Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Review Article
  • Published:

Expansion of the eukaryotic proteome by alternative splicing

Nature volume 463, pages 457–463 (2010)Cite this article

Subjects

Abstract

The collection of components required to carry out the intricate processes involved in generating and maintaining a living, breathing and, sometimes, thinking organism is staggeringly complex. Where do all of the parts come from? Early estimates stated that about 100,000 genes would be required to make up a mammal; however, the actual number is less than one-quarter of that, barely four times the number of genes in budding yeast. It is now clear that the 'missing' information is in large part provided by alternative splicing, the process by which multiple different functional messenger RNAs, and therefore proteins, can be synthesized from a single gene.

Access through your institution
Buy or subscribe

This is a preview of subscription content, access via your institution

Access options

Access through your institution

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Types of alternative splicing.
Figure 2: The generation of diverse mRNA repertoires.
Figure 3: Alternative splicing regulatory mechanisms.

Similar content being viewed by others

References

  1. Alt, F. W. et al. Synthesis of secreted and membrane-bound immunoglobulin μ heavy chains is directed by mRNAs that differ at their 3′ ends. Cell 20, 293–301 (1980).

    Article  PubMed  CAS  Google Scholar 

  2. Early, P. et al. Two mRNAs can be produced from a single immunoglobulin μ gene by alternative RNA processing pathways. Cell 20, 313–319 (1980).

    Article  PubMed  CAS  Google Scholar 

  3. Rosenfeld, M. G. et al. Calcitonin mRNA polymorphism: peptide switching associated with alternative RNA splicing events. Proc. Natl Acad. Sci. USA 79, 1717–1721 (1982).

    Article  ADS  PubMed  CAS  PubMed Central  Google Scholar 

  4. Pan, Q., Shai, O., Lee, L. J., Frey, B. J. & Blencowe, B. J. Deep surveying of alternative splicing complexity in the human transcriptome by high-throughput sequencing. Nature Genet. 40, 1413–1415 (2008).

    Article  PubMed  CAS  Google Scholar 

  5. Wang, E. T. et al. Alternative isoform regulation in human tissue transcriptomes. Nature 456, 470–476 (2008). References 4 and 5 provide detailed views of the human transcriptome as determined by using deep-sequencing data. The authors conclude that the pre-mRNAs from all multi-exon genes are alternatively spliced.

    Article  ADS  PubMed  PubMed Central  CAS  Google Scholar 

  6. Schmucker, D. et al. Drosophila Dscam is an axon guidance receptor exhibiting extraordinary molecular diversity. Cell 101, 671–684 (2000).

    Article  PubMed  CAS  Google Scholar 

  7. Navaratnam, D. S., Bell, T. J., Tu, T. D., Cohen, E. L. & Oberholtzer, J. C. Differential distribution of Ca2+-activated K+ channel splice variants among hair cells along the tonotopic axis of the chick cochlea. Neuron 19, 1077–1085 (1997).

    Article  PubMed  CAS  Google Scholar 

  8. Rosenblatt, K. P., Sun, Z. P., Heller, S. & Hudspeth, A. J. Distribution of Ca2+-activated K+ channel isoforms along the tonotopic gradient of the chicken's cochlea. Neuron 19, 1061–1075 (1997).

    Article  PubMed  CAS  Google Scholar 

  9. 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  ADS  PubMed  CAS  PubMed Central  Google Scholar 

  10. Labrador, M. et al. Protein encoding by both DNA strands. Nature 409, 1000 (2001). References 9 and 10 show that trans-splicing of two separate pre-mRNAs can generate a single protein-coding mRNA.

    Article  ADS  PubMed  CAS  Google Scholar 

  11. Sánchez, L. Sex-determining mechanisms in insects. Int. J. Dev. Biol. 52, 837–856 (2008).

    Article  PubMed  Google Scholar 

  12. Makeyev, E. V., Zhang, J., Carrasco, M. A. & Maniatis, T. The microRNA miR-124 promotes neuronal differentiation by triggering brain-specific alternative pre-mRNA splicing. Mol. Cell 27, 435–448 (2007).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  13. Boutz, P. L. et al. A post-transcriptional regulatory switch in polypyrimidine tract-binding proteins reprograms alternative splicing in developing neurons. Genes Dev. 21, 1636–1652 (2007).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  14. Xie, J. & Black, D. L. A CaMK IV responsive RNA element mediates depolarization-induced alternative splicing of ion channels. Nature 410, 936–939 (2001).

