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The molecular archaeology of a mitochondrial death effector: AIF in Drosophila.

ABSTRACT: Apoptosis-inducing factor (AIF) is a phylogenetically conserved redox-active flavoprotein that contributes to cell death and oxidative phosphorylation in Saccharomyces cerevisiae, Caenorhabditis elegans, mouse and humans. AIF has been characterized as a caspase-independent death effector that is activated by its translocation from mitochondria to the cytosol and nucleus. Here, we report the molecular characterization of AIF in Drosophila melanogaster, a species in which most cell deaths occur in a caspase-dependent manner. Interestingly, knockout of zygotic D. melanogaster AIF (DmAIF) expression using gene targeting resulted in decreased embryonic cell death and the persistence of differentiated neuronal cells at late embryonic stages. Although knockout embryos hatch, they undergo growth arrest at early larval stages, accompanied by mitochondrial respiratory dysfunction. Transgenic expression of DmAIF misdirected to the extramitochondrial compartment (DeltaN-DmAIF), but not wild-type DmAIF, triggered ectopic caspase activation and cell death. DeltaN-DmAIF-induced death was not blocked by removal of caspase activator Dark or transgenic expression of baculoviral caspase inhibitor p35, but was partially inhibited by Diap1 overexpression. Knockdown studies revealed that DeltaN-DmAIF interacts genetically with the redox protein thioredoxin-2. In conclusion, we show that Drosophila AIF is a mitochondrial effector of cell death that plays roles in developmentally regulated cell death and normal mitochondrial function.

SUBMITTER: Joza N

PROVIDER: S-EPMC2907157 | biostudies-literature | 2008 Jun

REPOSITORIES: biostudies-literature

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The molecular archaeology of a mitochondrial death effector: AIF in Drosophila.

Joza N N Galindo K K Pospisilik J A JA Benit P P Rangachari M M Kanitz E E EE Nakashima Y Y Neely G G GG Rustin P P Abrams J M JM Kroemer G G Penninger J M JM

Cell death and differentiation 20080229 6

Apoptosis-inducing factor (AIF) is a phylogenetically conserved redox-active flavoprotein that contributes to cell death and oxidative phosphorylation in Saccharomyces cerevisiae, Caenorhabditis elegans, mouse and humans. AIF has been characterized as a caspase-independent death effector that is activated by its translocation from mitochondria to the cytosol and nucleus. Here, we report the molecular characterization of AIF in Drosophila melanogaster, a species in which most cell deaths occur in ...[more]

PMID: 18309327

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Project description:ObjectivesApoptosis-Inducing Factor (AIF) is a protein involved in mitochondrial electron transport chain assembly/stability and programmed cell death. The relevant role of this protein is underlined because mutations altering mitochondrial AIF properties result in acute pediatric mitochondriopathies and tumor metastasis. By generating an original AIF-deficient mouse strain, this study attempted to analyze, in a single paradigm, the cellular and developmental metabolic consequences of AIF loss and the subsequent oxidative phosphorylation (OXPHOS) dysfunction.MethodsWe developed a novel AIF-deficient mouse strain and assessed, using molecular and cell biology approaches, the cellular, embryonic, and adult mice phenotypic alterations. Additionally, we conducted ex vivo assays with primary and immortalized AIF knockout mouse embryonic fibroblasts (MEFs) to establish the cell death characteristics and the metabolic adaptive responses provoked by the mitochondrial electron transport chain (ETC) breakdown.ResultsAIF deficiency destabilized mitochondrial ETC and provoked supercomplex disorganization, mitochondrial transmembrane potential loss, and high generation of mitochondrial reactive oxygen species (ROS). AIF-/Y MEFs counterbalanced these OXPHOS alterations by mitochondrial network reorganization and a metabolic reprogramming toward anaerobic glycolysis illustrated by the AMPK phosphorylation at Thr172, the overexpression of the glucose transporter GLUT-4, the subsequent enhancement of glucose uptake, and the anaerobic lactate generation. A late phenotype was characterized by the activation of P53/P21-mediated senescence. Notably, approximately 2% of AIF-/Y MEFs diminished both mitochondrial mass and ROS levels and spontaneously proliferated. These cycling AIF-/Y MEFs were resistant to caspase-independent cell death inducers. The AIF-deficient mouse strain was embryonic lethal between E11.5 and E13.5 with energy loss, proliferation arrest, and increased apoptotic levels. Contrary to AIF-/Y MEFs, the AIF KO embryos were unable to reprogram their metabolism toward anaerobic glycolysis. Heterozygous AIF+/- females displayed progressive bone marrow, thymus, and spleen cellular loss. In addition, approximately 10% of AIF+/- females developed perinatal hydrocephaly characterized by brain development impairment, meningeal fibrosis, and medullar hemorrhages; those mice died 5 weeks after birth. AIF+/- with hydrocephaly exhibited loss of ciliated epithelium in the ependymal layer. This phenotype was triggered by the ROS excess. Accordingly, it was possible to diminish the occurrence of hydrocephalus AIF+/- females by supplying dams and newborns with an antioxidant in drinking water.ConclusionsIn a single knockout model and at 3 different levels (cell, embryo, and adult mice) we demonstrated that by controlling the mitochondrial OXPHOS/metabolism, AIF is a key factor regulating cell differentiation and fate. Additionally, by providing new insights into the pathological consequences of mitochondrial OXPHOS dysfunction, our new findings pave the way for novel pharmacological strategies.

Project description:While death is universal, reactions to death and ways of dealing with the dead body are hugely diverse, and archaeological research reveals numerous ways of dealing with the dead through time and across the world. In this paper, findings are presented which not only demonstrate the power of archaeology to promote and aid discussion around this difficult and challenging topic, but also how our approach resulted in personal growth and professional development impacts for participants. In this interdisciplinary pilot study, archaeological case studies were used in 31 structured workshops with 187 participants from health and social care backgrounds in the UK, to explore their reactions to a diverse range of materials which documented wide and varied approaches to death and the dead. Our study supports the hypothesis that the past is a powerful instigator of conversation around challenging aspects of death, and after death care and practices: 93% of participants agreed with this. That exposure to archaeological case studies and artefacts stimulates multifaceted discourse, some of it difficult, is a theme that also emerges in our data from pre, post and follow-up questionnaires, and semi-structured interviews. The material prompted participants to reflect on their biases, expectations and norms around both treatment of the dead, and of bereavement, impacting on their values, attitudes and beliefs. Moreover, 87% of participants believed the workshop would have a personal effect through thinking differently about death and bereavement, and 57% thought it would impact on how they approached death and bereavement in their professional practice. This has huge implications today, where talk of death remains troublesome, and for some, has a near-taboo status-'taboo' being a theme evident in some participants' own words. The findings have an important role to play in facilitating and normalising discussions around dying and bereavement and in equipping professionals in their work with people with advanced illness.

Dataset Information

The molecular archaeology of a mitochondrial death effector: AIF in Drosophila.

Publications

The molecular archaeology of a mitochondrial death effector: AIF in Drosophila.

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