title: A conserved N-terminal sequence targets human DAP3 to mitochondria.
abstract: Human DAP3 (death-associated protein-3) has been identified as an essential positive mediator of programmed cell death. Structure-function studies have shown previously the N-terminal extremity of the protein to be required in apoptosis induction. Analysis of human DAP3 gene structure predicted 13 exons and subsequent targeting prediction by two software packages (MITOPROT and TargetP) gave a high probability for mitochondrial targeting. The predicted N-terminal targeting structure was also found in the mouse, Drosophila, and C. elegans orthologues with a strong sequence homology between mouse and human. Secondary structure analyses identified alpha-helical structures typical of mitochondrial target peptides. To confirm experimentally this targeting we constructed a fusion protein with N-terminal human DAP3 upstream of enhanced green fluorescent protein (EGFP). Confocal analysis of transfected human fibroblasts clearly demonstrated EGFP localization exclusive to mitochondria. The positioning of this key apoptotic factor at the heart of the mitochondrial pathway provides exciting insight into its role in programmed cell death.
authors.affiliation: Laboratoire de Biochimie et Biologie Moléculaire, INSERM EMI-U 00-18, CHU d'Angers, 4 rue Larrey, F-49033 Angers, France. morgan@med.univ-angers.fr
authors.forename: C J
authors.initials: CJ
authors.lastname: Morgan
authors.forename: C
authors.initials: C
authors.lastname: Jacques
authors.forename: F
authors.initials: F
authors.lastname: Savagner
authors.forename: Y
authors.initials: Y
authors.lastname: Tourmen
authors.forename: D P
authors.initials: DP
authors.lastname: Mirebeau
authors.forename: Y
authors.initials: Y
authors.lastname: Malthièry
authors.forename: P
authors.initials: P
authors.lastname: Reynier
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D017173:Caenorhabditis elegans; D002460:Cell Line; D017124:Conserved Sequence; D004330:Drosophila; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000032:analysis; D051379:Mice; D008928:Mitochondria / Q000378:metabolism*; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism*; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D012269:Ribosomal Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014162:Transfection
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; C097041:DAP3 protein, human; C490762:Dap3 protein, mouse; D008164:Luminescent Proteins; D011506:Proteins; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D012269:Ribosomal Proteins; D049452:Green Fluorescent Proteins