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Lessons from non-canonical splicing

Nature Reviews Genetics volume 17, pages 407–421 (2016)Cite this article

Subjects

Key Points

  • The development of new methods for preparing and sequencing RNA sequencing libraries, as well as new alignment algorithms, have revealed many thousands of previously unknown non-canonical splicing events.

  • Non-canonical splicing events are often tissue-specific and are particularly enriched in the central nervous system, thereby increasing proteome diversity or regulating gene expression.

  • Cryptic exons, microexons and recursive splice sites often require unconventional exon definition mechanisms.

  • Other non-canonical splicing events result from lower or higher splicing efficiency than normal (such as retained introns and exonic introns), changes in the usual order of splicing (circular RNAs and chimeric RNAs) or changes in the consensus sequence (atypical splice sites).

  • Transposable elements are a rich source of newly emerging cryptic exons, which can contribute to the evolution of gene regulatory networks.

  • Mutations that perturb functionally important non-canonical splicing events, or strongly increase the recognition of cryptic splice sites, can cause numerous diseases.

  • Non-canonical splicing mechanisms offer new therapeutic opportunities to treat disease.

Abstract

Recent improvements in experimental and computational techniques that are used to study the transcriptome have enabled an unprecedented view of RNA processing, revealing many previously unknown non-canonical splicing events. This includes cryptic events located far from the currently annotated exons and unconventional splicing mechanisms that have important roles in regulating gene expression. These non-canonical splicing events are a major source of newly emerging transcripts during evolution, especially when they involve sequences derived from transposable elements. They are therefore under precise regulation and quality control, which minimizes their potential to disrupt gene expression. We explain how non-canonical splicing can lead to aberrant transcripts that cause many diseases, and also how it can be exploited for new therapeutic strategies.

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Figure 1: Cryptic exons and microexons.
Figure 2: Recursive splicing of long introns.
Figure 3: Intron retention and exitrons.
Figure 4: Formation of circularRNAs and chimeric transcripts.
Figure 5: A summary of human splice site DNA consensus motifs.
Figure 6: Cryptic splicing in disease and therapeutic strategies.

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References

  1. Raj, B. & Blencowe, B. J. Alternative splicing in the mammalian nervous system: recent insights into mechanisms and functional roles. Neuron 87, 14–27 (2015).

    Article  CAS  PubMed  Google Scholar 

  2. Fu, X. D. & Ares, M. Jr. Context-dependent control of alternative splicing by RNA-binding proteins. Nat. Rev. Genet. 15, 689–701 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Derrien, T. et al. The GENCODE v7 catalog of human long noncoding RNAs: analysis of their gene structure, evolution, and expression. Genome Res. 22, 1775–1789 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Matera, A. G. & Wang, Z. A day in the life of the spliceosome. Nat. Rev. Mol. Cell Biol. 15, 108–121 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Scotti, M. M. & Swanson, M. S. RNA mis-splicing in disease. Nat. Rev. Genet. 17, 19–32 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  6. Jangi, M., Boutz, P. L., Paul, P. & Sharp, P. A. Rbfox2 controls autoregulation in RNA-binding protein networks. Genes Dev. 28, 637–651 (2014). In this study, RBFOX2 is found to cross-regulate conserved NMD exons within transcripts that encode nearly 70 RBPs, thus forming a broad auto- and cross-regulatory splicing network for fine-tuning the expression levels of RBPs.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Eom, T. et al. NOVA-dependent regulation of cryptic NMD exons controls synaptic protein levels after seizure. eLife 2, e00178 (2013). In this study, NOVA proteins are found to regulate the splicing of many cryptic NMD exons, which mediates the regulation of transcripts encoding synaptic proteins in response to excitation, indicating its role in the homeostasis of synaptic activity.

    Article  PubMed  PubMed Central  Google Scholar 

  8. Zarnack, K. et al. Direct competition between hnRNP C and U2AF65 protects the transcriptome from the exonization of Alu elements. Cell 152, 453–466 (2013). In this paper, U tracts within thousands of antisense Alu elements are found to act as a platform for competition between HNRNPC and U2AF2, and therefore the length of the U tract affects splicing efficiency of cryptic Alu exons.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Ling, J. P., Pletnikova, O., Troncoso, J. C. & Wong, P. C. TDP-43 repression of nonconserved cryptic exons is compromised in ALS-FTD. Science 349, 650–655 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Yan, Q. et al. Systematic discovery of regulated and conserved alternative exons in the mammalian brain reveals NMD modulating chromatin regulators. Proc. Natl Acad. Sci. USA 112, 3445–3450 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Wu, J., Anczukow, O., Krainer, A. R., Zhang, M. Q. & Zhang, C. OLego: fast and sensitive mapping of spliced mRNA-Seq reads using small seeds. Nucleic Acids Res. 41, 5149–5163 (2013). This paper describes a sequence mapping algorithm that incorporates short seed mapping and identified >500,000 non-canonical splicing events, including microexons, in humans and mice.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Irimia, M. et al. A highly conserved program of neuronal microexons is misregulated in autistic brains. Cell 159, 1511–1523 (2014). In this study, a new strategy to map RNA-seq data identified more than 400 ORF-modifying microexons, many of which are regulated by SRMM4 and are differentially spliced in patients with ASD.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Li, Y. I., Sanchez-Pulido, L., Haerty, W. & Ponting, C. P. RBFOX and PTBP1 proteins regulate the alternative splicing of micro-exons in human brain transcripts. Genome Res. 25, 1–13 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  14. Sibley, C. R. et al. Recursive splicing in long vertebrate genes. Nature 521, 371–375 (2015). This paper describes the identification of highly conserved vertebrate RS sites within extremely long introns that require an unusual exon definition mechanism for their splicing. Genes with such long introns are found to be more highly expressed in the brain.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Duff, M. O. et al. Genome-wide identification of zero nucleotide recursive splicing in Drosophila. Nature 521, 376–379 (2015). This paper identifies197 RS sites in Drosophila , one of which is also found in an orthologous human gene.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Hansen, T. B. et al. Natural RNA circles function as efficient microRNA sponges. Nature 495, 384–388 (2013). This study identifies a brain-enriched circRNA that acts as a miR-7 sponge and a testes-specific circRNA acting as a miR-138 sponge.

    Article  CAS  PubMed  Google Scholar 

  17. Memczak, S. et al. Circular RNAs are a large class of animal RNAs with regulatory potency. Nature 495, 333–338 (2013). This paper reports the discovery of thousands of new circRNAs across multiple tissues and species, including a circRNA that acts as a miR-7 sponge.

    Article  CAS  PubMed  Google Scholar 

  18. Salzman, J., Gawad, C., Wang, P. L., Lacayo, N. & Brown, P. O. Circular RNAs are the predominant transcript isoform from hundreds of human genes in diverse cell types. PLoS ONE 7, e30733 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Danan, M., Schwartz, S., Edelheit, S. & Sorek, R. Transcriptome-wide discovery of circular RNAs in Archaea. Nucleic Acids Res. 40, 3131–3142 (2012).

    Article  CAS  PubMed  Google Scholar 

  20. Chen, L. L. The biogenesis and emerging roles of circular RNAs. Nat. Rev. Mol. Cell Biol. 17, 205–211 (2016).

    Article  CAS  PubMed  Google Scholar 

  21. Braunschweig, U. et al. Widespread intron retention in mammals functionally tunes transcriptomes. Genome Res. 24, 1774–1786 (2014). This paper reports that intron retention can be detected in three-quarters of human and mouse multiexonic genes; it is often coupled to RNA polymerase II stalling to suppress inappropriately expressed transcripts.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Yap, K., Lim, Z. Q., Khandelia, P., Friedman, B. & Makeyev, E. V. Coordinated regulation of neuronal mRNA steady-state levels through developmentally controlled intron retention. Genes Dev. 26, 1209–1223 (2012). This study shows that PTBP1 regulates the expression of at least four neuron-specific genes by inhibiting splicing of 3′ terminal introns in non-neuronal cells, which promotes nuclear retention and degradation of the resulting transcripts.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Boutz, P. L., Bhutkar, A. & Sharp, P. A. Detained introns are a novel, widespread class of post-transcriptionally spliced introns. Genes Dev. 29, 63–80 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  24. Wong, J. J. et al. Orchestrated intron retention regulates normal granulocyte differentiation. Cell 154, 583–595 (2013). This paper reports that coordinated intron retention in 86 functionally related genes is used to regulate gene expression during granulopoiesis.

    Article  CAS  PubMed  Google Scholar 

  25. Marquez, Y., Brown, J. W., Simpson, C., Barta, A. & Kalyna, M. Transcriptome survey reveals increased complexity of the alternative splicing landscape in Arabidopsis. Genome Res. 22, 1184–1195 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Marquez, Y., Hopfler, M., Ayatollahi, Z., Barta, A. & Kalyna, M. Unmasking alternative splicing inside protein-coding exons defines exitrons and their role in proteome plasticity. Genome Res. 25, 995–1007 (2015). This paper reports the discovery of 923 alternative introns inside annotated human protein-coding exons, which are referred to as exitrons.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. De Conti, L., Baralle, M. & Buratti, E. Exon and intron definition in pre-mRNA splicing. Wiley Interdiscip. Rev. RNA 4, 49–60 (2013).

    CAS  PubMed  Google Scholar 

  28. Robberson, B. L., Cote, G. J. & Berget, S. M. Exon definition may facilitate splice site selection in RNAs with multiple exons. Mol. Cell. Biol. 10, 84–94 (1990).

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Trapnell, C., Pachter, L. & Salzberg, S. L. TopHat: discovering splice junctions with RNA-Seq. Bioinformatics 25, 1105–1111 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Dobin, A. et al. STAR: ultrafast universal RNA-seq aligner. Bioinformatics 29, 15–21 (2013).

    Article  CAS  PubMed  Google Scholar 

  31. Kelly, S. et al. Splicing of many human genes involves sites embedded within introns. Nucleic Acids Res. 43, 4721–4732 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Kapustin, Y. et al. Cryptic splice sites and split genes. Nucleic Acids Res. 39, 5837–5844 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Ni, J. Z. et al. Ultraconserved elements are associated with homeostatic control of splicing regulators by alternative splicing and nonsense-mediated decay. Genes Dev. 21, 708–718 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Lareau, L. F., Inada, M., Green, R. E., Wengrod, J. C. & Brenner, S. E. Unproductive splicing of SR genes associated with highly conserved and ultraconserved DNA elements. Nature 446, 926–929 (2007).

    Article  CAS  PubMed  Google Scholar 

  35. Jangi, M. & Sharp, P. A. Building robust transcriptomes with master splicing factors. Cell 159, 487–498 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Vaz-Drago, R. et al. Transcription-coupled RNA surveillance in human genetic diseases caused by splice site mutations. Hum. Mol. Genet. 24, 2784–2795 (2015).

    Article  CAS  PubMed  Google Scholar 

  37. Keren, H., Lev-Maor, G. & Ast, G. Alternative splicing and evolution: diversification, exon definition and function. Nat. Rev. Genet. 11, 345–355 (2010).

    Article  CAS  PubMed  Google Scholar 

  38. Quentin, Y. Origin of the Alu family: a family of Alu-like monomers gave birth to the left and the right arms of the Alu elements. Nucleic Acids Res. 20, 3397–3401 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Gal-Mark, N., Schwartz, S., Ram, O., Eyras, E. & Ast, G. The pivotal roles of TIA proteins in 5′ splice-site selection of Alu exons and across evolution. PLoS Genet. 5, e1000717 (2009).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  40. Konig, J., Zarnack, K., Luscombe, N. M. & Ule, J. Protein–RNA interactions: new genomic technologies and perspectives. Nat. Rev. Genet. 13, 77–83 (2011).

    Article  CAS  Google Scholar 

  41. Corvelo, A. & Eyras, E. Exon creation and establishment in human genes. Genome Biol. 9, R141 (2008).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  42. Dominski, Z. & Kole, R. Selection of splice sites in pre-mRNAs with short internal exons. Mol. Cell. Biol. 11, 6075–6083 (1991).

    CAS  PubMed  PubMed Central  Google Scholar 

  43. Black, D. L. Does steric interference between splice sites block the splicing of a short c-src neuron-specific exon in non-neuronal cells? Genes Dev. 5, 389–402 (1991).

    Article  CAS  PubMed  Google Scholar 

  44. Volfovsky, N., Haas, B. J. & Salzberg, S. L. Computational discovery of internal micro-exons. Genome Res. 13, 1216–1221 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Burnette, J. M., Miyamoto-Sato, E., Schaub, M. A., Conklin, J. & Lopez, A. J. Subdivision of large introns in Drosophila by recursive splicing at nonexonic elements. Genetics 170, 661–674 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Hatton, A. R., Subramaniam, V. & Lopez, A. J. Generation of alternative Ultrabithorax isoforms and stepwise removal of a large intron by resplicing at exon–exon junctions. Mol. Cell 2, 787–796 (1998).

    Article  CAS  PubMed  Google Scholar 

  47. Herzel, L. & Neugebauer, K. M. Quantification of co-transcriptional splicing from RNA-Seq data. Methods, 85, 36–43 (2015).

    Article  CAS  PubMed  Google Scholar 

  48. Ameur, A. et al. Total RNA sequencing reveals nascent transcription and widespread co-transcriptional splicing in the human brain. Nat. Struct. Mol. Biol. 18, 1435–1440 (2011).

    Article  CAS  PubMed  Google Scholar 

  49. Parra, M. K., Tan, J. S., Mohandas, N. & Conboy, J. G. Intrasplicing coordinates alternative first exons with alternative splicing in the protein 4.1R gene. EMBO J. 27, 122–131 (2008).

    Article  CAS  PubMed  Google Scholar 

  50. Ner-Gaon, H. et al. Intron retention is a major phenomenon in alternative splicing in Arabidopsis. Plant J. 39, 877–885 (2004).

    Article  CAS  PubMed  Google Scholar 

  51. Galante, P. A., Sakabe, N. J., Kirschbaum-Slager, N. & de Souza, S. J. Detection and evaluation of intron retention events in the human transcriptome. RNA 10, 757–765 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Kan, Z., States, D. & Gish, W. Selecting for functional alternative splices in ESTs. Genome Res. 12, 1837–1845 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Sakabe, N. J. & de Souza, S. J. Sequence features responsible for intron retention in human. BMC Genom. 8, 59 (2007).

    Article  CAS  Google Scholar 

  54. Martinez-Contreras, R. et al. Intronic binding sites for hnRNP A/B and hnRNP F/H proteins stimulate pre-mRNA splicing. PLoS Biol. 4, e21 (2006).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  55. Wickramasinghe, V. O. et al. Regulation of constitutive and alternative mRNA splicing across the human transcriptome by PRPF8 is determined by 5′ splice site strength. Genome Biol. 16, 201 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  56. Marinescu, V., Loomis, P. A., Ehmann, S., Beales, M. & Potashkin, J. A. Regulation of retention of FosB intron 4 by PTB. PLoS ONE 2, e828 (2007).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  57. Bergeron, D., Pal, G., Beaulieu, Y. B., Chabot, B. & Bachand, F. Regulated intron retention and nuclear pre-mRNA decay contribute to PABPN1 autoregulation. Mol. Cell. Biol. 35, 2503–2517 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Malone, C. D. et al. The exon junction complex controls transposable element activity by ensuring faithful splicing of the piwi transcript. Genes Dev. 28, 1786–1799 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  59. Hayashi, R., Handler, D., Ish-Horowicz, D. & Brennecke, J. The exon junction complex is required for definition and excision of neighboring introns in Drosophila. Genes Dev. 28, 1772–1785 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  60. Wang, Z., Murigneux, V. & Le Hir, H. Transcriptome-wide modulation of splicing by the exon junction complex. Genome Biol. 15, 551 (2014).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  61. Nigro, J. M. et al. Scrambled exons. Cell 64, 607–613 (1991).

    Article  CAS  PubMed  Google Scholar 

  62. Schindewolf, C., Braun, S. & Domdey, H. In vitro generation of a circular exon from a linear pre-mRNA transcript. Nucleic Acids Res. 24, 1260–1266 (1996).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  63. Pasman, Z., Been, M. D. & Garcia-Blanco, M. A. Exon circularization in mammalian nuclear extracts. RNA 2, 603–610 (1996).

    CAS  PubMed  PubMed Central  Google Scholar 

  64. Braun, S., Domdey, H. & Wiebauer, K. Inverse splicing of a discontinuous pre-mRNA intron generates a circular exon in a HeLa cell nuclear extract. Nucleic Acids Res. 24, 4152–4157 (1996).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  65. Suzuki, H. et al. Characterization of RNase R-digested cellular RNA source that consists of lariat and circular RNAs from pre-mRNA splicing. Nucleic Acids Res. 34, e63 (2006).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  66. Guo, J. U., Agarwal, V., Guo, H. & Bartel, D. P. Expanded identification and characterization of mammalian circular RNAs. Genome Biol. 15, 409 (2014).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  67. Jeck, W. R. & Sharpless, N. E. Detecting and characterizing circular RNAs. Nat. Biotechnol. 32, 453–461 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  68. You, X. et al. Neural circular RNAs are derived from synaptic genes and regulated by development and plasticity. Nat. Neurosci. 18, 603–610 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  69. Liang, D. & Wilusz, J. E. Short intronic repeat sequences facilitate circular RNA production. Genes Dev. 28, 2233–2247 (2014).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  70. Jeck, W. R. et al. Circular RNAs are abundant, conserved, and associated with ALU repeats. RNA 19, 141–157 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  71. Kramer, M. C. et al. Combinatorial control of Drosophila circular RNA expression by intronic repeats, hnRNPs, and SR proteins. Genes Dev. 29, 2168–2182 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  72. Zhang, X. O. et al. Complementary sequence-mediated exon circularization. Cell 159, 134–147 (2014).

    Article  CAS  PubMed  Google Scholar 

  73. Ivanov, A. et al. Analysis of intron sequences reveals hallmarks of circular RNA biogenesis in animals. Cell Rep. 10, 170–177 (2015).

    Article  CAS  PubMed  Google Scholar 

  74. Ashwal-Fluss, R. et al. circRNA biogenesis competes with pre-mRNA splicing. Mol. Cell 56, 55–66 (2014).

    Article  CAS  PubMed  Google Scholar 

  75. Conn, S. J. et al. The RNA binding protein quaking regulates formation of circRNAs. Cell 160, 1125–1134 (2015). This study shows that QKI facilitates certain circRNA back-splicing events across developmental pathways, possibly through its dimerization when bound to flanking intronic elements.

