title: FUBP1: a new protagonist in splicing regulation of the DMD gene.
abstract: We investigated the molecular mechanisms for in-frame skipping of DMD exon 39 caused by the nonsense c.5480T>A mutation in a patient with Becker muscular dystrophy. RNase-assisted pull down assay coupled with mass spectrometry revealed that the mutant RNA probe specifically recruits hnRNPA1, hnRNPA2/B1 and DAZAP1. Functional studies in a human myoblast cell line transfected with DMD minigenes confirmed the splicing inhibitory activity of hnRNPA1 and hnRNPA2/B1, and showed that DAZAP1, also known to activate splicing, acts negatively in the context of the mutated exon 39. Furthermore, we uncovered that recognition of endogenous DMD exon 39 in muscle cells is promoted by FUSE binding protein 1 (FUBP1), a multifunctional DNA- and RNA-binding protein whose role in splicing is largely unknown. By serial deletion and mutagenesis studies in minigenes, we delineated a functional intronic splicing enhancer (ISE) in intron 38. FUBP1 recruitment to the RNA sequence containing the ISE was established by RNA pull down and RNA EMSA, and further confirmed by RNA-ChIP on endogenous DMD pre-mRNA. This study provides new insights about the splicing regulation of DMD exon 39, highlighting the emerging role of FUBP1 in splicing and describing the first ISE for constitutive exon inclusion in the mature DMD transcript.
authors: Université Montpellier, UFR de Médecine, Montpellier F-34000, France Inserm U827, Laboratoire de Génétique de Maladies Rares, F-34000 Montpellier, France.
authors: Julie
authors: J
authors: Miro
authors: Université Montpellier, UFR de Médecine, Montpellier F-34000, France Inserm U827, Laboratoire de Génétique de Maladies Rares, F-34000 Montpellier, France.
authors: Abdelhamid Mahdi
authors: AM
authors: Laaref
authors: UR1199 Laboratoire de Protéomique Fonctionnelle, INRA, 34060 Montpellier cedex, France.
authors: Valérie
authors: V
authors: Rofidal
authors: Université Montpellier, UFR de Médecine, Montpellier F-34000, France Inserm U827, Laboratoire de Génétique de Maladies Rares, F-34000 Montpellier, France.
authors: Rosyne
authors: R
authors: Lagrafeuille
authors: UR1199 Laboratoire de Protéomique Fonctionnelle, INRA, 34060 Montpellier cedex, France.
authors: Sonia
authors: S
authors: Hem
authors: CHU Montpellier, Hôpital Arnaud de Villeneuve, Laboratoire de Génétique Moléculaire, F-34000 Montpellier, France.
authors: Delphine
authors: D
authors: Thorel
authors: CHU Montpellier, Hôpital Arnaud de Villeneuve, Laboratoire de Génétique Moléculaire, F-34000 Montpellier, France.
authors: Déborah
authors: D
authors: Méchin
authors: Institut de Myologie, UM76 Université Pierre et Marie Curie (UPMC), Paris, France INSERM U 974, Paris, France CNRS UMR 7215, Paris, France.
authors: Kamel
authors: K
authors: Mamchaoui
authors: Institut de Myologie, UM76 Université Pierre et Marie Curie (UPMC), Paris, France INSERM U 974, Paris, France CNRS UMR 7215, Paris, France.
authors: Vincent
authors: V
authors: Mouly
authors: Université Montpellier, UFR de Médecine, Montpellier F-34000, France Inserm U827, Laboratoire de Génétique de Maladies Rares, F-34000 Montpellier, France CHU Montpellier, Hôpital Arnaud de Villeneuve, Laboratoire de Génétique Moléculaire, F-34000 Montpellier, France.
authors: Mireille
authors: M
authors: Claustres
authors: Université Montpellier, UFR de Médecine, Montpellier F-34000, France Inserm U827, Laboratoire de Génétique de Maladies Rares, F-34000 Montpellier, France sylvie.tuffery@inserm.fr.
authors: Sylvie
authors: S
authors: Tuffery-Giraud
mesh_terms: D017398:Alternative Splicing; D001665:Binding Sites; D002460:Cell Line; D018389:Codon, Nonsense; D004265:DNA Helicases / Q000378:metabolism* / Q000502:physiology; D004268:DNA-Binding Proteins / Q000378:metabolism* / Q000502:physiology; D016189:Dystrophin / Q000235:genetics*; D005091:Exons; D006801:Humans; D007438:Introns; D016601:RNA-Binding Proteins / Q000378:metabolism* / Q000502:physiology; D038621:Regulatory Sequences, Ribonucleic Acid
chemical_list: D018389:Codon, Nonsense; C484258:DMD protein, human; D004268:DNA-Binding Proteins; D016189:Dystrophin; C492258:FUBP1 protein, human; D016601:RNA-Binding Proteins; D038621:Regulatory Sequences, Ribonucleic Acid; D004265:DNA Helicases