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Logo of jcinvestThe Journal of Clinical InvestigationCurrent IssueArchiveSubscriptionAbout the Journal
J Clin Invest. Nov 1, 1997; 100(9): 2204–2210.
PMCID: PMC508415

Disruption of the splicing enhancer sequence within exon 27 of the dystrophin gene by a nonsense mutation induces partial skipping of the exon and is responsible for Becker muscular dystrophy.


The mechanism of exon skipping induced by nonsense mutations has not been well elucidated. We now report results of in vitro splicing studies which disclosed that a particular example of exon skipping is due to disruption of a splicing enhancer sequence located within the exon. A nonsense mutation (E1211X) due to a G to T transversion at the 28th nucleotide of exon 27 (G3839T) was identified in the dystrophin gene of a Japanese Becker muscular dystrophy case. Partial skipping of the exon resulted in the production of truncated dystrophin mRNA, although the consensus sequences for splicing at both ends of exon 27 were unaltered. To determine how E1211X induced exon 27 skipping, the splicing enhancer activity of purine-rich region within exon 27 was examined in an in vitro splicing system using chimeric doublesex gene pre-mRNA. The mutant sequence containing G3839T abolished splicing enhancer activity of the wild-type purine-rich sequence for the upstream intron in this chimeric pre-mRNA. An artificial polypurine oligonucleotide mimicking the purine-rich sequence of exon 27 also showed enhancer activity that was suppressed by the introduction of a T nucleotide. Furthermore, the splicing enhancer activity was more markedly inhibited when a nonsense codon was created by the inserted T residue. This is the first evidence that partial skipping of an exon harboring a nonsense mutation is due to disruption of a splicing enhancer sequence.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Green MR. Pre-mRNA splicing. Annu Rev Genet. 1986;20:671–708. [PubMed]
  • Watakabe A, Tanaka K, Shimura Y. The role of exon sequences in splice site selection. Genes Dev. 1993 Mar;7(3):407–418. [PubMed]
  • Tanaka K, Watakabe A, Shimura Y. Polypurine sequences within a downstream exon function as a splicing enhancer. Mol Cell Biol. 1994 Feb;14(2):1347–1354. [PMC free article] [PubMed]
  • Tian M, Maniatis T. A splicing enhancer exhibits both constitutive and regulated activities. Genes Dev. 1994 Jul 15;8(14):1703–1712. [PubMed]
  • Yeakley JM, Morfin JP, Rosenfeld MG, Fu XD. A complex of nuclear proteins mediates SR protein binding to a purine-rich splicing enhancer. Proc Natl Acad Sci U S A. 1996 Jul 23;93(15):7582–7587. [PMC free article] [PubMed]
  • Tacke R, Manley JL. The human splicing factors ASF/SF2 and SC35 possess distinct, functionally significant RNA binding specificities. EMBO J. 1995 Jul 17;14(14):3540–3551. [PMC free article] [PubMed]
  • Krawczak M, Reiss J, Cooper DN. The mutational spectrum of single base-pair substitutions in mRNA splice junctions of human genes: causes and consequences. Hum Genet. 1992 Sep-Oct;90(1-2):41–54. [PubMed]
  • Dietz HC, Valle D, Francomano CA, Kendzior RJ, Jr, Pyeritz RE, Cutting GR. The skipping of constitutive exons in vivo induced by nonsense mutations. Science. 1993 Jan 29;259(5095):680–683. [PubMed]
  • Naylor JA, Green PM, Rizza CR, Giannelli F. Analysis of factor VIII mRNA reveals defects in everyone of 28 haemophilia A patients. Hum Mol Genet. 1993 Jan;2(1):11–17. [PubMed]
  • Gibson RA, Hajianpour A, Murer-Orlando M, Buchwald M, Mathew CG. A nonsense mutation and exon skipping in the Fanconi anaemia group C gene. Hum Mol Genet. 1993 Jun;2(6):797–799. [PubMed]