    Article  CAS  PubMed  Google Scholar 

  76. Grosso, A. R. et al. Pervasive transcription read-through promotes aberrant expression of oncogenes and RNA chimeras in renal carcinoma. eLife 4, e09214 (2015). This paper reports that transcription read-through beyond the termination site in clear cell renal cell carcinoma (ccRCC) leads to chimeric transcripts through cis -splicing that correlates with poor survival rates.

    Article  PubMed  PubMed Central  Google Scholar 

  77. Akiva, P. et al. Transcription-mediated gene fusion in the human genome. Genome Res. 16, 30–36 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  78. Qin, F. et al. Discovery of CTCF-sensitive cis-spliced fusion RNAs between adjacent genes in human prostate cells. PLoS Genet. 11, e1005001 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  79. Jividen, K. & Li, H. Chimeric RNAs generated by intergenic splicing in normal and cancer cells. Genes Chromosomes Cancer 53, 963–971 (2014).

    Article  CAS  PubMed  Google Scholar 

  80. Sutton, R. E. & Boothroyd, J. C. Evidence for trans splicing in trypanosomes. Cell 47, 527–535 (1986).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  81. Allen, M. A., Hillier, L. W., Waterston, R. H. & Blumenthal, T. A global analysis of C. elegans trans-splicing. Genome Res. 21, 255–264 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  82. McManus, C. J., Duff, M. O., Eipper-Mains, J. & Graveley, B. R. Global analysis of trans-splicing in Drosophila. Proc. Natl Acad. Sci. USA 107, 12975–12979 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  83. Dorn, R., Reuter, G. & Loewendorf, A. Transgene analysis proves mRNA trans-splicing at the complex mod(mdg4) locus in Drosophila. Proc. Natl Acad. Sci. USA 98, 9724–9729 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  84. Gabler, M. et al. Trans-splicing of the mod(mdg4) complex locus is conserved between the distantly related species Drosophila melanogaster and D. virilis. Genetics 169, 723–736 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  85. Kong, Y. et al. The evolutionary landscape of intergenic trans-splicing events in insects. Nat. Commun. 6, 8734 (2015).

    Article  CAS  PubMed  Google Scholar 

  86. Li, H., Wang, J., Mor, G. & Sklar, J. A neoplastic gene fusion mimics trans-splicing of RNAs in normal human cells. Science 321, 1357–1361 (2008). This paper reports that physiologically regulated trans -splicing between precursor mRNAs for JAZF1 and JJAZ1 forms a chimeric transcript and protein with anti-apoptotic activity identical to that produced from chromosomal rearrangements in human tumours.

    Article  CAS  PubMed  Google Scholar 

  87. Wu, C. S. et al. Integrative transcriptome sequencing identifies trans-splicing events with important roles in human embryonic stem cell pluripotency. Genome Res. 24, 25–36 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  88. Dietrich, R. C., Incorvaia, R. & Padgett, R. A. Terminal intron dinucleotide sequences do not distinguish between U2- and U12-dependent introns. Mol. Cell 1, 151–160 (1997).

    Article  CAS  PubMed  Google Scholar 

  89. Wu, Q. & Krainer, A. R. Splicing of a divergent subclass of AT-AC introns requires the major spliceosomal snRNAs. RNA 3, 586–601 (1997).

    CAS  PubMed  PubMed Central  Google Scholar 

  90. Sheth, N. et al. Comprehensive splice-site analysis using comparative genomics. Nucleic Acids Res. 34, 3955–3967 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  91. Parada, G. E., Munita, R., Cerda, C. A. & Gysling, K. A comprehensive survey of non-canonical splice sites in the human transcriptome. Nucleic Acids Res. 42, 10564–10578 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  92. Mercer, T. R. et al. Genome-wide discovery of human splicing branchpoints. Genome Res. 25, 290–303 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  93. DeBoever, C. et al. Transcriptome sequencing reveals potential mechanism of cryptic 3′ splice site selection in SF3B1-mutated cancers. PLoS Comput. Biol. 11, e1004105 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  94. Darman, R. B. et al. Cancer-associated SF3B1 hotspot mutations induce cryptic 3′ splice site selection through use of a different branch point. Cell Rep. 13, 1033–1045 (2015). This paper shows that mutations in SF3B1 lead to tumour-specific splicing changes by using an alternative branch point that induces aberrant 3′ splice site selection.

    Article  CAS  PubMed  Google Scholar 

  95. Alsafadi, S. et al. Cancer-associated SF3B1 mutations affect alternative splicing by promoting alternative branchpoint usage. Nat. Commun. 7, 10615 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  96. Roca, X. & Krainer, A. R. Recognition of atypical 5′ splice sites by shifted base-pairing to U1 snRNA. Nat. Struct. Mol. Biol. 16, 176–182 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  97. Roca, X. et al. Widespread recognition of 5′ splice sites by noncanonical base-pairing to U1 snRNA involving bulged nucleotides. Genes Dev. 26, 1098–1109 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  98. Rueter, S. M., Dawson, T. R. & Emeson, R. B. Regulation of alternative splicing by RNA editing. Nature 399, 75–80 (1999).

    Article  CAS  PubMed  Google Scholar 

  99. Shen, X. et al. Complementary signaling pathways regulate the unfolded protein response and are required for C. elegans development. Cell 107, 893–903 (2001).

    Article  CAS  PubMed  Google Scholar 

  100. Yoshida, H., Matsui, T., Yamamoto, A., Okada, T. & Mori, K. XBP1 mRNA is induced by ATF6 and spliced by IRE1 in response to ER stress to produce a highly active transcription factor. Cell 107, 881–891 (2001).

    Article  CAS  PubMed  Google Scholar 

  101. Filipowicz, W. Making ends meet: a role of RNA ligase RTCB in unfolded protein response. EMBO J. 33, 2887–2889 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  102. Dergai, M. et al. Microexon-based regulation of ITSN1 and Src SH3 domains specificity relies on introduction of charged amino acids into the interaction interface. Biochem. Biophys. Res. Commun. 399, 307–312 (2010).

    Article  CAS  PubMed  Google Scholar 

  103. Quesnel-Vallieres, M., Irimia, M., Cordes, S. P. & Blencowe, B. J. Essential roles for the splicing regulator nSR100/SRRM4 during nervous system development. Genes Dev. 29, 746–759 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  104. Wright, P. E. & Dyson, H. J. Intrinsically disordered proteins in cellular signalling and regulation. Nat. Rev. Mol. Cell Biol. 16, 18–29 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  105. Rossbach, O. et al. Auto- and cross-regulation of the hnRNP L proteins by alternative splicing. Mol. Cell. Biol. 29, 1442–1451 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  106. Buckley, P. T., Khaladkar, M., Kim, J. & Eberwine, J. Cytoplasmic intron retention, function, splicing, and the sentinel RNA hypothesis. Wiley Interdiscip Rev. RNA 5, 223–230 (2014).

    Article  CAS  PubMed  Google Scholar 

  107. Sibley, C. R. Regulation of gene expression through production of unstable mRNA isoforms. Biochem. Soc. Trans. 42, 1196–1205 (2014).

    Article  CAS  PubMed  Google Scholar 

  108. Jens, M. & Rajewsky, N. Competition between target sites of regulators shapes post-transcriptional gene regulation. Nat. Rev. Genet. 16, 113–126 (2015).

    Article  CAS  PubMed  Google Scholar 

  109. Dhir, A., Buratti, E., van Santen, M. A., Luhrmann, R. & Baralle, F. E. The intronic splicing code: multiple factors involved in ATM pseudoexon definition. EMBO J. 29, 749–760 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  110. Pagani, F. et al. A new type of mutation causes a splicing defect in ATM. Nat. Genet. 30, 426–429 (2002).

    Article  CAS  PubMed  Google Scholar 

  111. Liu, N. et al. N6-methyladenosine-dependent RNA structural switches regulate RNA–protein interactions. Nature 518, 560–564 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  112. Solomon, O. et al. Global regulation of alternative splicing by adenosine deaminase acting on RNA (ADAR). RNA 19, 591–604 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  113. Lovci, M. T. et al. Rbfox proteins regulate alternative mRNA splicing through evolutionarily conserved RNA bridges. Nat. Struct. Mol. Biol. 20, 1434–1442 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  114. Bitton, D. A. et al. Widespread exon skipping triggers degradation by nuclear RNA surveillance in fission yeast. Genome Res. 25, 884–896 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  115. de Koning, A. P., Gu, W., Castoe, T. A., Batzer, M. A. & Pollock, D. D. Repetitive elements may comprise over two-thirds of the human genome. PLoS Genet. 7, e1002384 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  116. Brouha, B. et al. Hot L1s account for the bulk of retrotransposition in the human population. Proc. Natl Acad. Sci. USA 100, 5280–5285 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  117. Jacob, F. Evolution and tinkering. Science 196, 1161–1166 (1977).

    Article  CAS  PubMed  Google Scholar 

  118. Cowley, M. & Oakey, R. J. Transposable elements re-wire and fine-tune the transcriptome. PLoS Genet. 9, e1003234 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  119. Ule, J. Alu elements: at the crossroads between disease and evolution. Biochem. Soc. Trans. 41, 1532–1535 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  120. Brunet, T. D. & Doolittle, W. F. Multilevel selection theory and the evolutionary functions of transposable elements. Genome Biol. Evol. 7, 2445–2457 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  121. Feschotte, C. Transposable elements and the evolution of regulatory networks. Nat. Rev. Genet. 9, 397–405 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  122. Roy, M., Kim, N., Xing, Y. & Lee, C. The effect of intron length on exon creation ratios during the evolution of mammalian genomes. RNA 14, 2261–2273 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  123. Pickrell, J. K., Pai, A. A., Gilad, Y. & Pritchard, J. K. Noisy splicing drives mRNA isoform diversity in human cells. PLoS Genet. 6, e1001236 (2010).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  124. Lopez-Bigas, N., Audit, B., Ouzounis, C., Parra, G. & Guigo, R. Are splicing mutations the most frequent cause of hereditary disease? FEBS Lett. 579, 1900–1903 (2005).

    Article  CAS  PubMed  Google Scholar 

  125. Daguenet, E., Dujardin, G. & Valcarcel, J. The pathogenicity of splicing defects: mechanistic insights into pre-mRNA processing inform novel therapeutic approaches. EMBO Rep. 16, 1640–1655 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  126. Singh, R. K. & Cooper, T. A. Pre-mRNA splicing in disease and therapeutics. Trends Mol. Med. 18, 472–482 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  127. Supek, F., Minana, B., Valcarcel, J., Gabaldon, T. & Lehner, B. Synonymous mutations frequently act as driver mutations in human cancers. Cell 156, 1324–1335 (2014).

    Article  CAS  PubMed  Google Scholar 

  128. Xiong, H. Y. et al. RNA splicing. The human splicing code reveals new insights into the genetic determinants of disease. Science 347, 1254806 (2015). In this study, machine learning reveals the effect of distal sequence variants on splicing outcome, the predictive value of which will be important when considering genomic variation at non-canonical splicing elements.

    Article  PubMed  CAS  Google Scholar 

  129. Meili, D. et al. Disease-causing mutations improving the branch site and polypyrimidine tract: pseudoexon activation of LINE-2 and antisense Alu lacking the poly(T)-tail. Hum. Mutat. 30, 823–831 (2009).

    Article  CAS  PubMed  Google Scholar 

  130. Ferlini, A. et al. A novel Alu-like element rearranged in the dystrophin gene causes a splicing mutation in a family with X-linked dilated cardiomyopathy. Am. J. Hum. Genet. 63, 436–446 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  131. Sowalsky, A. G. et al. Whole transcriptome sequencing reveals extensive unspliced mRNA in metastatic castration-resistant prostate cancer. Mol. Cancer Res. 13, 98–106 (2015).

    Article  CAS  PubMed  Google Scholar 

  132. Yuan, H. et al. A chimeric RNA characteristic of rhabdomyosarcoma in normal myogenesis process. Cancer Discov. 3, 1394–1403 (2013).

