, and Puma into microsomal membranes. An Escherichia coli complementation assay was used to validate the results in a cellular context, and peptide insertions were modeled using molecular dynamics simulations. We also found that some of the C-terminal domains were sufficient to direct green fluorescent protein fusion proteins to specific membranes in human cells, but the domains did not activate apoptosis. Thus, the hydrophobic regions in the C termini of BH3-only members associated in distinct ways with various biological membranes, suggesting that a detailed investigation of the entire process of apoptosis should include studying the membranes as a setting for protein-protein and protein-membrane interactions.
authors.affiliation: From the Departament de Bioquímica i Biologia Molecular, ERI BioTecMed, Universitat de València, E-46100 Burjassot, Spain.|the Laboratory of Peptide and Protein Chemistry, Centro de Investigación Príncipe Felipe, E-46012 Valencia, Spain, and.
authors.forename: Vicente
authors.initials: V
authors.lastname: Andreu-Fernández
authors.affiliation: From the Departament de Bioquímica i Biologia Molecular, ERI BioTecMed, Universitat de València, E-46100 Burjassot, Spain.
authors.forename: María J
authors.initials: MJ
authors.lastname: García-Murria
authors.affiliation: From the Departament de Bioquímica i Biologia Molecular, ERI BioTecMed, Universitat de València, E-46100 Burjassot, Spain.
authors.forename: Manuel
authors.identifier: http://orcid.org/0000-0002-4840-4480
authors.initials: M
authors.lastname: Bañó-Polo
authors.affiliation: the Bases Moléculaires et Structurales des Systèmes Infectieux (BMSSI), CNRS UMR 5086, 7 Passage du Vercors, 69007 Lyon, France.
authors.forename: Juliette
authors.initials: J
authors.lastname: Martin
authors.affiliation: the Bases Moléculaires et Structurales des Systèmes Infectieux (BMSSI), CNRS UMR 5086, 7 Passage du Vercors, 69007 Lyon, France.
authors.forename: Luca
authors.identifier: http://orcid.org/0000-0002-6352-4595
authors.initials: L
authors.lastname: Monticelli
authors.affiliation: the Laboratory of Peptide and Protein Chemistry, Centro de Investigación Príncipe Felipe, E-46012 Valencia, Spain, and.
authors.forename: Mar
authors.identifier: http://orcid.org/0000-0003-3231-5835
authors.initials: M
authors.lastname: Orzáez
authors.affiliation: From the Departament de Bioquímica i Biologia Molecular, ERI BioTecMed, Universitat de València, E-46100 Burjassot, Spain, Ismael.Mingarro@uv.es.
authors.forename: Ismael
authors.identifier: http://orcid.org/0000-0002-1910-1229
authors.initials: I
authors.lastname: Mingarro
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051017:Apoptosis Regulatory Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000072224:Bcl-2-Like Protein 11 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008861:Microsomes / Q000737:chemistry / Q000378:metabolism*; D024101:Mitochondrial Proteins; D000072417:Protein Domains; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; C438378:BBC3 protein, human; C096820:BIK protein, human; C486655:BMF protein, human; D000072224:Bcl-2-Like Protein 11; D008565:Membrane Proteins; D024101:Mitochondrial Proteins; C498702:PMAIP1 protein, human; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2
keywords: B-cell lymphoma 2 (Bcl-2) family; BH3-only; apoptosis; membrane insertion; membrane protein; mitochondrial apoptosis; transmembrane domain