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  • Review Article
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Epitranscriptome sequencing technologies: decoding RNA modifications

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An Erratum to this article was published on 28 February 2017

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Abstract

In recent years, major breakthroughs in RNA-modification-mediated regulation of gene expression have been made, leading to the emerging field of epitranscriptomics.Our understanding of the distribution, regulation and function of these dynamic RNA modifications is based on sequencing technologies. In this Review, we focus on the major mRNA modifications in the transcriptome of eukaryotic cells: N6-methyladenosine, N6, 2′-O-dimethyladenosine, 5-methylcytidine, 5-hydroxylmethylcytidine, inosine, pseudouridine and N1-methyladenosine. We discuss the sequencing technologies used to profile these epitranscriptomic marks, including scale, resolution, quantitative feature, pre-enrichment capability and the corresponding bioinformatics tools. We also discuss the challenges of epitranscriptome profiling and highlight the prospect of future detection tools. We aim to guide the choice of different detection methods and inspire new ideas in RNA biology.

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Figure 1: Chemical modifications in mRNA.
Figure 2: Transcriptome-wide sequencing methods for m6A.
Figure 3: Detecting m5C in the transcriptome.
Figure 4: ICE-seq.
Figure 5: Transcriptome-wide profiling of Ψ in the eukaryotic cells.
Figure 6: Mapping the m1A methylome.

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Change history

  • 10 February 2017

    In the version of this article initially published, author affiliation numbers were incorrect. Xiaoyu Li originally had affiliation 1; this has been changed to affiliations 1 and 2. Xushen Xiong originally had affiliations 1 and 2; these have been changed to affiliations 1–3. Chengqi Yi originally had affiliations 1 and 3; these have been changed to affiliations 2 and 4. The error has been corrected in the HTML and PDF versions of the article.

References

  1. Machnicka, M.A. et al. MODOMICS: a database of RNA modification pathways—2013 update. Nucleic Acids Res. 41, D262–D267 (2013).

    Article  CAS  PubMed  Google Scholar 

  2. He, C. Grand challenge commentary: RNA epigenetics? Nat. Chem. Biol. 6, 863–865 (2010). This paper termed the scope and mechanisms of dynamic RNA modifications 'RNA epigenetics' for the first time.

    Article  CAS  PubMed  Google Scholar 

  3. Saletore, Y. et al. The birth of the Epitranscriptome: deciphering the function of RNA modifications. Genome Biol. 13, 175 (2012). This paper first termed the multitude of RNA modifications as 'epitranscriptome'.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Fu, Y., Dominissini, D., Rechavi, G. & He, C. Gene expression regulation mediated through reversible m6A RNA methylation. Nat. Rev. Genet. 15, 293–306 (2014).

    Article  CAS  PubMed  Google Scholar 

  5. Liu, N. & Pan, T. N6-methyladenosine–encoded epitranscriptomics. Nat. Struct. Mol. Biol. 23, 98–102 (2016).

    Article  CAS  PubMed  Google Scholar 

  6. Gilbert, W.V., Bell, T.A. & Schaening, C. Messenger RNA modifications: form, distribution, and function. Science 352, 1408–1412 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Meyer, K.D. & Jaffrey, S.R. The dynamic epitranscriptome: N6-methyladenosine and gene expression control. Nat. Rev. Mol. Cell Biol. 15, 313–326 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Lee, M., Kim, B. & Kim, V.N. Emerging roles of RNA modification: m(6)A and U-tail. Cell 158, 980–987 (2014).

    Article  CAS  PubMed  Google Scholar 

  9. Saletore, Y., Chen-Kiang, S. & Mason, C.E. Novel RNA regulatory mechanisms revealed in the epitranscriptome. RNA Biol. 10, 342–346 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Bokar, J.A., Rath-Shambaugh, M.E., Ludwiczak, R., Narayan, P. & Rottman, F. Characterization and partial purification of mRNA N6-adenosine methyltransferase from HeLa cell nuclei. Internal mRNA methylation requires a multisubunit complex. J. Biol. Chem. 269, 17697–17704 (1994).

    Article  CAS  PubMed  Google Scholar 

  11. Liu, J. et al. A METTL3–METTL14 complex mediates mammalian nuclear RNA N6-adenosine methylation. Nat. Chem. Biol. 10, 93–95 (2014).

    Article  CAS  PubMed  Google Scholar 

  12. Wang, Y. et al. N6-methyladenosine modification destabilizes developmental regulators in embryonic stem cells. Nat. Cell Biol. 16, 191–198 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Bokar, J.A., Shambaugh, M.E., Polayes, D., Matera, A.G. & Rottman, F.M. Purification and cDNA cloning of the AdoMet-binding subunit of the human mRNA (N6-adenosine)-methyltransferase. RNA 3, 1233–1247 (1997).

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Ping, X.L. et al. Mammalian WTAP is a regulatory subunit of the RNA N6-methyladenosine methyltransferase. Cell Res. 24, 177–189 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Schwartz, S. et al. Perturbation of m6A writers reveals two distinct classes of mRNA methylation at internal and 5′ sites. Cell Reports 8, 284–296 (2014).

    Article  CAS  PubMed  Google Scholar 

  16. Jia, G. et al. N6-methyladenosine in nuclear RNA is a major substrate of the obesity-associated FTO. Nat. Chem. Biol. 7, 885–887 (2011). This work discovered the first m6A RNA demethylase ('eraser')—FTO, demonstrating reversible RNA methylation in the human transcriptome.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Zheng, G. et al. ALKBH5 is a mammalian RNA demethylase that impacts RNA metabolism and mouse fertility. Mol. Cell 49, 18–29 (2013).

    Article  CAS  PubMed  Google Scholar 

  18. Alarcón, C.R. et al. HNRNPA2B1 is a mediator of m(6)A-dependent nuclear RNA processing events. Cell 162, 1299–1308 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  19. Dominissini, D. et al. Topology of the human and mouse m6A RNA methylomes revealed by m6A-seq. Nature 485, 201–206 (2012). This is an important study that reports the first m6A methylome in the mammalian transcriptome.

    Article  CAS  PubMed  Google Scholar 

  20. Liu, N. et al. N(6)-methyladenosine-dependent RNA structural switches regulate RNA–protein interactions. Nature 518, 560–564 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Meyer, K.D. et al. 5′ UTR m(6)A promotes cap-independent translation. Cell 163, 999–1010 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Wang, X. et al. N6-methyladenosine-dependent regulation of messenger RNA stability. Nature 505, 117–120 (2014).

    Article  PubMed  CAS  Google Scholar 

  23. Wang, X. et al. N(6)-methyladenosine modulates messenger RNA translation efficiency. Cell 161, 1388–1399 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Xiao, W. et al. Nuclear m(6)A reader YTHDC1 regulates mRNA splicing. Mol. Cell 61, 507–519 (2016).

    Article  CAS  PubMed  Google Scholar 

  25. Lichinchi, G. et al. Dynamics of the human and viral m(6)A RNA methylomes during HIV-1 infection of T cells. Nat. Microbiol. 1, 16011 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Tirumuru, N. et al. N(6)-methyladenosine of HIV-1 RNA regulates viral infection and HIV-1 Gag protein expression. eLife 5, e15528 (2016).

    Article  PubMed  PubMed Central  Google Scholar 

  27. Gokhale, N.S. et al. N6-methyladenosine in Flaviviridae viral RNA genomes regulates infection. Cell Host Microbe 20, 654–665 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Kennedy, E.M. et al. Posttranscriptional m(6)A editing of HIV-1 mRNAs enhances viral gene expression. Cell Host Microbe 19, 675–685 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Lichinchi, G. et al. Dynamics of human and viral RNA methylation during zika virus infection. Cell Host Microbe 20, 666–673 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Desrosiers, R., Friderici, K. & Rottman, F. Identification of methylated nucleosides in messenger RNA from Novikoff hepatoma cells. Proc. Natl. Acad. Sci. USA 71, 3971–3975 (1974).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Csepany, T., Lin, A., Baldick, C.J. Jr. & Beemon, K. Sequence specificity of mRNA N6-adenosine methyltransferase. J. Biol. Chem. 265, 20117–20122 (1990).

    Article  CAS  PubMed  Google Scholar 

  32. Narayan, P. & Rottman, F.M. An in vitro system for accurate methylation of internal adenosine residues in messenger RNA. Science 242, 1159–1162 (1988).

    Article  CAS  PubMed  Google Scholar 

  33. Narayan, P., Ludwiczak, R.L., Goodwin, E.C. & Rottman, F.M. Context effects on N6-adenosine methylation sites in prolactin mRNA. Nucleic Acids Res. 22, 419–426 (1994).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Meyer, K.D. et al. Comprehensive analysis of mRNA methylation reveals enrichment in 3′ UTRs and near stop codons. Cell 149, 1635–1646 (2012). The other study that first reports the transcriptome-wide m6A methylome.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Schwartz, S. et al. High-resolution mapping reveals a conserved, widespread, dynamic mRNA methylation program in yeast meiosis. Cell 155, 1409–1421 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Chen, K. et al. High-resolution N(6) -methyladenosine (m(6)A) map using photo-crosslinking-assisted m(6)A sequencing. Angew. Chem. Int. Edn Engl. 54, 1587–1590 (2015).

    Article  CAS  Google Scholar 

  37. Ke, S. et al. A majority of m6A residues are in the last exons, allowing the potential for 3′ UTR regulation. Genes Dev. 29, 2037–2053 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Linder, B. et al. Single-nucleotide-resolution mapping of m6A and m6Am throughout the transcriptome. Nat. Methods 12, 767–772 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Liu, N. et al. Probing N6-methyladenosine RNA modification status at single nucleotide resolution in mRNA and long noncoding RNA. RNA 19, 1848–1856 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Molinie, B. et al. m(6)A-LAIC-seq reveals the census and complexity of the m(6)A epitranscriptome. Nat. Methods 13, 692–698 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Schibler, U. & Perry, R.P. The 5′-termini of heterogeneous nuclear RNA: a comparison among molecules of different sizes and ages. Nucleic Acids Res. 4, 4133–4149 (1977).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Keith, J.M., Ensinger, M.J. & Mose, B. HeLa cell RNA (2′-O-methyladenosine-N6-)-methyltransferase specific for the capped 5′-end of messenger RNA. J. Biol. Chem. 253, 5033–5039 (1978).

    Article  CAS  PubMed  Google Scholar 

  43. Wei, C., Gershowitz, A. & Moss, B. N6, O2′-dimethyladenosine a novel methylated ribonucleoside next to the 5′ terminal of animal cell and virus mRNAs. Nature 257, 251–253 (1975).

    Article  CAS  PubMed  Google Scholar 

  44. Munns, T.W., Oberst, R.J., Sims, H.F. & Liszewski, M.K. Antibody-nucleic acid complexes. Immunospecific recognition of 7-methylguanine- and N6-methyladenine-containing 5′-terminal oligonucleotides of mRNA. J. Biol. Chem. 254, 4327–4330 (1979).

    Article  CAS  PubMed  Google Scholar 

  45. Motorin, Y. & Helm, M. tRNA stabilization by modified nucleotides. Biochemistry 49, 4934–4944 (2010).

    Article  CAS  PubMed  Google Scholar 

  46. Squires, J.E. & Preiss, T. Function and detection of 5-methylcytosine in eukaryotic RNA. Epigenomics 2, 709–715 (2010).

    Article  CAS  PubMed  Google Scholar 

  47. Chow, C.S., Lamichhane, T.N. & Mahto, S.K. Expanding the nucleotide repertoire of the ribosome with post-transcriptional modifications. ACS Chem. Biol. 2, 610–619 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Brzezicha, B. et al. Identification of human tRNA:m5C methyltransferase catalysing intron-dependent m5C formation in the first position of the anticodon of the pre-tRNA Leu (CAA). Nucleic Acids Res. 34, 6034–6043 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Goll, M.G. et al. Methylation of tRNAAsp by the DNA methyltransferase homolog Dnmt2. Science 311, 395–398 (2006).

    Article  CAS  PubMed  Google Scholar 

  50. Schaefer, M., Pollex, T., Hanna, K. & Lyko, F. RNA cytosine methylation analysis by bisulfite sequencing. Nucleic Acids Res. 37, e12 (2009).

    Article  PubMed  CAS  Google Scholar 

  51. Squires, J.E. et al. Widespread occurrence of 5-methylcytosine in human coding and non-coding RNA. Nucleic Acids Res. 40, 5023–5033 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Hussain, S., Aleksic, J., Blanco, S., Dietmann, S. & Frye, M. Characterizing 5-methylcytosine in the mammalian epitranscriptome. Genome Biol. 14, 215 (2013).

    Article  PubMed  PubMed Central  Google Scholar 

  53. Shafik, A., Schumann, U., Evers, M., Sibbritt, T. & Preiss, T. The emerging epitranscriptomics of long noncoding RNAs. Biochim. Biophys. Acta 1859, 59–70 (2016).

    Article  CAS  PubMed  Google Scholar 

  54. Edelheit, S., Schwartz, S., Mumbach, M.R., Wurtzel, O. & Sorek, R. Transcriptome-wide mapping of 5-methylcytidine RNA modifications in bacteria, archaea, and yeast reveals m5C within archaeal mRNAs. PLoS Genet. 9, e1003602 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Khoddami, V. & Cairns, B.R. Identification of direct targets and modified bases of RNA cytosine methyltransferases. Nat. Biotechnol. 31, 458–464 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Hussain, S. et al. NSun2-mediated cytosine-5 methylation of vault noncoding RNA determines its processing into regulatory small RNAs. Cell Reports 4, 255–261 (2013).

    Article  CAS  PubMed  Google Scholar 

  57. Kriaucionis, S. & Heintz, N. The nuclear DNA base 5-hydroxymethylcytosine is present in Purkinje neurons and the brain. Science 324, 929–930 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Tahiliani, M. et al. Conversion of 5-methylcytosine to 5-hydroxymethylcytosine in mammalian DNA by MLL partner TET1. Science 324, 930–935 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  59. Li, S. & Mason, C.E. The pivotal regulatory landscape of RNA modifications. Annu. Rev. Genomics Hum. Genet. 15, 127–150 (2014).

    Article  CAS  PubMed  Google Scholar 

  60. Rácz, I., Király, I. & Lásztily, D. Effect of light on the nucleotide composition of rRNA of wheat seedlings. Planta 142, 263–267 (1978).

    Article  PubMed  Google Scholar 

  61. Delatte, B. et al. RNA biochemistry. Transcriptome-wide distribution and function of RNA hydroxymethylcytosine. Science 351, 282–285 (2016).

    Article  CAS  PubMed  Google Scholar 

  62. Fu, L. et al. Tet-mediated formation of 5-hydroxymethylcytosine in RNA. J. Am. Chem. Soc. 136, 11582–11585 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  63. Huber, S.M. et al. Formation and abundance of 5-hydroxymethylcytosine in RNA. ChemBioChem 16, 752–755 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  64. Booth, M.J. et al. Quantitative sequencing of 5-methylcytosine and 5-hydroxymethylcytosine at single-base resolution. Science 336, 934–937 (2012).

    Article  CAS  PubMed  Google Scholar 

  65. Yu, M. et al. Base-resolution analysis of 5-hydroxymethylcytosine in the mammalian genome. Cell 149, 1368–1380 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  66. Nishikura, K. Functions and regulation of RNA editing by ADAR deaminases. Annu. Rev. Biochem. 79, 321–349 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  67. Bass, B.L. RNA editing by adenosine deaminases that act on RNA. Annu. Rev. Biochem. 71, 817–846 (2002).

    Article  CAS  PubMed  Google Scholar 

  68. Jepson, J.E. & Reenan, R.A. RNA editing in regulating gene expression in the brain. Biochim. Biophys. Acta 1779, 459–470 (2008).

    Article  CAS  PubMed  Google Scholar 

  69. Levanon, E.Y. et al. Systematic identification of abundant A-to-I editing sites in the human transcriptome. Nat. Biotechnol. 22, 1001–1005 (2004).

    Article  CAS  PubMed  Google Scholar 

  70. Athanasiadis, A., Rich, A. & Maas, S. Widespread A-to-I RNA editing of alu-containing mRNAs in the human transcriptome. PLoS Biol. 2, e391 (2004).

    Article  PubMed  PubMed Central  Google Scholar 

  71. Wulff, B.E., Sakurai, M. & Nishikura, K. Elucidating the inosinome: global approaches to adenosine-to-inosine RNA editing. Nat. Rev. Genet. 12, 81–85 (2011).

    Article  CAS  PubMed  Google Scholar 

  72. Kim, D.D. et al. Widespread RNA editing of embedded alu elements in the human transcriptome. Genome Res. 14, 1719–1725 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  73. Li, J.B. et al. Genome-wide identification of human RNA editing sites by parallel DNA capturing and sequencing. Science 324, 1210–1213 (2009).

    Article  CAS  PubMed  Google Scholar 

  74. Li, M. et al. Widespread RNA and DNA sequence differences in the human transcriptome. Science 333, 53–58 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  75. Lin, W., Piskol, R., Tan, M.H. & Li, J.B. Comment on “Widespread RNA and DNA sequence differences in the human transcriptome”. Science 335, 1302-e (2012).

    Article  CAS  Google Scholar 

  76. Pickrell, J.K., Gilad, Y. & Pritchard, J.K. Comment on “Widespread RNA and DNA sequence differences in the human transcriptome”. Science 335, 1302-d (2012).

    Article  CAS  Google Scholar 

  77. Kleinman, C.L. & Majewski, J. Comment on “Widespread RNA and DNA sequence differences in the human transcriptome”. Science 335, 1302-c (2012).

    Article  CAS  Google Scholar 

  78. Ju, Y.S. et al. Extensive genomic and transcriptional diversity identified through massively parallel DNA and RNA sequencing of eighteen Korean individuals. Nat. Genet. 43, 745–752 (2011).

    Article  CAS  PubMed  Google Scholar 

  79. Bahn, J.H. et al. Accurate identification of A-to-I RNA editing in human by transcriptome sequencing. Genome Res. 22, 142–150 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  80. Ramaswami, G. et al. Accurate identification of human Alu and non-Alu RNA editing sites. Nat. Methods 9, 579–581 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  81. Peng, Z. et al. Comprehensive analysis of RNA-Seq data reveals extensive RNA editing in a human transcriptome. Nat. Biotechnol. 30, 253–260 (2012).

    Article  CAS  PubMed  Google Scholar 

  82. Ramaswami, G. et al. Identifying RNA editing sites using RNA sequencing data alone. Nat. Methods 10, 128–132 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  83. Sakurai, M., Yano, T., Kawabata, H., Ueda, H. & Suzuki, T. Inosine cyanoethylation identifies A-to-I RNA editing sites in the human transcriptome. Nat. Chem. Biol. 6, 733–740 (2010).

    Article  CAS  PubMed  Google Scholar 

  84. Sakurai, M. et al. A biochemical landscape of A-to-I RNA editing in the human brain transcriptome. Genome Res. 24, 522–534 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  85. Karijolich, J., Yi, C. & Yu, Y.T. Transcriptome-wide dynamics of RNA pseudouridylation. Nat. Rev. Mol. Cell Biol. 16, 581–585 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  86. Kiss, T., Fayet-Lebaron, E. & Jády, B.E. Box H/ACA small ribonucleoproteins. Mol. Cell 37, 597–606 (2010).

    Article  PubMed  Google Scholar 

  87. Hamma, T. & Ferré-D'Amaré, A.R. Pseudouridine synthases. Chem. Biol. 13, 1125–1135 (2006).

    Article  CAS  PubMed  Google Scholar 

  88. Li, X., Ma, S. & Yi, C. Pseudouridine: the fifth RNA nucleotide with renewed interests. Curr. Opin. Chem. Biol. 33, 108–116 (2016).

    Article  CAS  PubMed  Google Scholar 

  89. Decatur, W.A. & Fournier, M.J. rRNA modifications and ribosome function. Trends Biochem. Sci. 27, 344–351 (2002).

    Article  CAS  PubMed  Google Scholar 

  90. Baudin-Baillieu, A. et al. Nucleotide modifications in three functionally important regions of the Saccharomyces cerevisiae ribosome affect translation accuracy. Nucleic Acids Res. 37, 7665–7677 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  91. Jack, K. et al. rRNA pseudouridylation defects affect ribosomal ligand binding and translational fidelity from yeast to human cells. Mol. Cell 44, 660–666 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  92. Gilham, P.T. An addition reaction specific for uridine and guanosine nucleotides and its application to the modification of ribonuclease action. J. Am. Chem. Soc. 84, 687–688 (1962).

    Article  CAS  Google Scholar 

  93. Bakin, A. & Ofengand, J. Four newly located pseudouridylate residues in Escherichia coli 23S ribosomal RNA are all at the peptidyltransferase center: analysis by the application of a new sequencing technique. Biochemistry 32, 9754–9762 (1993).

    Article  CAS  PubMed  Google Scholar 

  94. Schwartz, S. et al. Transcriptome-wide mapping reveals widespread dynamic-regulated pseudouridylation of ncRNA and mRNA. Cell 159, 148–162 (2014). This work reports Ψ-seq to map Ψ sites in the yeast and human transcriptome.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  95. Carlile, T.M. et al. Pseudouridine profiling reveals regulated mRNA pseudouridylation in yeast and human cells. Nature 515, 143–146 (2014). This study reports Pseudo-seq to map Ψ sites in the yeast and human transcriptome.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  96. Lovejoy, A.F., Riordan, D.P. & Brown, P.O. Transcriptome-wide mapping of pseudouridines: pseudouridine synthases modify specific mRNAs in S. cerevisiae. PLoS One 9, e110799 (2014).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  97. Li, X. et al. Chemical pulldown reveals dynamic pseudouridylation of the mammalian transcriptome. Nat. Chem. Biol. 11, 592–597 (2015). This study reports CeU-Seq for the comprehensive identification of Ψ sites in the human and mouse transcriptome.

    Article  CAS  PubMed  Google Scholar 

  98. Zaringhalam, M. & Papavasiliou, F.N. Pseudouridylation meets next-generation sequencing. Methods 107, 63–72 (2016).

    Article  CAS  PubMed  Google Scholar 

  99. Kellner, S., Burhenne, J. & Helm, M. Detection of RNA modifications. RNA Biol. 7, 237–247 (2010).

    Article  CAS  PubMed  Google Scholar 

  100. Behm-Ansmant, I., Helm, M. & Motorin, Y. Use of specific chemical reagents for detection of modified nucleotides in RNA. J. Nucleic Acids 2011, 408053 (2011).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  101. Ozanick, S., Krecic, A., Andersland, J. & Anderson, J.T. The bipartite structure of the tRNA m1A58 methyltransferase from S. cerevisiae is conserved in humans. RNA 11, 1281–1290 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  102. Chujo, T. & Suzuki, T. Trmt61B is a methyltransferase responsible for 1-methyladenosine at position 58 of human mitochondrial tRNAs. RNA 18, 2269–2276 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  103. Schevitz, R.W. et al. Crystal structure of a eukaryotic initiator tRNA. Nature 278, 188–190 (1979).

    Article  CAS  PubMed  Google Scholar 

  104. Vilardo, E. et al. A subcomplex of human mitochondrial RNase P is a bifunctional methyltransferase—extensive moonlighting in mitochondrial tRNA biogenesis. Nucleic Acids Res. 40, 11583–11593 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  105. Hauenschild, R. et al. The reverse transcription signature of N-1-methyladenosine in RNA-Seq is sequence dependent. Nucleic Acids Res. 43, 9950–9964 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  106. Waku, T. et al. NML-mediated rRNA base methylation links ribosomal subunit formation to cell proliferation in a p53-dependent manner. J. Cell Sci. 129, 2382–2393 (2016).

    CAS  PubMed  Google Scholar 

  107. Dominissini, D. et al. The dynamic N(1)-methyladenosine methylome in eukaryotic messenger RNA. Nature 530, 441–446 (2016). This study reports the transcriptome-wide m1A methylome for the first time.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  108. Li, X. et al. Transcriptome-wide mapping reveals reversible and dynamic N(1)-methyladenosine methylome. Nat. Chem. Biol. 12, 311–316 (2016). This work reports the transcriptome-wide m1A methylome for the first time.

    Article  CAS  PubMed  Google Scholar 

  109. Garalde, D.R. et al. Highly parallel direct RNA sequencing on an array of nanopores. Preprint at http://biorxiv.org/content/early/2016/08/12/068809 (2016).

  110. Dong, Z.W. et al. RTL-P: a sensitive approach for detecting sites of 2′-O-methylation in RNA molecules. Nucleic Acids Res. 40, e157 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  111. Lacoux, C. et al. BC1-FMRP interaction is modulated by 2′-O-methylation: RNA-binding activity of the tudor domain and translational regulation at synapses. Nucleic Acids Res. 40, 4086–4096 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  112. Birkedal, U. et al. Profiling of ribose methylations in RNA by high-throughput sequencing. Angew. Chem. Int. Edn Engl. 54, 451–455 (2015).

    CAS  Google Scholar 

  113. Gumienny, R., Jedlinski, D., Martin, G., Vina-Villaseca, A. & Zavolan, M. High-throughput identification of C/D box snoRNA targets with CLIP and RiboMeth-seq. Preprint at http://biorxiv.org/content/early/2016/01/19/037259 (2016).

  114. Incarnato, D. et al. High-throughput single-base resolution mapping of RNA 2′-O-methylated residues. Nucleic Acids Res. Sep 9, gkw810 (2016).

    Google Scholar 

  115. Marchand, V., Blanloeil-Oillo, F., Helm, M. & Motorin, Y. Illumina-based RiboMethSeq approach for mapping of 2′-O-Me residues in RNA. Nucleic Acids Res. 44, e135 (2016).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

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Acknowledgements

We thank all members of the Yi laboratory for their insights and discussions. This work was supported by the National Basic Research Foundation of China (grant nos. MOST2016YFC0900300 and 2014CB964900) and the National Natural Science Foundation of China (grant no. 21522201).

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Li, X., Xiong, X. & Yi, C. Epitranscriptome sequencing technologies: decoding RNA modifications. Nat Methods 14, 23–31 (2017). https://doi.org/10.1038/nmeth.4110

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