title: Expanded CUG repeats Dysregulate RNA splicing by altering the stoichiometry of the muscleblind 1 complex.
abstract: To understand the role of the splice regulator muscleblind 1 (MBNL1) in the development of RNA splice defects in myotonic dystrophy I (DM1), we purified RNA-independent MBNL1 complexes from normal human myoblasts and examined the behavior of these complexes in DM1 myoblasts. Antibodies recognizing MBNL1 variants (MBNL1(CUG)), which can sequester in the toxic CUG RNA foci that develop in DM1 nuclei, were used to purify MBNL1(CUG) complexes from normal myoblasts. In normal myoblasts, MBNL1(CUG) bind 10 proteins involved in remodeling ribonucleoprotein complexes including hnRNP H, H2, H3, F, A2/B1, K, L, DDX5, DDX17, and DHX9. Of these proteins, only MBNL1(CUG) colocalizes extensively with DM1 CUG foci (>80% of foci) with its partners being present in <10% of foci. Importantly, the stoichiometry of MBNL1(CUG) complexes is altered in DM1 myoblasts, demonstrating an increase in the steady state levels of nine of its partner proteins. These changes are recapitulated by the expression of expanded CUG repeat RNA in Cos7 cells. Altered stoichiometry of MBNL1(CUG) complexes results from aberrant protein synthesis or stability and is unlinked to PKCα function. Modeling these changes in normal myoblasts demonstrates that increased levels of hnRNP H, H2, H3, F, and DDX5 independently dysregulate splicing in overlapping RNA subsets. Thus expression of expanded CUG repeats alters the stoichiometry of MBNL1(CUG) complexes to allow both the reinforcement and expansion of RNA processing defects.
authors: Department of Biochemistry and Molecular Biology, University of Southern California, Los Angeles, California 90033.
authors: Sharan
authors: S
authors: Paul
authors: Department of Biochemistry and Molecular Biology, University of Southern California, Los Angeles, California 90033.
authors: Warunee
authors: W
authors: Dansithong
authors: Department of Biochemistry and Molecular Biology, University of Southern California, Los Angeles, California 90033.
authors: Sonali P
authors: SP
authors: Jog
authors: Wolfson Centre for Inherited Neuromuscular Disease, Robert Jones and Agnes Hunt Orthopedic Hospital, Oswestry SY10 7AG, United Kingdom; Institute for Science and Technology in Medicine, Keele University, Keele ST5 5BG, United Kingdom.
authors: Ian
authors: I
authors: Holt
authors: Institute of Genetics, University of Nottingham, Queen's Medical Centre, Nottingham NG7 2UH, United Kingdom.
authors: Saloni
authors: S
authors: Mittal
authors: Institute of Genetics, University of Nottingham, Queen's Medical Centre, Nottingham NG7 2UH, United Kingdom.
authors: J David
authors: JD
authors: Brook
authors: Wolfson Centre for Inherited Neuromuscular Disease, Robert Jones and Agnes Hunt Orthopedic Hospital, Oswestry SY10 7AG, United Kingdom; Institute for Science and Technology in Medicine, Keele University, Keele ST5 5BG, United Kingdom.
authors: Glenn E
authors: GE
authors: Morris
authors: Department Molecular Microbiology and Immunology, Institute for Genetic Medicine, University of Southern California, Los Angeles, California 90033. Electronic address: comai@usc.edu.
authors: Lucio
authors: L
authors: Comai
authors: Department of Biochemistry and Molecular Biology, University of Southern California, Los Angeles, California 90033. Electronic address: sitaredd@usc.edu.
authors: Sita
authors: S
authors: Reddy
mesh_terms: D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D034441:Heterogeneous-Nuclear Ribonucleoproteins / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D013058:Mass Spectrometry / Q000379:methods; D018482:Muscle, Skeletal / Q000378:metabolism; D009223:Myotonic Dystrophy / Q000235:genetics / Q000378:metabolism*; D025941:Protein Interaction Mapping / Q000379:methods; D012326:RNA Splicing; D034741:RNA, Small Interfering / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism*; D013347:Subcellular Fractions
chemical_list: D034441:Heterogeneous-Nuclear Ribonucleoproteins; C493663:MBNL1 protein, human; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins