Dysferlin rescue by spliceosome-mediated pre-mRNA trans-splicing targeting introns harbouring weakly defined 3' splice sites. 2015

Susanne Philippi, and Stéphanie Lorain, and Cyriaque Beley, and Cécile Peccate, and Guillaume Précigout, and Simone Spuler, and Luis Garcia
Université de Versailles St-Quentin, INSERM U1179, LIA BAHN Centre Scientifique de Monaco, 2 Avenue de la Source de la Bievre, Montigny-le-Bretonneux 78180, France, Muscle Research Unit, Experimental and Clinical Research Center, a Joint Cooperation Between Max-Delbrück-Center for Molecular Medicine and Charité Medical Faculty, Berlin, Germany and Sorbonne Universités, UPMC Univ Paris 06, INSERM UMRS974, CNRS FRE3617, Myology Research Center, Paris, France.

The modification of the pre-mRNA cis-splicing process employing a pre-mRNA trans-splicing molecule (PTM) is an attractive strategy for the in situ correction of genes whose careful transcription regulation and full-length expression is determinative for protein function, as it is the case for the dysferlin (DYSF, Dysf) gene. Loss-of-function mutations of DYSF result in different types of muscular dystrophy mainly manifesting as limb girdle muscular dystrophy 2B (LGMD2B) and Miyoshi muscular dystrophy 1 (MMD1). We established a 3' replacement strategy for mutated DYSF pre-mRNAs induced by spliceosome-mediated pre-mRNA trans-splicing (SmaRT) by the use of a PTM. In contrast to previously established SmaRT strategies, we particularly focused on the identification of a suitable pre-mRNA target intron other than the optimization of the PTM design. By targeting DYSF pre-mRNA introns harbouring differentially defined 3' splice sites (3' SS), we found that target introns encoding weakly defined 3' SSs were trans-spliced successfully in vitro in human LGMD2B myoblasts as well as in vivo in skeletal muscle of wild-type and Dysf(-/-) mice. For the first time, we demonstrate rescue of Dysf protein by SmaRT in vivo. Moreover, we identified concordant qualities among the successfully targeted Dysf introns and targeted endogenous introns in previously reported SmaRT approaches that might facilitate a selective choice of target introns in future SmaRT strategies.

UI MeSH Term Description Entries
D007438 Introns Sequences of DNA in the genes that are located between the EXONS. They are transcribed along with the exons but are removed from the primary gene transcript by RNA SPLICING to leave mature RNA. Some introns code for separate genes. Intervening Sequences,Sequences, Intervening,Intervening Sequence,Intron,Sequence, Intervening
D008565 Membrane Proteins Proteins which are found in membranes including cellular and intracellular membranes. They consist of two types, peripheral and integral proteins. They include most membrane-associated enzymes, antigenic proteins, transport proteins, and drug, hormone, and lectin receptors. Cell Membrane Protein,Cell Membrane Proteins,Cell Surface Protein,Cell Surface Proteins,Integral Membrane Proteins,Membrane-Associated Protein,Surface Protein,Surface Proteins,Integral Membrane Protein,Membrane Protein,Membrane-Associated Proteins,Membrane Associated Protein,Membrane Associated Proteins,Membrane Protein, Cell,Membrane Protein, Integral,Membrane Proteins, Integral,Protein, Cell Membrane,Protein, Cell Surface,Protein, Integral Membrane,Protein, Membrane,Protein, Membrane-Associated,Protein, Surface,Proteins, Cell Membrane,Proteins, Cell Surface,Proteins, Integral Membrane,Proteins, Membrane,Proteins, Membrane-Associated,Proteins, Surface,Surface Protein, Cell
D008810 Mice, Inbred C57BL One of the first INBRED MOUSE STRAINS to be sequenced. This strain is commonly used as genetic background for transgenic mouse models. Refractory to many tumors, this strain is also preferred model for studying role of genetic variations in development of diseases. Mice, C57BL,Mouse, C57BL,Mouse, Inbred C57BL,C57BL Mice,C57BL Mice, Inbred,C57BL Mouse,C57BL Mouse, Inbred,Inbred C57BL Mice,Inbred C57BL Mouse
D009124 Muscle Proteins The protein constituents of muscle, the major ones being ACTINS and MYOSINS. More than a dozen accessory proteins exist including TROPONIN; TROPOMYOSIN; and DYSTROPHIN. Muscle Protein,Protein, Muscle,Proteins, Muscle
D009136 Muscular Dystrophies A heterogeneous group of inherited MYOPATHIES, characterized by wasting and weakness of the SKELETAL MUSCLE. They are categorized by the sites of MUSCLE WEAKNESS; AGE OF ONSET; and INHERITANCE PATTERNS. Muscular Dystrophy,Myodystrophica,Myodystrophy,Dystrophies, Muscular,Dystrophy, Muscular,Myodystrophicas,Myodystrophies
D002478 Cells, Cultured Cells propagated in vitro in special media conducive to their growth. Cultured cells are used to study developmental, morphologic, metabolic, physiologic, and genetic processes, among others. Cultured Cells,Cell, Cultured,Cultured Cell
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000073939 Dysferlin A membrane protein that contains multiple C2 DOMAINS. It is highly expressed in skeletal muscle and functions as a calcium ion sensor in SYNAPTIC VESICLE-PLASMA MEMBRANE fusion, as well as in SARCOLEMMA repair following mechanical stress. Mutations in the dysferlin (DYSF) gene are associated with several hereditary MUSCULAR DYSTROPHIES. Fer-1-like Protein 1,Fer 1 like Protein 1
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D012322 RNA Precursors RNA transcripts of the DNA that are in some unfinished stage of post-transcriptional processing (RNA PROCESSING, POST-TRANSCRIPTIONAL) required for function. RNA precursors may undergo several steps of RNA SPLICING during which the phosphodiester bonds at exon-intron boundaries are cleaved and the introns are excised. Consequently a new bond is formed between the ends of the exons. Resulting mature RNAs can then be used; for example, mature mRNA (RNA, MESSENGER) is used as a template for protein production. Precursor RNA,Primary RNA Transcript,RNA, Messenger, Precursors,RNA, Ribosomal, Precursors,RNA, Small Nuclear, Precursors,RNA, Transfer, Precursors,Pre-mRNA,Pre-rRNA,Pre-snRNA,Pre-tRNA,Primary Transcript, RNA,RNA Precursor,mRNA Precursor,rRNA Precursor,snRNA Precursor,tRNA Precursor,Pre mRNA,Pre rRNA,Pre snRNA,Pre tRNA,Precursor, RNA,Precursor, mRNA,Precursor, rRNA,Precursor, snRNA,Precursor, tRNA,Precursors, RNA,RNA Primary Transcript,RNA Transcript, Primary,RNA, Precursor,Transcript, Primary RNA,Transcript, RNA Primary