    Article  ADS  PubMed  CAS  Google Scholar 

  15. Lynch, K. W. Regulation of alternative splicing by signal transduction pathways. Adv. Exp. Med. Biol. 623, 161–174 (2007).

    Article  PubMed  Google Scholar 

  16. Shin, C. & Manley, J. L. Cell signalling and the control of pre-mRNA splicing. Nature Rev. Mol. Cell Biol. 5, 727–738 (2004).

    Article  CAS  Google Scholar 

  17. Matlin, A. J., Clark, F. & Smith, C. W. Understanding alternative splicing: towards a cellular code. Nature Rev. Mol. Cell Biol. 6, 386–398 (2005).

    Article  CAS  Google Scholar 

  18. Gabut, M., Chaudhry, S. & Blencowe, B. J. The splicing regulatory machinery. Cell 133, 192 (2008).

  19. Buckanovich, R. J., Posner, J. B. & Darnell, R. B. Nova, the paraneoplastic Ri antigen, is homologous to an RNA-binding protein and is specifically expressed in the developing motor system. Neuron 11, 657–672 (1993).

    Article  PubMed  CAS  Google Scholar 

  20. Polydorides, A. D., Okano, H. J., Yang, Y. Y., Stefani, G. & Darnell, R. B. A brain-enriched polypyrimidine tract-binding protein antagonizes the ability of Nova to regulate neuron-specific alternative splicing. Proc. Natl Acad. Sci. USA 97, 6350–6355 (2000).

    Article  ADS  PubMed  CAS  PubMed Central  Google Scholar 

  21. Markovtsov, V. et al. Cooperative assembly of an hnRNP complex induced by a tissue-specific homolog of polypyrimidine tract binding protein. Mol. Cell. Biol. 20, 7463–7479 (2000).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  22. Underwood, J. G., Boutz, P. L., Dougherty, J. D., Stoilov, P. & Black, D. L. Homologues of the Caenorhabditis elegans Fox-1 protein are neuronal splicing regulators in mammals. Mol. Cell. Biol. 25, 10005–10016 (2005).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  23. Jin, Y. et al. A vertebrate RNA-binding protein Fox-1 regulates tissue-specific splicing via the pentanucleotide GCAUG. EMBO J. 22, 905–912 (2003).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  24. Warzecha, C. C., Sato, T. K., Nabet, B., Hogenesch, J. B. & Carstens, R. P. ESRP1 and ESRP2 are epithelial cell-type-specific regulators of FGFR2 splicing. Mol. Cell 33, 591–601 (2009).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  25. Calarco, J. A. et al. Regulation of vertebrate nervous system-specific alternative splicing and development by an SR-related protein. Cell 138, 898–910 (2009).

    Article  PubMed  CAS  Google Scholar 

  26. Lin, S. & Fu, X. D. SR proteins and related factors in alternative splicing. Adv. Exp. Med. Biol. 623, 107–122 (2007).

    Article  PubMed  Google Scholar 

  27. Martinez-Contreras, R. et al. hnRNP proteins and splicing control. Adv. Exp. Med. Biol. 623, 123–147 (2007).

    Article  PubMed  Google Scholar 

  28. Fu, X. D. Towards a splicing code. Cell 119, 736–738 (2004).

    Article  PubMed  CAS  Google Scholar 

  29. Zhang, X. H., Arias, M. A., Ke, S. & Chasin, L. A. Splicing of designer exons reveals unexpected complexity in pre-mRNA splicing. RNA 15, 367–376 (2009).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  30. Blanchette, M. et al. Genome-wide analysis of alternative pre-mRNA splicing and RNA-binding specificities of the Drosophila hnRNP A/B family members. Mol. Cell 33, 438–449 (2009).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  31. Valcárcel, J., Singh, R., Zamore, P. D. & Green, M. R. The protein Sex-lethal antagonizes the splicing factor U2AF to regulate alternative splicing of transformer pre-mRNA. Nature 362, 171–175 (1993).

    Article  ADS  PubMed  Google Scholar 

  32. Zuo, P. & Maniatis, T. The splicing factor U2AF35 mediates critical protein–protein interactions in constitutive and enhancer-dependent splicing. Genes Dev. 10, 1356–1368 (1996).

    Article  PubMed  CAS  Google Scholar 

  33. House, A. E. & Lynch, K. W. An exonic splicing silencer represses spliceosome assembly after ATP-dependent exon recognition. Nature Struct. Mol. Biol. 13, 937–944 (2006).

