Résumé :
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Spinal muscular atrophy (SMA) is an autosomal recessive neuromuscular disorder characterized by degeneration of alpha motor neurons in the anterior horn cells of the spinal cord, often appearing during early childhood. This disease results in most cases from large homozygous deletions in SMN1 gene or, in rare cases, from hemizygous SMN1 intragenic subtle mutations. In this study we present the identification and characterization of a new SMA-causing mutation: c.835-3C>T. Located on the intron 6 of SMN1, we identified this variation in the single SMN1 copy of a patient with type IV SMA. This patient carries only 2 copies of SMN2, the partially functional SMN1 paralog. Because of its location within the SMN1 exon 7 acceptor site and despite the absence of strong prediction of splicing alteration using in silico tools, we raised the hypothesis that this variation could alter SMN1 exon 7 splicing. We demonstrated both ex vivo, using SMN minigene splicing assays, and in vivo, by a comparative multiplex RT-PCR analysis of the patient RNA, that the c.835-3C>T variation induces a modest but unequivocal exclusion of exon 7 from SMN1 mRNA. We suggest that this splicing alteration could be the consequence of the creation of a high-affinity binding site for the splicing repressor protein hnRNP A1 overlapping the splice acceptor site of exon 7. This "leaky" splicing mutation explains the exceptionnally mild phenotype of an 70-year-old ambulatory SMA patient who carries only two copies of the SMN2 gene. Importantly, our study supports the current therapeutic strategies aiming at improving exon 7 inclusion in SMN transcripts, and bring new clues about the level of inclusion required to achieve an effect at the therapeutic scale.
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