• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of molcellbPermissionsJournals.ASM.orgJournalMCB ArticleJournal InfoAuthorsReviewers
Mol Cell Biol. Feb 1994; 14(2): 1347–1354.
PMCID: PMC358489

Polypurine sequences within a downstream exon function as a splicing enhancer.

Abstract

We have previously shown that a purine-rich sequence located within exon M2 of the mouse immunoglobulin mu gene functions as a splicing enhancer, as judged by its ability to stimulate splicing of a distant upstream intron. This sequence element has been designated ERS (exon recognition sequence). In this study, we investigated the stimulatory effects of various ERS-like sequences, using the in vitro splicing system with HeLa cell nuclear extracts. Here, we show that purine-rich sequences of several natural exons that have previously been shown to be required for splicing function as a splicing enhancer like the ERS of the immunoglobulin mu gene. Moreover, even synthetic polypurine sequences had stimulatory effects on the upstream splicing. Evaluation of the data obtained from the analyses of both natural and synthetic purine-rich sequences shows that (i) alternating purine sequences can stimulate splicing, while poly(A) or poly(G) sequences cannot, and (ii) the presence of U residues within the polypurine sequence greatly reduces the level of stimulation. Competition experiments strongly suggest that the stimulatory effects of various purine-rich sequences are mediated by the same trans-acting factor(s). We conclude from these results that the purine-rich sequences that we examined in this study also represent examples of ERS. Thus, ERS is considered a general splicing element that is present in various exons and plays an important role in splice site selection.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.9M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Black DL. Does steric interference between splice sites block the splicing of a short c-src neuron-specific exon in non-neuronal cells? Genes Dev. 1991 Mar;5(3):389–402. [PubMed]
  • Brunak S, Engelbrecht J, Knudsen S. Prediction of human mRNA donor and acceptor sites from the DNA sequence. J Mol Biol. 1991 Jul 5;220(1):49–65. [PubMed]
  • Burtis KC, Baker BS. Drosophila doublesex gene controls somatic sexual differentiation by producing alternatively spliced mRNAs encoding related sex-specific polypeptides. Cell. 1989 Mar 24;56(6):997–1010. [PubMed]
  • Cooper TA. In vitro splicing of cardiac troponin T precursors. Exon mutations disrupt splicing of the upstream intron. J Biol Chem. 1992 Mar 15;267(8):5330–5338. [PubMed]
  • Cooper TA, Ordahl CP. Nucleotide substitutions within the cardiac troponin T alternative exon disrupt pre-mRNA alternative splicing. Nucleic Acids Res. 1989 Oct 11;17(19):7905–7921. [PMC free article] [PubMed]
  • Cote GJ, Stolow DT, Peleg S, Berget SM, Gagel RF. Identification of exon sequences and an exon binding protein involved in alternative RNA splicing of calcitonin/CGRP. Nucleic Acids Res. 1992 May 11;20(9):2361–2366. [PMC free article] [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]
  • Dignam JD, Lebovitz RM, Roeder RG. Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei. Nucleic Acids Res. 1983 Mar 11;11(5):1475–1489. [PMC free article] [PubMed]
  • Dominski Z, Kole R. Selection of splice sites in pre-mRNAs with short internal exons. Mol Cell Biol. 1991 Dec;11(12):6075–6083. [PMC free article] [PubMed]
  • Eperon LP, Graham IR, Griffiths AD, Eperon IC. Effects of RNA secondary structure on alternative splicing of pre-mRNA: is folding limited to a region behind the transcribing RNA polymerase? Cell. 1988 Jul 29;54(3):393–401. [PubMed]
  • Estes PA, Cooke NE, Liebhaber SA. A native RNA secondary structure controls alternative splice-site selection and generates two human growth hormone isoforms. J Biol Chem. 1992 Jul 25;267(21):14902–14908. [PubMed]
  • Fu XD, Katz RA, Skalka AM, Maniatis T. The role of branchpoint and 3'-exon sequences in the control of balanced splicing of avian retrovirus RNA. Genes Dev. 1991 Feb;5(2):211–220. [PubMed]
  • Furdon PJ, Kole R. The length of the downstream exon and the substitution of specific sequences affect pre-mRNA splicing in vitro. Mol Cell Biol. 1988 Feb;8(2):860–866. [PMC free article] [PubMed]
  • Green MR. Biochemical mechanisms of constitutive and regulated pre-mRNA splicing. Annu Rev Cell Biol. 1991;7:559–599. [PubMed]
  • Hampson RK, La Follette L, Rottman FM. Alternative processing of bovine growth hormone mRNA is influenced by downstream exon sequences. Mol Cell Biol. 1989 Apr;9(4):1604–1610. [PMC free article] [PubMed]
  • Helfman DM, Ricci WM, Finn LA. Alternative splicing of tropomyosin pre-mRNAs in vitro and in vivo. Genes Dev. 1988 Dec;2(12A):1627–1638. [PubMed]
  • Hoffman BE, Grabowski PJ. U1 snRNP targets an essential splicing factor, U2AF65, to the 3' splice site by a network of interactions spanning the exon. Genes Dev. 1992 Dec;6(12B):2554–2568. [PubMed]
  • Hoshijima K, Inoue K, Higuchi I, Sakamoto H, Shimura Y. Control of doublesex alternative splicing by transformer and transformer-2 in Drosophila. Science. 1991 May 10;252(5007):833–836. [PubMed]
  • Jackson IJ. A reappraisal of non-consensus mRNA splice sites. Nucleic Acids Res. 1991 Jul 25;19(14):3795–3798. [PMC free article] [PubMed]
  • Kakizuka A, Ingi T, Murai T, Nakanishi S. A set of U1 snRNA-complementary sequences involved in governing alternative RNA splicing of the kininogen genes. J Biol Chem. 1990 Jun 15;265(17):10102–10108. [PubMed]
  • Katz RA, Skalka AM. Control of retroviral RNA splicing through maintenance of suboptimal processing signals. Mol Cell Biol. 1990 Feb;10(2):696–704. [PMC free article] [PubMed]
  • Kuo HC, Nasim FH, Grabowski PJ. Control of alternative splicing by the differential binding of U1 small nuclear ribonucleoprotein particle. Science. 1991 Mar 1;251(4997):1045–1050. [PubMed]
  • Libri D, Goux-Pelletan M, Brody E, Fiszman MY. Exon as well as intron sequences are cis-regulating elements for the mutually exclusive alternative splicing of the beta tropomyosin gene. Mol Cell Biol. 1990 Oct;10(10):5036–5046. [PMC free article] [PubMed]
  • Libri D, Piseri A, Fiszman MY. Tissue-specific splicing in vivo of the beta-tropomyosin gene: dependence on an RNA secondary structure. Science. 1991 Jun 28;252(5014):1842–1845. [PubMed]
  • Ligtenberg MJ, Gennissen AM, Vos HL, Hilkens J. A single nucleotide polymorphism in an exon dictates allele dependent differential splicing of episialin mRNA. Nucleic Acids Res. 1991 Jan 25;19(2):297–301. [PMC free article] [PubMed]
  • Mardon HJ, Sebastio G, Baralle FE. A role for exon sequences in alternative splicing of the human fibronectin gene. Nucleic Acids Res. 1987 Oct 12;15(19):7725–7733. [PMC free article] [PubMed]
  • Naeger LK, Schoborg RV, Zhao Q, Tullis GE, Pintel DJ. Nonsense mutations inhibit splicing of MVM RNA in cis when they interrupt the reading frame of either exon of the final spliced product. Genes Dev. 1992 Jun;6(6):1107–1119. [PubMed]
  • Nelson KK, Green MR. Splice site selection and ribonucleoprotein complex assembly during in vitro pre-mRNA splicing. Genes Dev. 1988 Mar;2(3):319–329. [PubMed]
  • Nelson KK, Green MR. Mechanism for cryptic splice site activation during pre-mRNA splicing. Proc Natl Acad Sci U S A. 1990 Aug;87(16):6253–6257. [PMC free article] [PubMed]
  • Ohshima Y, Gotoh Y. Signals for the selection of a splice site in pre-mRNA. Computer analysis of splice junction sequences and like sequences. J Mol Biol. 1987 May 20;195(2):247–259. [PubMed]
  • Reed R, Maniatis T. A role for exon sequences and splice-site proximity in splice-site selection. Cell. 1986 Aug 29;46(5):681–690. [PubMed]
  • Ricketts MH, Simons MJ, Parma J, Mercken L, Dong Q, Vassart G. A nonsense mutation causes hereditary goitre in the Afrikander cattle and unmasks alternative splicing of thyroglobulin transcripts. Proc Natl Acad Sci U S A. 1987 May;84(10):3181–3184. [PMC free article] [PubMed]
  • Robberson BL, Cote GJ, Berget SM. Exon definition may facilitate splice site selection in RNAs with multiple exons. Mol Cell Biol. 1990 Jan;10(1):84–94. [PMC free article] [PubMed]
  • Sakamoto H, Ohno M, Yasuda K, Mizumoto K, Shimura Y. In vitro splicing of a chicken delta-crystallin pre-mRNA in a mammalian nuclear extract. J Biochem. 1987 Nov;102(5):1289–1301. [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]
  • Solnick D. Alternative splicing caused by RNA secondary structure. Cell. 1985 Dec;43(3 Pt 2):667–676. [PubMed]
  • Solnick D, Lee SI. Amount of RNA secondary structure required to induce an alternative splice. Mol Cell Biol. 1987 Sep;7(9):3194–3198. [PMC free article] [PubMed]
  • Somasekhar MB, Mertz JE. Exon mutations that affect the choice of splice sites used in processing the SV40 late transcripts. Nucleic Acids Res. 1985 Aug 12;13(15):5591–5609. [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]
  • Tacke R, Goridis C. Alternative splicing in the neural cell adhesion molecule pre-mRNA: regulation of exon 18 skipping depends on the 5'-splice site. Genes Dev. 1991 Aug;5(8):1416–1429. [PubMed]
  • Talerico M, Berget SM. Effect of 5' splice site mutations on splicing of the preceding intron. Mol Cell Biol. 1990 Dec;10(12):6299–6305. [PMC free article] [PubMed]
  • Tian M, Maniatis T. Positive control of pre-mRNA splicing in vitro. Science. 1992 Apr 10;256(5054):237–240. [PubMed]
  • Wakamatsu N, Kobayashi H, Miyatake T, Tsuji S. A novel exon mutation in the human beta-hexosaminidase beta subunit gene affects 3' splice site selection. J Biol Chem. 1992 Feb 5;267(4):2406–2413. [PubMed]
  • Watakabe A, Inoue K, Sakamoto H, Shimura Y. A secondary structure at the 3' splice site affects the in vitro splicing reaction of mouse immunoglobulin mu chain pre-mRNAs. Nucleic Acids Res. 1989 Oct 25;17(20):8159–8169. [PMC free article] [PubMed]
  • Watakabe A, Sakamoto H, Shimura Y. Repositioning of an alternative exon sequence of mouse IgM pre-mRNA activates splicing of the preceding intron. Gene Expr. 1991;1(3):175–184. [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]
  • 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]
  • Zamore PD, Patton JG, Green MR. Cloning and domain structure of the mammalian splicing factor U2AF. Nature. 1992 Feb 13;355(6361):609–614. [PubMed]

Articles from Molecular and Cellular Biology are provided here courtesy of American Society for Microbiology (ASM)

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

  • MedGen
    MedGen
    Related information in MedGen
  • PubMed
    PubMed
    PubMed citations for these articles
  • Substance
    Substance
    PubChem Substance links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...