A splicing silencer in SMN2 intron 6 is critical in spinal muscular atrophy

Hum Mol Genet. 2023 Mar 6;32(6):971-983. doi: 10.1093/hmg/ddac260.


Spinal muscular atrophy (SMA) is a fatal neuromuscular disease caused by homozygous deletions or mutations of the SMN1 gene. SMN2 is a paralogous gene of SMN1 and a modifying gene of SMA. A better understanding of how SMN2 exon 7 splicing is regulated helps discover new therapeutic targets for SMA therapy. Based on an antisense walk method to map exonic and intronic splicing silencers (ESSs and ISSs) in SMN2 exon 7 and the proximal regions of its flanking introns, we identified one ISS (ISS6-KH) at upstream of the branch point site in intron 6. By using mutagenesis-coupled RT-PCR with SMN1/2 minigenes, immunochromatography, overexpression and siRNA-knockdown, we found this ISS consists of a bipartite hnRNP A1 binding cis-element and a poly-U sequence located between the proximal hnRNP A1 binding site (UAGCUA) and the branch site. Both HuR and hnRNP C1 proteins promote exon 7 skipping through the poly-U stretch. Mutations or deletions of these motifs lead to efficient SMN2 exon 7 inclusion comparable to SMN1 gene. Furthermore, we identified an optimal antisense oligonucleotide that binds the intron six ISS and causes striking exon 7 inclusion in the SMN2 gene in patient fibroblasts and SMA mouse model. Our findings demonstrate that this novel ISS plays an important role in SMN2 exon 7 skipping and highlight a new therapeutic target for SMA therapy.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Heterogeneous Nuclear Ribonucleoprotein A1 / genetics
  • Heterogeneous Nuclear Ribonucleoprotein A1 / metabolism
  • Introns / genetics
  • Mice
  • Muscular Atrophy, Spinal* / genetics
  • Muscular Atrophy, Spinal* / therapy
  • RNA Splicing / genetics
  • RNA-Binding Proteins* / genetics
  • RNA-Binding Proteins* / metabolism


  • RNA-Binding Proteins
  • Heterogeneous Nuclear Ribonucleoprotein A1