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.

  • Letter
  • Published:

Dynamic repertoire of a eukaryotic transcriptome surveyed at single-nucleotide resolution

Nature volume 453, pages 1239–1243 (2008)Cite this article

Abstract

Recent data from several organisms indicate that the transcribed portions of genomes are larger and more complex than expected, and that many functional properties of transcripts are based not on coding sequences but on regulatory sequences in untranslated regions or non-coding RNAs1,2,3,4,5,6,7,8,9. Alternative start and polyadenylation sites and regulation of intron splicing add additional dimensions to the rich transcriptional output10,11. This transcriptional complexity has been sampled mainly using hybridization-based methods under one or few experimental conditions. Here we applied direct high-throughput sequencing of complementary DNAs (RNA-Seq), supplemented with data from high-density tiling arrays, to globally sample transcripts of the fission yeast Schizosaccharomyces pombe, independently from available gene annotations. We interrogated transcriptomes under multiple conditions, including rapid proliferation, meiotic differentiation and environmental stress, as well as in RNA processing mutants to reveal the dynamic plasticity of the transcriptional landscape as a function of environmental, developmental and genetic factors. High-throughput sequencing proved to be a powerful and quantitative method to sample transcriptomes deeply at maximal resolution. In contrast to hybridization, sequencing showed little, if any, background noise and was sensitive enough to detect widespread transcription in >90% of the genome, including traces of RNAs that were not robustly transcribed or rapidly degraded. The combined sequencing and strand-specific array data provide rich condition-specific information on novel, mostly non-coding transcripts, untranslated regions and gene structures, thus improving the existing genome annotation. Sequence reads spanning exon–exon or exon–intron junctions give unique insight into a surprising variability in splicing efficiency across introns, genes and conditions. Splicing efficiency was largely coordinated with transcript levels, and increased transcription led to increased splicing in test genes. Hundreds of introns showed such regulated splicing during cellular proliferation or differentiation.

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

Access options

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: Quantitation of sequence coverage.
Figure 2: Analyses of novel transcripts.
Figure 3: Analyses of transcript structures.
Figure 4: Dynamics of splicing efficiency reflects transcription.

Similar content being viewed by others

Accession codes

Primary accessions

ArrayExpress

Data deposits

Raw data are available from ArrayExpress (http://www.ebi.ac.uk/arrayexpress) under accession numbers E-MTAB-5 (sequence data) and E-MTAB-18 (array data). Transcript data-plots are available from our TranscriptomeViewer at http://www.sanger.ac.uk/PostGenomics/S_pombe/.

References

  1. Yamada, K. et al. Empirical analysis of transcriptional activity in the Arabidopsis genome. Science 302, 842–846 (2003)

    Article  ADS  CAS  PubMed  Google Scholar 

  2. Bertone, P. et al. Global identification of human transcribed sequences with genome tiling arrays. Science 306, 2242–2246 (2004)

    Article  ADS  CAS  PubMed  Google Scholar 

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

    Article  ADS  CAS  PubMed  Google Scholar 

  4. Carninci, P. et al. The transcriptional landscape of the mammalian genome. Science 309, 1559–1563 (2005)

    Article  ADS  CAS  PubMed  Google Scholar 

  5. David, L. et al. A high-resolution map of transcription in the yeast genome. Proc. Natl Acad. Sci. USA 103, 5320–5325 (2006)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  6. Li, L. et al. Genome-wide transcription analyses in rice using tiling microarrays. Nature Genet. 38, 124–129 (2006)

    Article  CAS  PubMed  Google Scholar 

  7. The Encode Project Consortium. Identification and analysis of functional elements in 1% of the human genome by the ENCODE pilot project. Nature 447, 799–816 (2007)

  8. Kapranov, P. et al. RNA maps reveal new RNA classes and a possible function for pervasive transcription. Science 316, 1484–1488 (2007)

    Article  ADS  CAS  PubMed  Google Scholar 

  9. Kapranov, P., Willingham, A. T. & Gingeras, T. R. Genome-wide transcription and the implications for genomic organization. Nature Rev. Genet. 8, 413–423 (2007)

