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2009-11-09 - Colloque/Présentation - communication orale - Anglais - 1 page(s)

Tassin Alexandra , Coppée Frédérique , Leroy Baptiste , Sauvage Sébastien, Preillon J, Erculisse V., Charron Sébastien , Vanderplanck Céline, Barro Marietta, Vander Elst Luce , Muller Robert , Laoudj-Chenivesse Dalila, Colet Jean-Marie , Wattiez Ruddy , Belayew Alexandra , "Investigations on the molecular mechanism of a muscular dystrophy (FSHD) by proteomic and metabonomic analyses of primary myoblast cultures" in FSH Society International Research Consortium and Research Planning Meetings, Boston, USA, 2009

  • Codes CREF : Biologie moléculaire (DI3111), Pathologies particulières (DI3370)
  • Unités de recherche UMONS : Chimie générale, organique et biomédicale (M108), Biologie moléculaire (M122), Biologie humaine et Toxicologie (M125), Protéomie et Microbiologie (S828)

Abstract(s) :

(Anglais) We identified a double homeobox gene (DUX4) within each D4Z4 unit at the FSHD locus that is activated in muscle cells following the FSHD deletion (Gabriels et al., 1999; Kowaljow et al., 2007; Dixit et al., 2007). The DUX4 protein is a transcription factor that targets a number of genes, some of which encode other transcription factors. Among others, DUX4 inhibits MYOD gene expression, leading to myoblast differentiation defects, activates PITX1 that induces muscle atrophy, and inhibits genes involved in response to oxidative stress (Bosnakovski et al., 2008; Dixit et al., 2007). Several studies investigated gene and/or protein expression in FSHD primary myoblasts or muscle biopsies. Since DUX4 is induced upon myoblast differentiation to myotubes, we chose to compare the proteome of FSHD and control myotubes by gel free differential mass spectrometry analysis using an isotope coded protein labelling (ICPL). In keeping with an FSHD differentiation defect we observed increased amounts of vimentin, an intermediate filament only expressed in early myogenesis. Co-transfection of C2C12 cells with a DUX4 expression vector and a luciferase reporter gene fused to the vimentin promoter yields a 4-fold induction of luciferase activity in keeping with the presence of a DUX4 consensus sequence. In addition, galectin-1 that is normally decreased during differentiation remains at higher levels in FSHD myotubes. These observations were confirmed by specific immunodetection on Western blots. We found a general decrease of cytoskeletal and contractile proteins that could reflect an atrophy-associated proteolysis as well as differentiation defects. Our data also confirmed a mitochondrial dysfunction in FSHD: enzymes involved in fatty acid ß-oxidation, Kreb’s cycle and the electron transport chain were decreased. There was no change in glycolytic enzymes but an increase of lactate dehydrogenase that correlated with lactate accumulation in the culture medium as observed by a metabonomics analysis (proton Nuclear Magnetic Resonance spectra). Bosnakovski D, Xu Z, Gang EJ, Galindo CL, Liu M, Simsek T, Garner HR, gha-Mohammadi S, Tassin A, Coppee F, Belayew A, Perlingeiro RR, Kyba M. 2008 Octb. An isogenetic myoblast expression screen identifies DUX4-mediated FSHD-associated molecular pathologies. EMBO J 27(20):2766-2779. Dixit M, Ansseau E, Tassin A, Winokur S, Shi R, Qian H, Sauvage S, Matteotti C, van Acker AM, Leo O, Figlewicz D, Barro M, Laoudj-Chenivesse D, Belayew A, Coppee F, Chen YW. 2007 Nova. DUX4, a candidate gene of facioscapulohumeral muscular dystrophy, encodes a transcriptional activator of PITX1. Proc Natl Acad Sci U S A 104(46):18157-18162. Gabriels J, Beckers MC, Ding H, De Vriese A, Plaisance S, van der Maarel SM, Padberg GW, Frants RR, Hewitt JE, Collen D, Belayew A. 1999 Aug. Nucleotide sequence of the partially deleted D4Z4 locus in a patient with FSHD identifies a putative gene within each 3.3 kb element. Gene 236(1):25-32. Kowaljow V, Marcowycz A, Ansseau E, Conde CB, Sauvage S, Matteotti C, Arias C, Corona ED, Nunez NG, Leo O, Wattiez R, Figlewicz D, Laoudj-Chenivesse D, Belayew A, Coppee F, Rosa AL. 2007 Aug. The DUX4 gene at the FSHD1A locus encodes a pro-apoptotic protein. Neuromuscul Disord 17(8):611-623.