Abstract
Myotonic dystrophy is the most common muscular dystrophy in adults and the first recognized example of an RNA-mediated disease. Congenital myotonic dystrophy (CDM1) and myotonic dystrophy of type 1 (DM1) or of type 2 (DM2) are caused by the expression of mutant RNAs containing expanded CUG or CCUG repeats, respectively. These mutant RNAs sequester the splicing regulator Muscleblind-like-1 (MBNL1), resulting in specific misregulation of the alternative splicing of other pre-mRNAs. We found that alternative splicing of the bridging integrator-1 (BIN1) pre-mRNA is altered in skeletal muscle samples of people with CDM1, DM1 and DM2. BIN1 is involved in tubular invaginations of membranes and is required for the biogenesis of muscle T tubules, which are specialized skeletal muscle membrane structures essential for excitation-contraction coupling. Mutations in the BIN1 gene cause centronuclear myopathy, which shares some histopathological features with myotonic dystrophy. We found that MBNL1 binds the BIN1 pre-mRNA and regulates its alternative splicing. BIN1 missplicing results in expression of an inactive form of BIN1 lacking phosphatidylinositol 5-phosphate–binding and membrane-tubulating activities. Consistent with a defect of BIN1, muscle T tubules are altered in people with myotonic dystrophy, and membrane structures are restored upon expression of the normal splicing form of BIN1 in muscle cells of such individuals. Finally, reproducing BIN1 splicing alteration in mice is sufficient to promote T tubule alterations and muscle weakness, a predominant feature of myotonic dystrophy.
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Acknowledgements
We thank T. Cooper (Baylor College of Medicine) for the gift of the β-globin 4.11.12, DMPKS, DT960 and tgCUGBP1 plasmids, L. Ranum (University of Minnesota) for the gift of the CCTG300 expression plasmid, C. Branlant (CNRS) for the gift of the pGEX-MBNL1Δ101 vector, M. Swanson (University of Florida) for the gift of the pGEX-6P-MBNL1-His vector, P. Hawkins (Babraham Research Campus) for the gift of the GFP-PH domain of PLCg and GFP-PX domain of P40 constructs, O. Gozani (Stanford University) for the gift of the GFP-PHD domain of ING2 vector, Z. Xue (Université Paris 7) for the gift of the synemin-specific antibody, I. Marty (INSERM) for the gift of the RYR1-specific antibody, M. Swanson (University of Florida), G. Gourdon (INSERM) and C. Thornton (University of Rochester) for the gift of RNA from skeletal muscles of MBNL1ΔE3, DM300 and HSALR mice, respectively, the IGBMC facilities for assistance, J. Lainé, G. Butler-Browne and all members of the French DM Network for fruitful discussion. This work was supported by INSERM AVENIR (N.C.-B.), Agence nationale de la recherche GENOPAT 07-942 (N.C.-B.), 08-005 (J.L.) and BLANC 07-065 (J.L.), Association Française contre les Myopathies MNM1 12982 (N.C.-B.), 12570 and 14269 (D.F.), 12576 and 14058 (J.L.), Fondation pour la Recherche Médicale 20071210538 (J.L.), Japan Society for the Promotion of Science KAKENHI 20590998 (M.P.T.) and a European Molecular Biology Organization long-term fellowship (Y.I.).
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Experiments were carried out by C.F., A.F.K., C.H., S.V., Y.I., A.T., V.T., A.V., A.F., N.M., Y.K., R.T., V.F., G.P., C.B.-L. F.D. and M.-C.H. Bioinformatic analyses were done by P.d.l.G., D.D., C.T. and D.A. Clinical samples and patient data were from A.L.d.M., N.S., A.L., B.U., B.S., M.P.T., I.N., G.B. and D.F. Data were collected and analyzed by N.M., C.T., D.A., L.G., P.Z., I.N., M.P.T., B.U., B.S., G.B., J.L., D.F. and N.C.-B. The study was designed, coordinated and written by D.F. and N.C.-B.
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Fugier, C., Klein, A., Hammer, C. et al. Misregulated alternative splicing of BIN1 is associated with T tubule alterations and muscle weakness in myotonic dystrophy. Nat Med 17, 720–725 (2011). https://doi.org/10.1038/nm.2374
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DOI: https://doi.org/10.1038/nm.2374
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