  • Bach G, Moskowitz SM, Tieu PT, Matynia A, Neufeld EF. Molecular analysis of Hurler syndrome in Druze and Muslim Arab patients in Israel: multiple allelic mutations of the IDUA gene in a small geographic area. Am J Hum Genet. 1993 Aug;53(2):330–338. [PMC free article] [PubMed]
  • Das S, Levinson B, Whitney S, Vulpe C, Packman S, Gitschier J. Diverse mutations in patients with Menkes disease often lead to exon skipping. Am J Hum Genet. 1994 Nov;55(5):883–889. [PMC free article] [PubMed]
  • Hull J, Shackleton S, Harris A. The stop mutation R553X in the CFTR gene results in exon skipping. Genomics. 1994 Jan 15;19(2):362–364. [PubMed]
  • Santisteban I, Arredondo-Vega FX, Kelly S, Loubser M, Meydan N, Roifman C, Howell PL, Bowen T, Weinberg KI, Schroeder ML, et al. Three new adenosine deaminase mutations that define a splicing enhancer and cause severe and partial phenotypes: implications for evolution of a CpG hotspot and expression of a transduced ADA cDNA. Hum Mol Genet. 1995 Nov;4(11):2081–2087. [PubMed]
  • Will K, Dörk T, Stuhrmann M, von der Hardt H, Ellemunter H, Tümmler B, Schmidtke J. Transcript analysis of CFTR nonsense mutations in lymphocytes and nasal epithelial cells from cystic fibrosis patients. Hum Mutat. 1995;5(3):210–220. [PubMed]
  • Okamoto Y, Yamazaki T, Katsumi A, Kojima T, Takamatsu J, Nishida M, Saito H. A novel nonsense mutation associated with an exon skipping in a patient with hereditary protein S deficiency type I. Thromb Haemost. 1996 Jun;75(6):877–882. [PubMed]
  • Bienvenu T, Beldjord C, Chelly J, Fonknechten N, Hubert D, Dusser D, Kaplan JC. Analysis of alternative splicing patterns in the cystic fibrosis transmembrane conductance regulator gene using mRNA derived from lymphoblastoid cells of cystic fibrosis patients. Eur J Hum Genet. 1996;4(3):127–134. [PubMed]
  • Dietz HC, Kendzior RJ., Jr Maintenance of an open reading frame as an additional level of scrutiny during splice site selection. Nat Genet. 1994 Oct;8(2):183–188. [PubMed]
  • Ahn AH, Kunkel LM. The structural and functional diversity of dystrophin. Nat Genet. 1993 Apr;3(4):283–291. [PubMed]
  • Nishio H, Takeshima Y, Narita N, Yanagawa H, Suzuki Y, Ishikawa Y, Ishikawa Y, Minami R, Nakamura H, Matsuo M. Identification of a novel first exon in the human dystrophin gene and of a new promoter located more than 500 kb upstream of the nearest known promoter. J Clin Invest. 1994 Sep;94(3):1037–1042. [PMC free article] [PubMed]
  • Matsuo M. Duchenne/Becker muscular dystrophy: from molecular diagnosis to gene therapy. Brain Dev. 1996 May-Jun;18(3):167–172. [PubMed]
  • Matsuo M, Masumura T, Nishio H, Nakajima T, Kitoh Y, Takumi T, Koga J, Nakamura H. Exon skipping during splicing of dystrophin mRNA precursor due to an intraexon deletion in the dystrophin gene of Duchenne muscular dystrophy kobe. J Clin Invest. 1991 Jun;87(6):2127–2131. [PMC free article] [PubMed]
  • Narita N, Nishio H, Kitoh Y, Ishikawa Y, Ishikawa Y, Minami R, Nakamura H, Matsuo M. Insertion of a 5' truncated L1 element into the 3' end of exon 44 of the dystrophin gene resulted in skipping of the exon during splicing in a case of Duchenne muscular dystrophy. J Clin Invest. 1993 May;91(5):1862–1867. [PMC free article] [PubMed]
  • Takeshima Y, Nishio H, Sakamoto H, Nakamura H, Matsuo M. Modulation of in vitro splicing of the upstream intron by modifying an intra-exon sequence which is deleted from the dystrophin gene in dystrophin Kobe. J Clin Invest. 1995 Feb;95(2):515–520. [PMC free article] [PubMed]
  • Pramono ZA, Takeshima Y, Alimsardjono H, Ishii A, Takeda S, Matsuo M. Induction of exon skipping of the dystrophin transcript in lymphoblastoid cells by transfecting an antisense oligodeoxynucleotide complementary to an exon recognition sequence. Biochem Biophys Res Commun. 1996 Sep 13;226(2):445–449. [PubMed]
  • Beggs AH, Koenig M, Boyce FM, Kunkel LM. Detection of 98% of DMD/BMD gene deletions by polymerase chain reaction. Hum Genet. 1990 Nov;86(1):45–48. [PubMed]
  • Boyce FM, Beggs AH, Feener C, Kunkel LM. Dystrophin is transcribed in brain from a distant upstream promoter. Proc Natl Acad Sci U S A. 1991 Feb 15;88(4):1276–1280. [PMC free article] [PubMed]
  • Roberts RG, Bobrow M, Bentley DR. Point mutations in the dystrophin gene. Proc Natl Acad Sci U S A. 1992 Mar 15;89(6):2331–2335. [PMC free article] [PubMed]
  • Hagiwara Y, Nishio H, Kitoh Y, Takeshima Y, Narita N, Wada H, Yokoyama M, Nakamura H, Matsuo M. A novel point mutation (G-1 to T) in a 5' splice donor site of intron 13 of the dystrophin gene results in exon skipping and is responsible for Becker muscular dystrophy. Am J Hum Genet. 1994 Jan;54(1):53–61. [PMC free article] [PubMed]
  • Isegawa Y, Sheng J, Sokawa Y, Yamanishi K, Nakagomi O, Ueda S. Selective amplification of cDNA sequence from total RNA by cassette-ligation mediated polymerase chain reaction (PCR): application to sequencing 6.5 kb genome segment of hantavirus strain B-1. Mol Cell Probes. 1992 Dec;6(6):467–475. [PubMed]
  • Lynch KW, Maniatis T. Synergistic interactions between two distinct elements of a regulated splicing enhancer. Genes Dev. 1995 Feb 1;9(3):284–293. [PubMed]
  • Maquat LE. When cells stop making sense: effects of nonsense codons on RNA metabolism in vertebrate cells. RNA. 1995 Jul;1(5):453–465. [PMC free article] [PubMed]
  • Steingrimsdottir H, Rowley G, Dorado G, Cole J, Lehmann AR. Mutations which alter splicing in the human hypoxanthine-guanine phosphoribosyltransferase gene. Nucleic Acids Res. 1992 Mar 25;20(6):1201–1208. [PMC free article] [PubMed]
  • Jin Y, Dietz HC, Montgomery RA, Bell WR, McIntosh I, Coller B, Bray PF. Glanzmann thrombasthenia. Cooperation between sequence variants in cis during splice site selection. J Clin Invest. 1996 Oct 15;98(8):1745–1754. [PMC free article] [PubMed]
  • Willing MC, Deschenes SP, Slayton RL, Roberts EJ. Premature chain termination is a unifying mechanism for COL1A1 null alleles in osteogenesis imperfecta type I cell strains. Am J Hum Genet. 1996 Oct;59(4):799–809. [PMC free article] [PubMed]
  • Xu R, Teng J, Cooper TA. The cardiac troponin T alternative exon contains a novel purine-rich positive splicing element. Mol Cell Biol. 1993 Jun;13(6):3660–3674. [PMC free article] [PubMed]
  • Morisaki H, Morisaki T, Newby LK, Holmes EW. Alternative splicing: a mechanism for phenotypic rescue of a common inherited defect. J Clin Invest. 1993 May;91(5):2275–2280. [PMC free article] [PubMed]
  • Monaco AP, Bertelson CJ, Liechti-Gallati S, Moser H, Kunkel LM. An explanation for the phenotypic differences between patients bearing partial deletions of the DMD locus. Genomics. 1988 Jan;2(1):90–95. [PubMed]
  • Prior TW, Bartolo C, Papp AC, Snyder PJ, Sedra MS, Burghes AH, Mendell JR. Nonsense mutations in a Becker muscular dystrophy and an intermediate patient. Hum Mutat. 1996;7(1):72–75. [PubMed]
  • Barbieri AM, Soriani N, Ferlini A, Michelato A, Ferrari M, Carrera P. Seven novel additional small mutations and a new alternative splicing in the human dystrophin gene detected by heteroduplex analysis and restricted RT-PCR heteroduplex analysis of illegitimate transcripts. Eur J Hum Genet. 1996;4(3):183–187. [PubMed]
  • Shapiro MB, Senapathy P. RNA splice junctions of different classes of eukaryotes: sequence statistics and functional implications in gene expression. Nucleic Acids Res. 1987 Sep 11;15(17):7155–7174. [PMC free article] [PubMed]
  • Koenig M, Monaco AP, Kunkel LM. The complete sequence of dystrophin predicts a rod-shaped cytoskeletal protein. Cell. 1988 Apr 22;53(2):219–228. [PubMed]

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