    Article  CAS  PubMed  Google Scholar 

  133. Greer, K. et al. Pseudoexon activation increases phenotype severity in a Becker muscular dystrophy patient. Mol. Genet. Genom. Med. 3, 320–326 (2015).

    Article  CAS  Google Scholar 

  134. Buratti, E., Dhir, A., Lewandowska, M. A. & Baralle, F. E. RNA structure is a key regulatory element in pathological ATM and CFTR pseudoexon inclusion events. Nucleic Acids Res. 35, 4369–4383 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  135. Highsmith, W. E. et al. A novel mutation in the cystic fibrosis gene in patients with pulmonary disease but normal sweat chloride concentrations. N. Engl. J. Med. 331, 974–980 (1994).

    Article  CAS  PubMed  Google Scholar 

  136. Chen, X. et al. Intronic alterations in BRCA1 and BRCA2: effect on mRNA splicing fidelity and expression. Hum. Mutat. 27, 427–435 (2006).

    Article  CAS  PubMed  Google Scholar 

  137. Lualdi, S. et al. Multiple cryptic splice sites can be activated by IDS point mutations generating misspliced transcripts. J. Mol. Med. (Berl.) 84, 692–700 (2006).

    Article  CAS  Google Scholar 

  138. Sathasivam, K. et al. Aberrant splicing of HTT generates the pathogenic exon 1 protein in Huntington disease. Proc. Natl Acad. Sci. USA 110, 2366–2370 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  139. Ghosal, S., Das, S., Sen, R., Basak, P. & Chakrabarti, J. Circ2Traits: a comprehensive database for circular RNA potentially associated with disease and traits. Frontiers Genet. 4, 283 (2013).

    Article  CAS  Google Scholar 

  140. Akker, S. A. et al. Pre-spliceosomal binding of U1 small nuclear ribonucleoprotein (RNP) and heterogenous nuclear RNP E1 is associated with suppression of a growth hormone receptor pseudoexon. Mol. Endocrinol. 21, 2529–2540 (2007).

    Article  CAS  PubMed  Google Scholar 

  141. Vorechovsky, I. Transposable elements in disease-associated cryptic exons. Hum. Genet. 127, 135–154 (2010).

    Article  CAS  PubMed  Google Scholar 

  142. Madan, V. et al. Aberrant splicing of U12-type introns is the hallmark of ZRSR2 mutant myelodysplastic syndrome. Nat. Commun. 6, 6042 (2015).

    Article  CAS  PubMed  Google Scholar 

  143. Edery, P. et al. Association of TALS developmental disorder with defect in minor splicing component U4atac snRNA. Science 332, 240–243 (2011).

    Article  CAS  PubMed  Google Scholar 

  144. He, H. et al. Mutations in U4atac snRNA, a component of the minor spliceosome, in the developmental disorder MOPD I. Science 332, 238–240 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  145. Merico, D. et al. Compound heterozygous mutations in the noncoding RNU4ATAC cause Roifman Syndrome by disrupting minor intron splicing. Nat. Commun. 6, 8718 (2015).

    Article  CAS  PubMed  Google Scholar 

  146. Yoshida, K. et al. Frequent pathway mutations of splicing machinery in myelodysplasia. Nature 478, 64–69 (2011).

    Article  CAS  PubMed  Google Scholar 

  147. Menzies, F. M., Fleming, A. & Rubinsztein, D. C. Compromised autophagy and neurodegenerative diseases. Nat. Rev. Neurosci. 16, 345–357 (2015).

    Article  CAS  PubMed  Google Scholar 

  148. Argente, J. et al. Defective minor spliceosome mRNA processing results in isolated familial growth hormone deficiency. EMBO Mol. Med. 6, 299–306 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  149. Bachmayr-Heyda, A. et al. Correlation of circular RNA abundance with proliferation — exemplified with colorectal and ovarian cancer, idiopathic lung fibrosis, and normal human tissues. Scientif. Rep. 5, 8057 (2015).

    Article  CAS  Google Scholar 

  150. Wang, Y. H., Yu, X. H., Luo, S. S. & Han, H. Comprehensive circular RNA profiling reveals that circular RNA100783 is involved in chronic CD28-associated CD8+ T cell ageing. Immun. Ageing 12, 17 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  151. Li, J. et al. Circular RNAs in cancer: novel insights into origins, properties, functions and implications. Am. J. Cancer Res. 5, 472–480 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  152. Memczak, S., Papavasileiou, P., Peters, O. & Rajewsky, N. Identification and characterization of circular RNAs as a new class of putative biomarkers in human blood. PLoS ONE 10, e0141214 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  153. Dvinge, H. & Bradley, R. K. Widespread intron retention diversifies most cancer transcriptomes. Genome Med. 7, 45 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  154. Jung, H. et al. Intron retention is a widespread mechanism of tumor-suppressor inactivation. Nat. Genet. 47, 1242–1248 (2015).

    Article  CAS  PubMed  Google Scholar 

  155. Romano, M., Buratti, E. & Baralle, D. Role of pseudoexons and pseudointrons in human cancer. Int. J. Cell Biol. 2013, 810572 (2013).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  156. Hsu, T. Y. et al. The spliceosome is a therapeutic vulnerability in MYC-driven cancer. Nature 525, 384–388 (2015). In this study, the spliceosome is found to be a target of oncogenic stress in MYC-dependent breast cancers, providing an opportunity for genetic or pharmacological spliceosome inhibition.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  157. Ilagan, J. O. et al. U2AF1 mutations alter splice site recognition in hematological malignancies. Genome Res. 25, 14–26 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  158. Milde-Langosch, K., Kappes, H., Riethdorf, S., Loning, T. & Bamberger, A. M. FosB is highly expressed in normal mammary epithelia, but down-regulated in poorly differentiated breast carcinomas. Breast Cancer Res. Treat. 77, 265–275 (2003).

    Article  CAS  PubMed  Google Scholar 

  159. Rickman, D. S. et al. SLC45A3–ELK4 is a novel and frequent erythroblast transformation-specific fusion transcript in prostate cancer. Cancer Res. 69, 2734–2738 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  160. Zhang, Y. et al. Chimeric transcript generated by cis-splicing of adjacent genes regulates prostate cancer cell proliferation. Cancer Discov. 2, 598–607 (2012).

    Article  CAS  PubMed  Google Scholar 

  161. Bonnal, S., Vigevani, L. & Valcarcel, J. The spliceosome as a target of novel antitumour drugs. Nat. Rev. Drug Discov. 11, 847–859 (2012).

    Article  CAS  PubMed  Google Scholar 

  162. Koh, C. M. et al. MYC regulates the core pre-mRNA splicing machinery as an essential step in lymphomagenesis. Nature 523, 96–100 (2015).

    Article  CAS  PubMed  Google Scholar 

  163. Dominski, Z. & Kole, R. Restoration of correct splicing in thalassemic pre-mRNA by antisense oligonucleotides. Proc. Natl Acad. Sci. USA 90, 8673–8677 (1993).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  164. Hua, Y. et al. Peripheral SMN restoration is essential for long-term rescue of a severe spinal muscular atrophy mouse model. Nature 478, 123–126 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  165. McClorey, G. & Wood, M. J. An overview of the clinical application of antisense oligonucleotides for RNA-targeting therapies. Curr. Opin. Pharmacol. 24, 52–58 (2015).

    Article  CAS  PubMed  Google Scholar 

  166. Goyenvalle, A. et al. Rescue of dystrophic muscle through U7 snRNA-mediated exon skipping. Science 306, 1796–1799 (2004).

    Article  CAS  PubMed  Google Scholar 

  167. Gorman, L., Suter, D., Emerick, V., Schumperli, D. & Kole, R. Stable alteration of pre-mRNA splicing patterns by modified U7 small nuclear RNAs. Proc. Natl Acad. Sci. USA 95, 4929–4934 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  168. Uchikawa, H. et al. U7 snRNA-mediated correction of aberrant splicing caused by activation of cryptic splice sites. J. Hum. Genet. 52, 891–897 (2007).

    Article  CAS  PubMed  Google Scholar 

  169. Blazquez, L. et al. In vitro correction of a pseudoexon-generating deep intronic mutation in LGMD2A by antisense oligonucleotides and modified small nuclear RNAs. Hum. Mutat. 34, 1387–1395 (2013).

    Article  CAS  PubMed  Google Scholar 

  170. Goyenvalle, A., Babbs, A., van Ommen, G. J., Garcia, L. & Davies, K. E. Enhanced exon-skipping induced by U7 snRNA carrying a splicing silencer sequence: promising tool for DMD therapy. Mol. Ther. 17, 1234–1240 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  171. Garcia-Blanco, M. A., Baraniak, A. P. & Lasda, E. L. Alternative splicing in disease and therapy. Nat. Biotechnol. 22, 535–546 (2004).

    Article  CAS  PubMed  Google Scholar 

  172. Xu, L. et al. CRISPR-mediated genome editing restores dystrophin expression and function in mdx mice. Mol. Ther. 24, 564–569 (2015).

    Article  PubMed  CAS  Google Scholar 

  173. Nelson, C. E. et al. In vivo genome editing improves muscle function in a mouse model of Duchenne muscular dystrophy. Science 351, 403–407 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  174. Tabebordbar, M. et al. In vivo gene editing in dystrophic mouse muscle and muscle stem cells. Science 351, 407–411 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  175. Puttaraju, M., DiPasquale, J., Baker, C. C., Mitchell, L. G. & Garcia-Blanco, M. A. Messenger RNA repair and restoration of protein function by spliceosome-mediated RNA trans-splicing. Mol. Ther. 4, 105–114 (2001).

    Article  CAS  PubMed  Google Scholar 

  176. Koller, U., Wally, V., Bauer, J. W. & Murauer, E. M. Considerations for a successful RNA trans-splicing repair of genetic disorders. Mol. Ther. Nucleic Acids 3, e157 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  177. Chao, H. et al. Phenotype correction of hemophilia A mice by spliceosome-mediated RNA trans-splicing. Nat. Med. 9, 1015–1019 (2003).

    Article  CAS  PubMed  Google Scholar 

  178. Petkovic, S. & Muller, S. RNA circularization strategies in vivo and in vitro. Nucleic Acids Res. 43, 2454–2465 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  179. Davidson, L., Kerr, A. & West, S. Co-transcriptional degradation of aberrant pre-mRNA by Xrn2. EMBO J. 31, 2566–2578 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  180. Shen, S. et al. Widespread establishment and regulatory impact of Alu exons in human genes. Proc. Natl Acad. Sci. USA 108, 2837–2842 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  181. Tajnik, M. et al. Intergenic Alu exonisation facilitates the evolution of tissue-specific transcript ends. Nucleic Acids Res. 43, 10492–10505 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  182. Rybak-Wolf, A. et al. Circular RNAs in the mammalian brain are highly abundant, conserved, and dynamically expressed. Mol. Cell 58, 870–885 (2015).

    Article  CAS  PubMed  Google Scholar 

  183. Kellis, M. et al. Defining functional DNA elements in the human genome. Proc. Natl Acad. Sci. USA 111, 6131–6138 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  184. Naftelberg, S., Schor, I. E., Ast, G. & Kornblihtt, A. R. Regulation of alternative splicing through coupling with transcription and chromatin structure. Annu. Rev. Biochem. 84, 165–198 (2015).

    Article  CAS  PubMed  Google Scholar 

  185. Pulyakhina, I. et al. SplicePie: a novel analytical approach for the detection of alternative, non-sequential and recursive splicing. Nucleic Acids Res. 43, e80 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  186. Chuang, T. J. et al. NCLscan: accurate identification of non-co-linear transcripts (fusion, trans-splicing and circular RNA) with a good balance between sensitivity and precision. Nucleic Acids Res. 44, e29 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  187. Szabo, L. et al. Statistically based splicing detection reveals neural enrichment and tissue-specific induction of circular RNA during human fetal development. Genome Biol. 16, 126 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  188. McPherson, A. et al. deFuse: an algorithm for gene fusion discovery in tumor RNA-seq data. PLoS Comput. Biol. 7, e1001138 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  189. Maher, C. A. et al. Transcriptome sequencing to detect gene fusions in cancer. Nature 458, 97–101 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  190. Konig, J. et al. iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nature Struct. Mol. Biol. 17, 909–915 (2010).

    Article  CAS  Google Scholar 

  191. Alioto, T. S. U12DB: a database of orthologous U12-type spliceosomal introns. Nucleic Acids Res. 35, D110–D115 (2007).

    Article  CAS  PubMed  Google Scholar 

  192. Crooks, G. E., Hon, G., Chandonia, J. M. & Brenner, S. E. WebLogo: a sequence logo generator. Genome Res. 14, 1188–1190 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

The authors thank K. Zarnack for comments on the manuscript. This work was supported by the European Research Council (617837-Translate) and a Marie Curie Post-doctoral Research Fellowship (627783-NeuroCRYSP) to L.B., and an Edmond and Lily Safra Fellowship to C.R.S.

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  1. Christopher R. Sibley and Lorea Blazquez: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Molecular Neuroscience, University College London Institute of Neurology, Russell Square House, Russell Square, WC1B 5EH, London, UK

    Christopher R. Sibley, Lorea Blazquez & Jernej Ule

  2. Department of Medicine, Division of Brain Sciences, Imperial College London, Burlington Danes, DuCane Road, W12 0NN, London, UK

    Christopher R. Sibley

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  1. Christopher R. Sibley
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Correspondence to Jernej Ule.

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FURTHER INFORMATION

RepeatMasker

PowerPoint slides

Glossary

Splice sites

Sequences at the boundary of exons and introns, which contain motifs that recruit small nuclear ribonucleoproteins and RNA-binding proteins to initiate the splicing reaction. 3′ and 5′ splice sites are located upstream and downstream of exons, respectively.

Cryptic exons

Exons that are not annotated by current genomic databases, such as ENSEMBL, and are often only revealed after removing a repressive RNA-binding protein or after a genomic mutation that increases their splicing efficiency.

Microexons

Exons that are shorter than 30 nt.

Recursive splicing

A mechanism that allows an intron to be spliced in two or more steps.

Circular RNAs

(circRNAs). RNA molecules that have become circularized owing to intramolecular ligation of their 5′ and 3′ ends.

Exonic introns

(Exitrons). Introns located within annotated exons.

Exon definition mechanism

The process by which exons are recognized and defined as functional units through interactions between multiple small nuclear ribonucleoproteins (snRNPs) and RNA-binding proteins (RBPs), especially U1 and U2 snRNPs and serine/arginine-rich proteins.

Small nuclear ribonucleoproteins

(snRNPs). Ribonucleoprotein complexes assembled around the small nuclear RNAs that interact with splice sites or the branch point on pre-mRNA and thereby coordinate and catalyse the splicing reaction.

U2 auxiliary factor complex

(U2AF complex). A complex of two U2AF RNA-binding proteins (RBPs) that bind the 3′ splice site and facilitate the recruitment of the U2 small nuclear ribonucleoprotein (snRNP) to the branch point.

Serine/arginine-rich proteins

(SR proteins). A family of RNA-binding proteins (RBPs) containing a protein domain with long repeats of serine and arginine that generally promote exon definition when binding to exons.

Recursive splice sites

(RS sites). The sites of recursive splicing, which consists of a 3′ splice site that is followed by a sequence that reconstitutes a 5′ splice site after the first splicing event.

Seed sequence

The section of a sequencing read that is used to align the read to the genome or transcriptome

Nonsense-mediated decay

(NMD). A pathway that initiates decay of certain transcripts, especially those containing a premature termination codon.

Alu elements

Retrotransposons, ~300 nt long, belonging to the family of short interspersed elements (SINEs), which originally derived from 7SL signal recognition particle RNA.

Crosslinking and immunoprecipitation

(CLIP). A method used to identify the RNA targets bound by an RNA-binding proteinof-interest that employs crosslinking, immunoprecipitation and stringent purification of protein–RNA complexes by SDS-PAGE.

NOVA RBPs

A joint name for RNA-binding proteins encoded by two partially redundant genes that are expressed in the brain, neuro-oncological ventral antigen 1 (NOVA1) and NOVA2.

NMD exons

(Nonsense-mediated decay exons). Exons that contain a premature termination codon and are therefore targeted for NMD.

Intrasplicing

An unconventional splicing mechanism in which splicing to a 3′ splice site reconstitutes a new 3′ splice to be used in a subsequent splicing step.

RS site exons

(Recursive splice site exons). Exons that follow an RS site and which are required for the exon definition mechanism that initiates splicing at the RS site.

Spliceosome

A macromolecular machine consisting of small nuclear ribonucleoproteins (snRNPs) and additional RNA-binding proteins (RBPs) that coordinate and catalyse the splicing reaction.

Chimeric transcripts

Transcripts that are formed when sections of two or more different genes are joined together in a new transcript either by splicing or as a result of chromosomal fusions.

Axonogenesis

The generation and outgrowth of axons during neuronal development.

Ataxia telangiectasia

An autosomal recessive disorder involving cerebellar degeneration, immunodeficiency, chromosomal instability, radiosensitivity and cancer predisposition. It is caused by mutations in ataxia telangiectasia mutated (ATM).

Laron syndrome

An autosomal recessive disorder characterized by short stature caused by mutations in growth hormone receptor (GHR).

Duchenne muscular dystrophy

A progressive proximal muscular dystrophy caused by mutations in dystrophin (DMD).

Hyperphenylalaninaemia

A neurologic disorder caused by autosomal recessive mutations in the genes encoding enzymes involved in the synthesis or regenerationof the BH4 (tetrahydrobiopterin) cofactor. The most common form is caused by mutations in PTS (6-pyruvoyltetrahydropterin synthase).

Autophagy

Intracellular pathway responsible for regulated disassembly of unnecessary or dysfunctional cellular components after their targeting to lysosomes.

β-thalassaemia

A genetic blood disorder characterized by a defective synthesis of the β-globin chains of haemoglobin, thus causing abnormal erythropoiesis and anaemia.

Aptamers

Oligonucleotide (or peptide) molecules that have secondary and tertiary structures that strongly bind to specific proteins or other cellular targets.

Co-transcriptional decay

RNA surveillance mechanism that acts in the nucleus while transcripts are still associated with the chromatin template.

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Sibley, C., Blazquez, L. & Ule, J. Lessons from non-canonical splicing. Nat Rev Genet 17, 407–421 (2016). https://doi.org/10.1038/nrg.2016.46

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