title: 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.
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.
authors.affiliation: Department of Medical Rehabilitation, Faculty of Rehabilitation, Kobegakuin University, 518 Arise, Ikawadani, Nishi, Kobe, 651-2180, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Nishida
authors.forename: Maki
authors.initials: M
authors.lastname: Minegishi
authors.forename: Atsuko
authors.initials: A
authors.lastname: Takeuchi
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Awano
authors.forename: Emma Tabe Eko
authors.initials: ET
authors.lastname: Niba
authors.forename: Masafumi
authors.initials: M
authors.lastname: Matsuo
mesh_terms: D017398:Alternative Splicing; D001483:Base Sequence; D045744:Cell Line, Tumor; D004251:DNA Transposable Elements / Q000235:genetics; D016189:Dystrophin / Q000235:genetics*; D005091:Exons; D023281:Genomics; D006801:Humans; D007438:Introns; D008969:Molecular Sequence Data; D020388:Muscular Dystrophy, Duchenne / Q000235:genetics; D009154:Mutation; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D022821:RNA Splice Sites / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA
chemical_list: D004251:DNA Transposable Elements; D016189:Dystrophin; D022821:RNA Splice Sites; D012333:RNA, Messenger