    Article  CAS  PubMed  Google Scholar 

  10. Blencowe, B. J. Alternative splicing: new insights from global analyses. Cell 126, 37–47 (2006)

    Article  CAS  PubMed  Google Scholar 

  11. Hughes, T. A. Regulation of gene expression by alternative untranslated regions. Trends Genet. 22, 119–122 (2006)

    Article  CAS  PubMed  Google Scholar 

  12. Lackner, D. H. et al. A network of multiple regulatory layers shapes gene expression in fission yeast. Mol. Cell 26, 145–155 (2007)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Wood, V. et al. The genome sequence of Schizosaccharomyces pombe . Nature 415, 871–880 (2002)

    Article  ADS  CAS  PubMed  Google Scholar 

  14. Lyne, R. et al. Whole-genome microarrays of fission yeast: characteristics, accuracy, reproducibility, and processing of array data. BMC Genomics 4, 27 (2003)

    Article  PubMed  PubMed Central  Google Scholar 

  15. Chen, D. et al. Global transcriptional responses of fission yeast to environmental stress. Mol. Biol. Cell 14, 214–229 (2003)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Mata, J. & Bähler, J. Correlations between gene expression and gene conservation in fission yeast. Genome Res. 13, 2686–2690 (2003)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Struhl, K. Transcriptional noise and the fidelity of initiation by RNA polymerase II. Nature Struct. Mol. Biol. 14, 103–105 (2007)

    Article  CAS  Google Scholar 

  18. Potashkin, J., Li, R. & Frendewey, D. Pre-mRNA splicing mutants of Schizosaccharomyces pombe . EMBO J. 8, 551–559 (1989)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Leonardi, J., Box, J. A., Bunch, J. T. & Baumann, P. TER1, the RNA subunit of fission yeast telomerase. Nature Struct. Mol. Biol. 15, 26–33 (2008)

    Article  CAS  Google Scholar 

  20. Webb, C. J. & Zakian, V. A. Identification and characterization of the Schizosaccharomyces pombe TER1 telomerase RNA. Nat. Struct. Mol. Biol. 15, 34–42 (2008)

    Article  CAS  PubMed  Google Scholar 

  21. Bickel, K. S. & Morris, D. R. Silencing the transcriptome’s dark matter: mechanisms for suppressing translation of intergenic transcripts. Mol. Cell 22, 309–316 (2006)

    Article  CAS  PubMed  Google Scholar 

  22. Harigaya, Y. et al. Selective elimination of messenger RNA prevents an incidence of untimely meiosis. Nature 442, 45–50 (2006)

    Article  ADS  CAS  PubMed  Google Scholar 

  23. Borgne, A., Murakami, H., Ayté, J. & Nurse, P. The G1/S cyclin Cig2p during meiosis in fission yeast. Mol. Biol. Cell 13, 2080–2090 (2002)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Munoz, M. J., Daga, R. R., Garzon, A., Thode, G. & Jimenez, J. Poly(A) site choice during mRNA 3′-end formation in the Schizosaccharomyces pombe wos2 gene. Mol. Genet. Genomics 267, 792–796 (2002)

    Article  CAS  PubMed  Google Scholar 

  25. Averbeck, N., Sunder, S., Sample, N., Wise, J. A. & Leatherwood, J. Negative control contributes to an extensive program of meiotic splicing in fission yeast. Mol. Cell 18, 491–498 (2005)

    Article  CAS  PubMed  Google Scholar 

  26. Mata, J., Lyne, R., Burns, G. & Bähler, J. The transcriptional program of meiosis and sporulation in fission yeast. Nature Genet. 32, 143–147 (2002)

    Article  CAS  PubMed  Google Scholar 

  27. Mata, J., Wilbrey, A. & Bähler, J. Transcriptional regulatory network for sexual differentiation in fission yeast. Genome Biol. 8, R217 (2007)

    Article  PubMed  PubMed Central  Google Scholar 

  28. Malapeira, J. et al. A meiosis-specific cyclin regulated by splicing is required for proper progression through meiosis. Mol. Cell. Biol. 25, 6330–6337 (2005)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Huber, W., Toedling, J. & Steinmetz, L. M. Transcript mapping with high-density oligonucleotide tiling arrays. Bioinformatics 22, 1963–1970 (2006)

    Article  CAS  PubMed  Google Scholar 

  30. Watt, S. et al. urg1: a uracil-regulatable promoter system for fission yeast with short induction and repression times. PLoS ONE 3, e1428 (2008)

    Article  ADS  PubMed  PubMed Central  Google Scholar 

  31. Bernstein, B. E., Humphrey, E. L., Liu, C. L. & Schreiber, S. L. The use of chromatin immunoprecipitation assays in genome-wide analyses of histone modifications. Methods Enzymol. 376, 349–360 (2004)

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank K. Gould and M. Yamamoto for strains, W. Huber and R. Durbin for advice, and J. Mata, W. Huber, V. Pancaldi, D. Stemple, J.-R. Landry and D. Lackner for comments on the manuscript. B.T.W. was supported by Sanger Postdoctoral and Canadian NSERC fellowships, and S.M. by a fellowship for Advanced Researchers from the Swiss National Science Foundation. This research was funded by Cancer Research UK grant number C9546/A6517 by the Wellcome Trust, and by DIAMONDS, an EC FP6 Lifescihealth STREP (LSHB-CT-2004-512143).

Author Contributions B.T.W., S.M. and J.B. designed and supervised the research and discussed the results; S.W. performed most experiments with help of B.T.W. and S.M.; B.T.W. and S.M. analysed the data with help of F.S., V.W., C.J.P. and J.B.; I.G. and J.R. helped with sequencing; and J.B. drafted the manuscript.

Author information

Author notes

  1. Brian T. Wilhelm, Samuel Marguerat, Stephen Watt, Falk Schubert, Ian Goodhead, Christopher J. Penkett & Jürg Bähler

    Present address: Present addresses: Institut de Recherche en Immunologie et en Cancérologie (IRIC), Montreal, H3C 3J7, Canada (B.T.W.); Department of Genetics, Evolution and Environment, University College London, London WC1E 6BT, UK (S.M., S.W., F.S. and J.B); School of Biological Sciences, University of Liverpool, L69 7ZB, UK (I.G.); EMBL-European Bioinformatics Institute, Hinxton, Cambridge, CB10 1SD, UK (C.J.P.).,

  2. Brian T. Wilhelm and Samuel Marguerat: These authors contributed equally to this work.

Authors and Affiliations

  1. Cancer Research UK Fission Yeast Functional Genomics Group, Wellcome Trust Sanger Institute, Hinxton, Cambridge CB10 1HH, UK ,

    Brian T. Wilhelm, Samuel Marguerat, Stephen Watt, Falk Schubert, Valerie Wood, Ian Goodhead, Christopher J. Penkett, Jane Rogers & Jürg Bähler

Authors
  1. Brian T. Wilhelm
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Samuel Marguerat
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Stephen Watt
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Falk Schubert
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Valerie Wood
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Ian Goodhead
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Christopher J. Penkett
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Jane Rogers
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Jürg Bähler
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jürg Bähler.

Supplementary information

Supplementary Figures

The file contains Supplementary Figures 1 to 7 with Legends (PDF 244 kb)

Table 1

The file contains Supplementary Table 1 (XLS 19 kb)

Table 2

The file contains Supplementary Table 2 (XLS 19 kb)

Table 3

The file contains Supplementary Table 3 (XLS 88 kb)

Table 4

The file contains Supplementary Table 4 (XLS 2247 kb)

Table 5

The file contains Supplementary Table 5 (XLS 69 kb)

Table 6

The file contains Supplementary Table 6 (XLS 45 kb)

Table 7

The file contains Supplementary Table 7 (XLS 847 kb)

Table 8

The file contains Supplementary Table 8 (XLS 60 kb)

Table 9

The file contains Supplementary Table 9 (XLS 17 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Wilhelm, B., Marguerat, S., Watt, S. et al. Dynamic repertoire of a eukaryotic transcriptome surveyed at single-nucleotide resolution. Nature 453, 1239–1243 (2008). https://doi.org/10.1038/nature07002

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature07002

This article is cited by

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