Related Publications

Susanne Philippi, and Stéphanie Lorain, and Cyriaque Beley, and Cécile Peccate, and Guillaume Précigout, and Simone Spuler, and Luis Garcia
September 2018, Molecular therapy. Nucleic acids,
Susanne Philippi, and Stéphanie Lorain, and Cyriaque Beley, and Cécile Peccate, and Guillaume Précigout, and Simone Spuler, and Luis Garcia
November 2000, Gene therapy,
Susanne Philippi, and Stéphanie Lorain, and Cyriaque Beley, and Cécile Peccate, and Guillaume Précigout, and Simone Spuler, and Luis Garcia
July 2001, Human gene therapy,
Susanne Philippi, and Stéphanie Lorain, and Cyriaque Beley, and Cécile Peccate, and Guillaume Précigout, and Simone Spuler, and Luis Garcia
September 2016, Molecular therapy. Nucleic acids,
Susanne Philippi, and Stéphanie Lorain, and Cyriaque Beley, and Cécile Peccate, and Guillaume Précigout, and Simone Spuler, and Luis Garcia
December 2005, Molecular therapy : the journal of the American Society of Gene Therapy,
Susanne Philippi, and Stéphanie Lorain, and Cyriaque Beley, and Cécile Peccate, and Guillaume Précigout, and Simone Spuler, and Luis Garcia
February 2011, Current biology : CB,
Susanne Philippi, and Stéphanie Lorain, and Cyriaque Beley, and Cécile Peccate, and Guillaume Précigout, and Simone Spuler, and Luis Garcia
December 2019, Current opinion in structural biology,
Susanne Philippi, and Stéphanie Lorain, and Cyriaque Beley, and Cécile Peccate, and Guillaume Précigout, and Simone Spuler, and Luis Garcia
July 2008, Journal of nuclear medicine : official publication, Society of Nuclear Medicine,
Susanne Philippi, and Stéphanie Lorain, and Cyriaque Beley, and Cécile Peccate, and Guillaume Précigout, and Simone Spuler, and Luis Garcia
August 2003, The Journal of clinical investigation,
Susanne Philippi, and Stéphanie Lorain, and Cyriaque Beley, and Cécile Peccate, and Guillaume Précigout, and Simone Spuler, and Luis Garcia
June 2014, Molecular therapy : the journal of the American Society of Gene Therapy,
Copied contents to your clipboard!