    Article  CAS  Google Scholar 

  34. Giles, K. E. & Beemon, K. L. Retroviral splicing suppressor sequesters a 3′ splice site in a 50S aberrant splicing complex. Mol. Cell. Biol. 25, 4397–4405 (2005).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  35. Siebel, C. W., Fresco, L. D. & Rio, D. C. The mechanism of somatic inhibition of Drosophila P-element pre-mRNA splicing: multiprotein complexes at an exon pseudo-5′ splice site control U1 snRNP binding. Genes Dev. 6, 1386–1401 (1992).

    Article  PubMed  CAS  Google Scholar 

  36. Sharma, S., Kohlstaedt, L. A., Damianov, A., Rio, D. C. & Black, D. L. Polypyrimidine tract binding protein controls the transition from exon definition to an intron defined spliceosome. Nature Struct. Mol. Biol. 15, 183–191 (2008).

    Article  CAS  Google Scholar 

  37. 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 

  38. Smith, D. J., Query, C. C. & Konarska, M. M. “Nought may endure but mutability”: spliceosome dynamics and the regulation of splicing. Mol. Cell 30, 657–666 (2008).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  39. Kornblihtt, A. R. Coupling transcription and alternative splicing. Adv. Exp. Med. Biol. 623, 175–189 (2007).

    Article  PubMed  Google Scholar 

  40. Park, J. W., Parisky, K., Celotto, A. M., Reenan, R. A. & Graveley, B. R. Identification of alternative splicing regulators by RNA interference in Drosophila. Proc. Natl Acad. Sci. USA 101, 15974–15979 (2004).

    Article  ADS  PubMed  CAS  PubMed Central  Google Scholar 

  41. Pleiss, J. A., Whitworth, G. B., Bergkessel, M. & Guthrie, C. Transcript specificity in yeast pre-mRNA splicing revealed by mutations in core spliceosomal components. PLoS Biol. 5, e90 (2007).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  42. Fox-Walsh, K. L. et al. The architecture of pre-mRNAs affects mechanisms of splice-site pairing. Proc. Natl Acad. Sci. USA 102, 16176–16181 (2005).

    Article  ADS  PubMed  CAS  PubMed Central  Google Scholar 

  43. Yu, Y. et al. Dynamic regulation of alternative splicing by silencers that modulate 5′ splice site competition. Cell 135, 1224–1236 (2008).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  44. Dye, M. J., Gromak, N. & Proudfoot, N. J. Exon tethering in transcription by RNA polymerase II. Mol. Cell 21, 849–859 (2006).

    Article  PubMed  CAS  Google Scholar 

  45. Alló, M. et al. Control of alternative splicing through siRNA-mediated transcriptional gene silencing. Nature Struct. Mol. Biol. 16, 717–724 (2009).

    Article  CAS  Google Scholar 

  46. Kolasinska-Zwierz, P. et al. Differential chromatin marking of introns and expressed exons by H3K36me3. Nature Genet. 41, 376–381 (2009).

    Article  PubMed  CAS  Google Scholar 

  47. Schwartz, S., Meshorer, E. & Ast, G. Chromatin organization marks exon–intron architecture. Nature Struct. Mol. Biol. 16, 990–995 (2009).

    Article  CAS  Google Scholar 

  48. Tilgner, H. et al. Nucleosome positioning as a determinant of exon recognition. Nature Struct. Mol. Biol. 16, 996–1001 (2009).

    Article  CAS  Google Scholar 

  49. Neves, G., Zucker, J., Daly, M. & Chess, A. Stochastic yet biased expression of multiple Dscam splice variants by individual cells. Nature Genet. 36, 240–246 (2004).

    Article  PubMed  CAS  Google Scholar 

  50. Graveley, B. R. Mutually exclusive splicing of the insect Dscam pre-mRNA directed by competing intronic RNA secondary structures. Cell 123, 65–73 (2005).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  51. Anastassiou, D., Liu, H. & Varadan, V. Variable window binding for mutually exclusive alternative splicing. Genome Biol. 7, R2 (2006).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  52. Olson, S. et al. A regulator of Dscam mutually exclusive splicing fidelity. Nature Struct. Mol. Biol. 14, 1134–1140 (2007).

    Article  CAS  Google Scholar 

  53. Ule, J. et al. An RNA map predicting Nova-dependent splicing regulation. Nature 444, 580–586 (2006). This paper shows that the neural-specific splicing regulator NOVA functions as either an activator or a repressor depending on the context.

