Neuronal SH-SY5Y cells use the C-dystrophin promoter coupled with exon 78 skipping and display multiple patterns of alternative splicing including two intronic insertion events

Hum Genet. 2015 Sep;134(9):993-1001. doi: 10.1007/s00439-015-1581-2. Epub 2015 Jul 8.

Abstract

Duchenne muscular dystrophy (DMD) is a progressive muscle wasting disease caused by mutations in the dystrophin gene. One-third of DMD cases are complicated by mental retardation. Here, we used reverse transcription PCR to analyze the pattern of dystrophin transcripts in neuronal SH-SY5Y cells. Among the three alternative promoters/first exons at the 5'-end, only transcripts containing the brain cortex-specific C1 exon could be amplified. The C-transcript appeared as two products: a major product of the expected size and a minor larger product that contained the cryptic exon 1a between exons C1 and 2. At the 3'-end there was complete exon 78 skipping. Together, these findings indicate that SH-SY5Y cells have neuron-specific characteristics with regard to both promoter activation and alternative splicing. We also revealed partial skipping of exons 9 and 71. Four amplified products were obtained from a fragment covering exons 36-41: a strong expected product, two weak products lacking either exon 37 or exon 38, and a second strong larger product with a 568-bp insertion between exons 40 and 41. The inserted sequence matched the 3'-end of intron 40 perfectly. We concluded that a cryptic splice site was activated in SH-SY5Y cells to create the novel, unusually large, exon 41e (751 bp). In total, we identified seven alternative splicing events in neuronal SH-SY5Y cells, and calculated that 32 dystrophin transcripts could be produced. Our results may provide clues in the analysis of transcriptype-phenotype correlations as regards mental retardation in DMD.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alternative Splicing*
  • Base Sequence
  • Cell Line, Tumor
  • DNA Transposable Elements / genetics
  • Dystrophin / genetics*
  • Exons*
  • Genomics
  • Humans
  • Introns*
  • Molecular Sequence Data
  • Muscular Dystrophy, Duchenne / genetics
  • Mutation
  • Polymerase Chain Reaction
  • Promoter Regions, Genetic*
  • RNA Splice Sites / genetics
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Sequence Analysis, DNA

Substances

  • DNA Transposable Elements
  • Dystrophin
  • RNA Splice Sites
  • RNA, Messenger