    Article  ADS  PubMed  CAS  Google Scholar 

  54. Modrek, B. & Lee, C. J. Alternative splicing in the human, mouse and rat genomes is associated with an increased frequency of exon creation and/or loss. Nature Genet. 34, 177–180 (2003).

    Article  PubMed  CAS  Google Scholar 

  55. Sorek, R. & Ast, G. Intronic sequences flanking alternatively spliced exons are conserved between human and mouse. Genome Res. 13, 1631–1637 (2003).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  56. Sugnet, C. W., Kent, W. J., Ares, M. J. & Haussler, D. Transcriptome and genome conservation of alternative splicing events in humans and mice. Pac. Symp. Biocomput. 2004, 66–77 (2004).

    Google Scholar 

  57. Ohler, U., Shomron, N. & Burge, C. B. Recognition of unknown conserved alternatively spliced exons. PLoS Comput. Biol. 1, 113–122 (2005). This paper shows that previously unannotated alternatively spliced exons can be identified accurately by using DNA sequence alone.

    Article  PubMed  CAS  Google Scholar 

  58. Fairbrother, W. G., Yeh, R. F., Sharp, P. A. & Burge, C. B. Predictive identification of exonic splicing enhancers in human genes. Science 297, 1007–1013 (2002). This paper describes one of the first genome-wide predictions of splicing regulatory sequences.

    Article  ADS  PubMed  CAS  Google Scholar 

  59. Goren, A. et al. Comparative analysis identifies exonic splicing regulatory sequences the complex definition of enhancers and silencers. Mol. Cell 22, 769–781 (2006).

    Article  PubMed  CAS  Google Scholar 

  60. Shepard, P. J. & Hertel, K. J. Conserved RNA secondary structures promote alternative splicing. RNA 14, 1463–1469 (2008).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  61. Hasselmann, M. et al. Evidence for the evolutionary nascence of a novel sex determination pathway in honeybees. Nature 454, 519–522 (2008).

    Article  ADS  PubMed  CAS  Google Scholar 

  62. Beye, M., Hasselmann, M., Fondrk, M. K., Page, R. E. & Omholt, S. W. The gene csd is the primary signal for sexual development in the honeybee and encodes an SR-type protein. Cell 114, 419–429 (2003).

    Article  CAS  PubMed  Google Scholar 

  63. Cartegni, L., Chew, S. L. & Krainer, A. R. Listening to silence and understanding nonsense: exonic mutations that affect splicing. Nature Rev. Genet. 3, 285–298 (2002).

    Article  PubMed  CAS  Google Scholar 

  64. Xing, Y. & Lee, C. Alternative splicing and RNA selection pressure — evolutionary consequences for eukaryotic genomes. Nature Rev. Genet. 7, 499–509 (2006).

    Article  PubMed  CAS  Google Scholar 

  65. Zhang, X. H. & Chasin, L. A. Comparison of multiple vertebrate genomes reveals the birth and evolution of human exons. Proc. Natl Acad. Sci. USA 103, 13427–13432 (2006).

    Article  ADS  PubMed  CAS  PubMed Central  Google Scholar 

  66. Calarco, J. A. et al. Global analysis of alternative splicing differences between humans and chimpanzees. Genes Dev. 21, 2963–2975 (2007).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  67. Graveley, B. R. The haplo-spliceo-transcriptome: common variations in alternative splicing in the human population. Trends Genet. 24, 5–7 (2008).

    Article  PubMed  CAS  Google Scholar 

  68. C. elegans Sequencing Consortium. Genome sequence of the nematode C. elegans: a platform for investigating biology. Science 282, 2012–2018 (1998).

  69. Adams, M. D. et al. The genome sequence of Drosophila melanogaster. Science 287, 2185–2195 (2000).

    Article  PubMed  Google Scholar 

  70. Clamp, M. et al. Distinguishing protein-coding and noncoding genes in the human genome. Proc. Natl Acad. Sci. USA 104, 19428–19433 (2007).

    Article  ADS  PubMed  CAS  PubMed Central  Google Scholar 

  71. Hillier, L. W. et al. Massively parallel sequencing of the polyadenylated transcriptome of C. elegans. Genome Res. 19, 657–666 (2009).

    Article  PubMed  CAS  Google Scholar 

  72. Stolc, V. et al. A gene expression map for the euchromatic genome of Drosophila melanogaster. Science 306, 655–660 (2004).

    Article  ADS  PubMed  CAS  Google Scholar 

  73. Celniker, S. E. et al. Unlocking the secrets of the genome. Nature 459, 927–930 (2009).

    Article  ADS  PubMed  PubMed Central  CAS  Google Scholar 

  74. Nern, A. et al. An isoform-specific allele of Drosophila N-cadherin disrupts a late step of R7 targeting. Proc. Natl Acad. Sci. USA 102, 12944–12949 (2005).

    Article  ADS  PubMed  CAS  PubMed Central  Google Scholar 

  75. Wang, Z. & Burge, C. B. Splicing regulation: from a parts list of regulatory elements to an integrated splicing code. RNA 14, 802–813 (2008).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  76. Wojtowicz, W. M. et al. A vast repertoire of Dscam binding specificities arises from modular interactions of variable Ig domains. Cell 130, 1134–1145 (2007).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  77. Demir, E. & Dickson, B. J. fruitless splicing specifies male courtship behavior in Drosophila. Cell 121, 785–794 (2005).

    Article  PubMed  CAS  Google Scholar 

  78. Kwon, S. Y., Xiao, H., Wu, C. & Badenhorst, P. Alternative splicing of NURF301 generates distinct NURF chromatin remodeling complexes with altered modified histone binding specificities. PLoS Genet. 5, e1000574 (2009).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  79. Goodman, S. J., Branda, C. S., Robinson, M. K., Burdine, R. D. & Stern, M. J. Alternative splicing affecting a novel domain in the C. elegans EGL-15 FGF receptor confers functional specificity. Development 130, 3757–3766 (2003).

    Article  PubMed  CAS  Google Scholar 

  80. Muriel, J. M., Dong, C., Hutter, H. & Vogel, B. E. Fibulin-1C and Fibulin-1D splice variants have distinct functions and assemble in a hemicentin-dependent manner. Development 132, 4223–4234 (2005).

    Article  PubMed  CAS  Google Scholar 

  81. Ono, K., Yamashiro, S. & Ono, S. Essential role of ADF/cofilin for assembly of contractile actin networks in the C. elegans somatic gonad. J. Cell Sci. 121, 2662–2670 (2008).

    PubMed  CAS  Google Scholar 

  82. Boucard, A. A., Chubykin, A. A., Comoletti, D., Taylor, P. & Südhof, T. C. A splice code for trans-synaptic cell adhesion mediated by binding of neuroligin 1 to α- and β-neurexins. Neuron 48, 229–236 (2005).

    Article  PubMed  CAS  Google Scholar 

  83. Chen, H. et al. The N-terminal domain of Slack determines the formation and trafficking of Slick/Slack heteromeric sodium-activated potassium channels. J. Neurosci. 29, 5654–5665 (2009).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  84. Sorensen, J. B. et al. Differential control of the releasable vesicle pools by SNAP-25 splice variants and SNAP-23. Cell 114, 75–86 (2003).

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgements

We apologize to the many authors whose publications are not cited directly because of space limitations. Research in our laboratories is supported by grants from the National Institutes of Health (T.W.N. and B.R.G.), the Raymond and Beverly Sackler Fund for the Arts and Sciences (B.R.G.) and the State of Connecticut's Stem Cell Research Fund (B.R.G.).

Author information

Authors and Affiliations

  1. Department of Biochemistry, Center for RNA Molecular Biology, Case Western Reserve University, School of Medicine, 10900 Euclid Avenue, Cleveland, 44106, Ohio, USA

    Timothy W. Nilsen

  2. Department of Genetics and Developmental Biology, University of Connecticut Stem Cell Institute, University of Connecticut Health Center, 263 Farmington Avenue, Farmington, 06030, Connecticut, USA

    Brenton R. Graveley

Authors
  1. Timothy W. Nilsen
    View author publications

    You can also search for this author inPubMed Google Scholar

  2. Brenton R. Graveley
    View author publications

    You can also search for this author inPubMed Google Scholar

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Additional information

Reprints and permissions information is available at http://www.nature.com/reprints. Correspondence should be addressed to T.W.N. (twn@case.edu) or B.R.G. (graveley@neuron.uchc.edu).

Rights and permissions

Reprints and permissions

About this article

Cite this article

Nilsen, T., Graveley, B. Expansion of the eukaryotic proteome by alternative splicing. Nature 463, 457–463 (2010). https://doi.org/10.1038/nature08909

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature08909

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing