Detecting RNA modifications in the epitranscriptome: predict and validate (original) (raw)

• Frye, M., Jaffrey, S. R., Pan, T., Rechavi, G. & Suzuki, T. RNA modifications: what have we learned and where are we headed? Nat. Rev. Genet. 17, 365–372 (2016).
  Article CAS PubMed Google Scholar
• Saletore, Y. et al. The birth of the epitranscriptome: deciphering the function of RNA modifications. Genome Biol. 13, 175 (2012).
  Article CAS PubMed PubMed Central Google Scholar
• Schwartz, S. Cracking the epitranscriptome. RNA 22, 169–174 (2016).
  Article CAS PubMed PubMed Central Google Scholar
• Witkin, K. L. et al. RNA editing, epitranscriptomics, and processing in cancer progression. Cancer Biol. Ther. 16, 21–27 (2015).
  Article CAS PubMed Google Scholar
• Roundtree, I. A. & He, C. RNA epigenetics — chemical messages for posttranscriptional gene regulation. Curr. Opin. Chem. Biol. 30, 46–51 (2016).
  Article CAS PubMed Google Scholar
• Liu, N. & Pan, T. RNA epigenetics. Transl Res. 165, 28–35 (2015).
  Article CAS PubMed Google Scholar
• Spenkuch, F., Motorin, Y. & Helm, M. Pseudouridine: still mysterious, but never a fake (uridine)! RNA Biol. 11, 1540–1554 (2014).
  Article PubMed Google Scholar
• 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
• Helm, M. & Alfonzo, J. D. Posttranscriptional RNA modifications: playing metabolic games in a cell's chemical Legoland. Chem. Biol. 21, 174–185 (2014).
  Article CAS PubMed Google Scholar
• Holley, R. W. et al. Structure of a ribonucleic acid. Science 147, 1462–1465 (1965).
  Article CAS PubMed Google Scholar
• Ban, E. & Song, E. J. Recent developments and applications of capillary electrophoresis with laser-induced fluorescence detection in biological samples. J. Chromatogr. B Analyt. Technol. Biomed. Life Sci. 929, 180–186 (2013).
  Article CAS PubMed Google Scholar
• Karijolich, J. & Yu, Y. T. Spliceosomal snRNA modifications and their function. RNA Biol. 7, 192–204 (2010).
  Article CAS PubMed Google Scholar
• Zinshteyn, B. & Nishikura, K. Adenosine-to-inosine RNA editing. Wiley Interdiscip. Rev. Syst. Biol. Med. 1, 202–209 (2009).
  Article CAS PubMed PubMed Central Google Scholar
• Yu, B. et al. Methylation as a crucial step in plant microRNA biogenesis. Science 307, 932–935 (2005).
  Article CAS PubMed PubMed Central Google Scholar
• Motorin, Y. & Helm, M. RNA nucleotide methylation. Wiley Interdiscip. Rev. RNA 2, 611–631 (2011).
  Article CAS PubMed Google Scholar
• Moss, B., Gershowitz, A., Stringer, J. R., Holland, L. E. & Wagner, E. K. 5′-terminal and internal methylated nucleosides in herpes simplex virus type 1 mRNA. J. Virol. 23, 234–239 (1977).
  CAS PubMed PubMed Central Google Scholar
• Nichols, J. L. & Welder, L. The modified nucleotide constituents of human prostatic cancer cell (MA-160) poly(A)-containing RNA. Biochim. Biophys. Acta 608, 1–18 (1980).
  Article CAS PubMed Google Scholar
• Rottman, F. M., Desrosiers, R. C. & Friderici, K. Nucleotide methylation patterns in eukaryotic mRNA. Prog. Nucleic Acid. Res. Mol. Biol. 19, 21–38 (1976).
  Article CAS PubMed Google Scholar
• Limbach, P. A. & Paulines, M. J. Going global: the new era of mapping modifications in RNA. Wiley Interdiscip. Rev. RNA 8, e1367 (2016).
  Article Google Scholar
• Song, C. X., Yi, C. & He, C. Mapping recently identified nucleotide variants in the genome and transcriptome. Nat. Biotechnol. 30, 1107–1116 (2012).
  Article CAS PubMed PubMed Central Google Scholar
• Jia, G. et al. _N_6-methyladenosine in nuclear RNA is a major substrate of the obesity-associated FTO. Nat. Chem. Biol. 7, 885–887 (2011). This is the first demonstration of dynamics in mRNA modification by enzymatic demethylation.
  Article CAS PubMed PubMed Central Google Scholar
• Ebhardt, H. A. et al. Meta-analysis of small RNA-sequencing errors reveals ubiquitous post-transcriptional RNA modifications. Nucleic Acids Res. 37, 2461–2470 (2009).
  Article CAS PubMed PubMed Central Google Scholar
• Findeiss, S., Langenberger, D., Stadler, P. F. & Hoffmann, S. Traces of post-transcriptional RNA modifications in deep sequencing data. Biol. Chem. 392, 305–313 (2011).
  Article CAS PubMed Google Scholar
• Ryvkin, P. et al. HAMR: high-throughput annotation of modified ribonucleotides. RNA 19, 1684–1692 (2013).
  Article CAS PubMed PubMed Central Google Scholar
• Alon, S. et al. Systematic identification of edited microRNAs in the human brain. Genome Res. 22, 1533–1540 (2012).
  Article CAS PubMed PubMed Central Google Scholar
• Dominissini, D., Moshitch-Moshkovitz, S., Amariglio, N. & Rechavi, G. Adenosine-to-inosine RNA editing meets cancer. Carcinogenesis 32, 1569–1577 (2011).
  Article CAS PubMed Google Scholar
• 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). This is a combination of the cDNA signature concept with machine learning to predict modification sites.
  CAS PubMed PubMed Central Google Scholar
• Tserovski, L. et al. High-throughput sequencing for 1-methyladenosine (m1A) mapping in RNA. Methods 107, 110–121 (2016).
  Article CAS PubMed Google Scholar
• Gustafsson, C. & Persson, B. C. Identification of the rrmA gene encoding the 23S rRNA m1G745 methyltransferase in Escherichia coli and characterization of an m1G745-deficient mutant. J. Bacteriol. 180, 359–365 (1998).
  CAS PubMed PubMed Central Google Scholar
• Li, X. et al. Transcriptome-wide mapping reveals reversible and dynamic N1-methyladenosine methylome. Nat. Chem. Biol. 12, 311–316 (2016).
  Article CAS PubMed Google Scholar
• Dominissini, D. et al. The dynamic N1-methyladenosine methylome in eukaryotic messenger RNA. Nature 530, 441–446 (2016).
  Article CAS PubMed PubMed Central Google Scholar
• Talkish, J., May, G., Lin, Y., Woolford, J. L. Jr & McManus, C. J. Mod-seq: high-throughput sequencing for chemical probing of RNA structure. RNA 20, 713–720 (2014).
  Article CAS PubMed PubMed Central Google Scholar
• Weeks, K. M. Advances in RNA structure analysis by chemical probing. Curr. Opin. Struct. Biol. 20, 295–304 (2010).
  Article CAS PubMed PubMed Central Google Scholar
• 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). This is pioneering work on the use of selective chemicals for transcriptome-wide detection of modifications.
  Article CAS PubMed Google Scholar
• Suzuki, T., Ueda, H., Okada, S. & Sakurai, M. Transcriptome-wide identification of adenosine-to-inosine editing using the ICE-seq method. Nat. Protoc. 10, 715–732 (2015).
  Article CAS PubMed Google Scholar
• 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
• Schwartz, S. et al. Transcriptome-wide mapping reveals widespread dynamic-regulated pseudouridylation of ncRNA and mRNA. Cell 159, 148–162 (2014).
  Article CAS PubMed PubMed Central Google Scholar
• 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 CAS PubMed PubMed Central Google Scholar
• Carlile, T. M. et al. Pseudouridine profiling reveals regulated mRNA pseudouridylation in yeast and human cells. Nature 515, 143–146 (2014).
  Article CAS PubMed PubMed Central Google Scholar
• Li, X. et al. Chemical pulldown reveals dynamic pseudouridylation of the mammalian transcriptome. Nat. Chem. Biol. 11, 592–597 (2015). In this paper, a chemical pulldown is combined with thorough validation of pseudouridine in mRNA.
  Article CAS PubMed Google Scholar
• Amort, T. et al. Long non-coding RNAs as targets for cytosine methylation. RNA Biol. 10, 1003–1008 (2013).
  Article CAS PubMed Google Scholar
• Schaefer, M., Pollex, T., Hanna, K. & Lyko, F. RNA cytosine methylation analysis by bisulfite sequencing. Nucleic Acids Res. 37, e12 (2009).
  Article CAS PubMed Google Scholar
• 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
• 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
• Burgess, A. L., David, R. & Searle, I. R. Conservation of tRNA and rRNA 5-methylcytosine in the kingdom Plantae. BMC Plant Biol. 15, 199 (2015).
  Article CAS PubMed PubMed Central Google Scholar
• Militello, K. T., Chen, L. M., Ackerman, S. E., Mandarano, A. H. & Valentine, E. L. A map of 5-methylcytosine residues in Trypanosoma brucei tRNA revealed by sodium bisulfite sequencing. Mol. Biochem. Parasitol. 193, 122–126 (2014).
  Article CAS PubMed PubMed Central Google Scholar
• Schaefer, M. RNA 5-methylcytosine analysis by bisulfite sequencing. Methods Enzymol. 560, 297–329 (2015). The adaptation of bisulfite sequencing from DNA to RNA has strongly affected the field by pointing out the possibility for transcriptome-wide studies.
  Article CAS PubMed Google Scholar
• Krueger, F., Kreck, B., Franke, A. & Andrews, S. R. DNA methylome analysis using short bisulfite sequencing data. Nat. Methods 9, 145–151 (2012).
  Article CAS PubMed Google Scholar
• Booth, M. J., Marsico, G., Bachman, M., Beraldi, D. & Balasubramanian, S. Quantitative sequencing of 5-formylcytosine in DNA at single-base resolution. Nat. Chem. 6, 435–440 (2014).
  Article CAS PubMed PubMed Central Google Scholar
• Van Haute, L. et al. Deficient methylation and formylation of mt-tRNAMet wobble cytosine in a patient carrying mutations in NSUN3. Nat. Commun. 7, 12039 (2016). This shows the combined application of three different methods for the detection of m 5 C and derivatives in mitochondrial RNA.
  Article CAS PubMed PubMed Central Google Scholar
• 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 CAS PubMed PubMed Central Google Scholar
• Zheng, G. et al. Efficient and quantitative high-throughput tRNA sequencing. Nat. Methods 12, 835–837 (2015).
  Article CAS PubMed PubMed Central Google Scholar
• Clark, W. C., Evans, M. E., Dominissini, D., Zheng, G. & Pan, T. tRNA base methylation identification and quantification via high-throughput sequencing. RNA 22, 1771–1784 (2016).
  Article CAS PubMed PubMed Central Google Scholar
• Cozen, A. E. et al. ARM-seq: AlkB-facilitated RNA methylation sequencing reveals a complex landscape of modified tRNA fragments. Nat. Methods 12, 879–884 (2015).
  Article CAS PubMed PubMed Central Google Scholar
• Zaringhalam, M. & Papavasiliou, F. N. Pseudouridylation meets next-generation sequencing. Methods 107, 63–72 (2016).
  Article CAS PubMed Google Scholar
• 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
• Jeltsch, A. et al. Mechanism and biological role of Dnmt2 in nucleic acid methylation. RNA Biol. http://dx.doi.org/10.1080/15476286.2016.1191737 (2016).
• Poirier, M. C. Antisera specific for carcinogen-DNA adducts and carcinogen-modified DNA: applications for detection of xenobiotics in biological samples. Mutat. Res. 288, 31–38 (1993).
  Article CAS PubMed Google Scholar
• Strickland, P. T. & Boyle, J. M. Immunoassay of carcinogen-modified DNA. Prog. Nucleic Acid. Res. Mol. Biol. 31, 1–58 (1984).
  Article CAS PubMed Google Scholar
• Sawicki, D. L., Erlanger, B. F. & Beiser, S. M. Immunochemical detection of minor bases in nucleic acids. Science 174, 70–72 (1971).
  Article CAS PubMed Google Scholar
• Vold, B. S., Longmire, M. E. & Keith, D. E. Jr. Thiolation and 2-methylthio- modification of Bacillus subtilis transfer ribonucleic acids. J. Bacteriol. 148, 869–876 (1981).
  CAS PubMed PubMed Central Google Scholar
• Milstone, D. S., Vold, B. S., Glitz, D. G. & Shutt, N. Antibodies to _N_6-(Δ2-isopentenyl) adenosine and its nucleotide: interaction with purified tRNAs and with bases, nucleosides and nucleotides of the isopentenyladenosine family. Nucleic Acids Res. 5, 3439–3455 (1978).
  Article CAS PubMed PubMed Central Google Scholar
• Munns, T. W., Liszewski, M. K. & Hahn, B. H. Antibody-nucleic acid complexes. Antigenic domains within nucleosides as defined by solid-phase immunoassay. Biochemistry 23, 2958–2964 (1984).
  Article CAS PubMed Google Scholar
• Horowitz, S., Horowitz, A., Nilsen, T. W., Munns, T. W. & Rottman, F. M. Mapping of _N_6-methyladenosine residues in bovine prolactin mRNA. Proc. Natl Acad. Sci. USA 81, 5667–5671 (1984). This paper uses an antibody to show the presence of m 6 A in mammalian mRNA.
  Article CAS PubMed PubMed Central Google Scholar
• Dominissini, D. et al. Topology of the human and mouse m6A RNA methylomes revealed by m6A-seq. Nature 485, 201–206 (2012).
  Article CAS PubMed Google Scholar
• Meyer, K. D. et al. Comprehensive analysis of mRNA methylation reveals enrichment in 3′ UTRs and near stop codons. Cell 149, 1635–1646 (2012). References 65 and 66 are two pioneering studies combining antibody-based pulldown and high-throughput sequencing.
  Article CAS PubMed PubMed Central Google Scholar
• Mishima, E. et al. Immuno-northern blotting: detection of RNA modifications by using antibodies against modified nucleosides. PLoS ONE 10, e0143756 (2015).
  Article CAS PubMed PubMed Central Google Scholar
• Delatte, B. et al. RNA biochemistry. Transcriptome- wide distribution and function of RNA hydroxymethylcytosine. Science 351, 282–285 (2016).
  Article CAS PubMed Google Scholar
• 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
• 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
• Jacinto, F. V., Ballestar, E. & Esteller, M. Methyl-DNA immunoprecipitation (MeDIP): hunting down the DNA methylome. Biotechniques 44, 35–43 (2008).
  Article CAS PubMed Google Scholar
• Chen, K. et al. High-resolution N6 -methyladenosine (m6A) map using photo-crosslinking-assisted m6A sequencing. Angew. Chem. Int. Ed. 54, 1587–1590 (2015).
  Article CAS Google Scholar
• Arluison, V., Buckle, M. & Grosjean, H. Pseudouridine synthetase Pus1 of Saccharomyces cerevisiae: kinetic characterisation, tRNA structural requirement and real-time analysis of its complex with tRNA. J. Mol. Biol. 289, 491–502 (1999).
  Article CAS PubMed Google Scholar
• Müller, S. et al. Target recognition, RNA methylation activity and transcriptional regulation of the Dictyostelium discoideum Dnmt2-homologue (DnmA). Nucleic Acids Res. 41, 8615–8627 (2013).
  Article CAS PubMed PubMed Central Google Scholar
• Motorin, Y., Lyko, F. & Helm, M. 5-Methylcytosine in RNA: detection, enzymatic formation and biological functions. Nucleic Acids Res. 38, 1415–1430 (2010).
  Article CAS PubMed Google Scholar
• Hussain, S. et al. NSun2-mediated cytosine-5 methylation of vault noncoding RNA determines its processing into regulatory small RNAs. Cell Rep. 4, 255–261 (2013).
  Article CAS PubMed PubMed Central Google Scholar
• Metodiev, M. D. et al. NSUN4 is a dual function mitochondrial protein required for both methylation of 12S rRNA and coordination of mitoribosomal assembly. PLoS Genet. 10, e1004110 (2014).
  Article CAS PubMed PubMed Central Google Scholar
• 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
• Khoddami, V. & Cairns, B. R. Transcriptome-wide target profiling of RNA cytosine methyltransferases using the mechanism-based enrichment procedure Aza-IP. Nat. Protoc. 9, 337–361 (2014).
  Article CAS PubMed Google Scholar
• Haag, S. et al. NSUN6 is a human RNA methyltransferase that catalyzes formation of m5C72 in specific tRNAs. RNA 21, 1532–1543 (2015).
  Article CAS PubMed PubMed Central Google Scholar
• Hafner, M. et al. Transcriptome-wide identification of RNA-binding protein and microRNA target sites by PAR-CLIP. Cell 141, 129–141 (2010).
  Article CAS PubMed PubMed Central Google Scholar
• Gu, X., Liu, Y. & Santi, D. V. The mechanism of pseudouridine synthase I as deduced from its interaction with 5-fluorouracil-tRNA. Proc. Natl Acad. Sci. USA 96, 14270–14275 (1999).
  Article CAS PubMed PubMed Central Google Scholar
• Spedaliere, C. J. & Mueller, E. G. Not all pseudouridine synthases are potently inhibited by RNA containing 5-fluorouridine. RNA 10, 192–199 (2004).
  Article CAS PubMed PubMed Central Google Scholar
• Cahova, H., Winz, M. L., Hofer, K., Nubel, G. & Jaschke, A. NAD captureSeq indicates NAD as a bacterial cap for a subset of regulatory RNAs. Nature 519, 374–377 (2015).
  Article CAS PubMed Google Scholar
• Kivioja, T. et al. Counting absolute numbers of molecules using unique molecular identifiers. Nat. Methods 9, 72–74 (2011).
  Article CAS PubMed Google Scholar
• Qin, Y. et al. High-throughput sequencing of human plasma RNA by using thermostable group II intron reverse transcriptases. RNA 22, 111–128 (2016).
  Article CAS PubMed PubMed Central Google Scholar
• Nottingham, R. M. et al. RNA-seq of human reference RNA samples using a thermostable group II intron reverse transcriptase. RNA 22, 597–613 (2016).
  Article CAS PubMed PubMed Central Google Scholar
• Kielpinski, L. J., Boyd, M., Sandelin, A. & Vinther, J. Detection of reverse transcriptase termination sites using cDNA ligation and massive parallel sequencing. Methods Mol. Biol. 1038, 213–231 (2013).
  Article CAS PubMed Google Scholar
• Sooknanan, R., Pease, J. & Doyle, K. Novel methods for rRNA removal and directional, ligation-free RNA-seq library preparation. Nat. Methods 7 http://www.nature.com/app_notes/nmeth/2010/101310/full/nmeth.f.313.html (2010).
• 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 CAS PubMed PubMed Central Google Scholar
• Birkedal, U. et al. Profiling of ribose methylations in RNA by high-throughput sequencing. Angew. Chem. Int. Ed. 54, 451–455 (2015).
  CAS Google Scholar
• Taoka, M. et al. The complete chemical structure of Saccharomyces cerevisiae rRNA: partial pseudouridylation of U2345 in 25S rRNA by snoRNA snR9. Nucleic Acids Res. 44, 8951–8961 (2016).
  Article CAS PubMed PubMed Central Google Scholar
• Hafner, M. et al. RNA-ligase-dependent biases in miRNA representation in deep-sequenced small RNA cDNA libraries. RNA 17, 1697–1712 (2011).
  Article CAS PubMed PubMed Central Google Scholar
• Munafo, D. B. & Robb, G. B. Optimization of enzymatic reaction conditions for generating representative pools of cDNA from small RNA. RNA 16, 2537–2552 (2010).
  Article CAS PubMed PubMed Central Google Scholar
• Raabe, C. A. et al. The rocks and shallows of deep RNA sequencing: examples in the Vibrio cholerae RNome. RNA 17, 1357–1366 (2011).
  Article CAS PubMed PubMed Central Google Scholar
• Shen, Y., Zheng, K. X., Duan, D., Jiang, L. & Li, J. Label-free microRNA profiling not biased by 3′ end 2′-O-methylation. Anal. Chem. 84, 6361–6365 (2012).
  Article CAS PubMed Google Scholar
• Mourato, L. L., Beland, F. A. & Marques, M. M. 32P-postlabeling of N-(deoxyguanosin-8-yl)arylamine adducts: a comparative study of labeling efficiencies. Chem. Res. Toxicol. 12, 661–669 (1999).
  Article CAS PubMed Google Scholar
• Sun, W. J. et al. RMBase: a resource for decoding the landscape of RNA modifications from high-throughput sequencing data. Nucleic Acids Res. 44, D259–D265 (2016).
  Article CAS PubMed Google Scholar
• Bantscheff, M., Schirle, M., Sweetman, G., Rick, J. & Kuster, B. Quantitative mass spectrometry in proteomics: a critical review. Anal. Bioanal. Chem. 389, 1017–1031 (2007).
  Article CAS PubMed Google Scholar
• Addepalli, B., Lesner, N. P. & Limbach, P. A. Detection of RNA nucleoside modifications with the uridine-specific ribonuclease MC1 from Momordica charantia. RNA 21, 1746–1756 (2015).
  Article CAS PubMed PubMed Central Google Scholar
• Cao, X. & Limbach, P. A. Enhanced detection of post-transcriptional modifications using a mass-exclusion list strategy for RNA modification mapping by LC-MS/MS. Anal. Chem. 87, 8433–8440 (2015).
  Article CAS PubMed PubMed Central Google Scholar
• Wetzel, C. & Limbach, P. A. Mass spectrometry of modified RNAs: recent developments. Analyst 141, 16–23 (2016).
  Article CAS PubMed Google Scholar
• Ross, R., Cao, X., Yu, N. & Limbach, P. A. Sequence mapping of transfer RNA chemical modifications by liquid chromatography tandem mass spectrometry. Methods 107, 73–78 (2016).
  Article CAS PubMed PubMed Central Google Scholar
• Wetzel, C. & Limbach, P. A. The global identification of tRNA isoacceptors by targeted tandem mass spectrometry. Analyst 138, 6063–6072 (2013).
  Article CAS PubMed Google Scholar
• Kellner, S. et al. Absolute and relative quantification of RNA modifications via biosynthetic isotopomers. Nucleic Acids Res. 42, e142 (2014).
  Article CAS PubMed PubMed Central Google Scholar
• Sakaguchi, Y., Miyauchi, K., Kang, B. I. & Suzuki, T. Nucleoside analysis by hydrophilic interaction liquid chromatography coupled with mass spectrometry. Methods Enzymol. 560, 19–28 (2015).
  Article CAS PubMed Google Scholar
• Fu, L. et al. Simultaneous quantification of methylated cytidine and adenosine in cellular and tissue RNA by nano-flow liquid chromatography-tandem mass spectrometry coupled with the stable isotope-dilution method. Anal. Chem. 87, 7653–7659 (2015).
  Article CAS PubMed PubMed Central Google Scholar
• Cai, W. M. et al. A platform for discovery and quantification of modified ribonucleosides in RNA: application to stress-induced reprogramming of tRNA modifications. Methods Enzymol. 560, 29–71 (2015).
  Article CAS PubMed PubMed Central Google Scholar
• Basanta-Sanchez, M., Temple, S., Ansari, S. A., D'Amico, A. & Agris, P. F. Attomole quantification and global profile of RNA modifications: epitranscriptome of human neural stem cells. Nucleic Acids Res. 44, e26 (2016).
  Article CAS PubMed Google Scholar
• Huang, W. et al. Determination of DNA and RNA methylation in circulating tumor cells by mass spectrometry. Anal. Chem. 88, 1378–1384 (2016).
  Article CAS PubMed Google Scholar
• Helm, M., Florentz, C., Chomyn, A. & Attardi, G. Search for differences in post-transcriptional modification patterns of mitochondrial DNA-encoded wild-type and mutant human tRNALys and tRNALeu(UUR). Nucleic Acids Res. 27, 756–763 (1999).
  Article CAS PubMed PubMed Central Google Scholar
• Suzuki, T. & Suzuki, T. Chaplet column chromatography: isolation of a large set of individual RNAs in a single step. Methods Enzymol. 425, 231–239 (2007).
  Article CAS PubMed Google Scholar
• Sharma, S., Watzinger, P., Kotter, P. & Entian, K. D. Identification of a novel methyltransferase, Bmt2, responsible for the N-1-methyl-adenosine base modification of 25S rRNA in Saccharomyces cerevisiae. Nucleic Acids Res. 41, 5428–5443 (2013).
  Article CAS PubMed PubMed Central Google Scholar
• Buchhaupt, M. et al. Partial methylation at Am100 in 18S rRNA of baker's yeast reveals ribosome heterogeneity on the level of eukaryotic rRNA modification. PLoS ONE 9, e89640 (2014).
  Article CAS PubMed PubMed Central Google Scholar
• Bourgeois, G. et al. Eukaryotic rRNA modification by yeast 5-methylcytosine-methyltransferases and human proliferation-associated antigen p120. PLoS ONE 10, e0133321 (2015).
  Article CAS PubMed PubMed Central Google Scholar
• Kellner, S., Burhenne, J. & Helm, M. Detection of RNA modifications. RNA Biol. 7, 237–247 (2010).
  Article CAS PubMed Google Scholar
• Grosjean, H., Droogmans, L., Roovers, M. & Keith, G. Detection of enzymatic activity of transfer RNA modification enzymes using radiolabeled tRNA substrates. Methods Enzymol. 425, 55–101 (2007).
  Article CAS PubMed Google Scholar
• Keith, G. Mobilities of modified ribonucleotides on two-dimensional cellulose thin-layer chromatography. Biochimie 77, 142–144 (1995).
  Article CAS PubMed Google Scholar
• Stanley, J. & Vassilenko, S. A different approach to RNA sequencing. Nature 274, 87–89 (1978).
  Article CAS PubMed Google Scholar
• Juhling, F. et al. tRNAdb 2009: compilation of tRNA sequences and tRNA genes. Nucleic Acids Res. 37, D159–D162 (2009).
  Article CAS PubMed Google Scholar
• Yu, Y. T., Shu, M. D. & Steitz, J. A. A new method for detecting sites of 2′-O-methylation in RNA molecules. RNA 3, 324–331 (1997).
  CAS PubMed PubMed Central Google Scholar
• Zhao, X. & Yu, Y. T. Detection and quantitation of RNA base modifications. RNA 10, 996–1002 (2004).
  Article CAS PubMed PubMed Central Google Scholar
• Hengesbach, M., Meusburger, M., Lyko, F. & Helm, M. Use of DNAzymes for site-specific analysis of ribonucleotide modifications. RNA 14, 180–187 (2008).
  Article CAS PubMed PubMed Central Google Scholar
• Pyle, A. M., Chu, V. T., Jankowsky, E. & Boudvillain, M. Using DNAzymes to cut, process, and map RNA molecules for structural studies or modification. Methods Enzymol. 317, 140–146 (2000).
  Article CAS PubMed Google Scholar
• Liu, N. et al. Probing _N_6-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
• Liu, N. & Pan, T. Probing RNA modification status at single-nucleotide resolution in total RNA. Methods Enzymol. 560, 149–159 (2015).
  Article CAS PubMed Google Scholar
• Liu, N. & Pan, T. Probing N6-methyladenosine (m6A) RNA modification in total RNA with SCARLET. Methods Mol. Biol. 1358, 285–292 (2016).
  Article CAS PubMed Google Scholar
• Saikia, M. et al. A systematic, ligation-based approach to study RNA modifications. RNA 12, 2025–2033 (2006).
  Article CAS PubMed PubMed Central Google Scholar
• Dai, Q. et al. Identification of recognition residues for ligation-based detection and quantitation of pseudouridine and _N_6-methyladenosine. Nucleic Acids Res. 35, 6322–6329 (2007).
  Article CAS PubMed PubMed Central Google Scholar
• Hiley, S. L. et al. Detection and discovery of RNA modifications using microarrays. Nucleic Acids Res. 33, e2 (2005).
  Article CAS PubMed PubMed Central Google Scholar
• Maden, B. E., Corbett, M. E., Heeney, P. A., Pugh, K. & Ajuh, P. M. Classical and novel approaches to the detection and localization of the numerous modified nucleotides in eukaryotic ribosomal RNA. Biochimie 77, 22–29 (1995).
  Article CAS PubMed Google Scholar
• Aschenbrenner, J. & Marx, A. Direct and site-specific quantification of RNA 2′-O-methylation by PCR with an engineered DNA polymerase. Nucleic Acids Res. 44, 3495–3502 (2016).
  Article CAS PubMed PubMed Central Google Scholar
• 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
• Kellner, S., Kollar, L. B., Ochel, A., Ghate, M. & Helm, M. Structure-function relationship of substituted bromomethylcoumarins in nucleoside specificity of RNA alkylation. PLoS ONE 8, e67945 (2013).
  Article CAS PubMed PubMed Central Google Scholar
• Kellner, S., Seidu-Larry, S., Burhenne, J., Motorin, Y. & Helm, M. A multifunctional bioconjugate module for versatile photoaffinity labeling and click chemistry of RNA. Nucleic Acids Res. 39, 7348–7360 (2011).
  Article CAS PubMed PubMed Central Google Scholar
• 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
• Müller, M. et al. Dynamic modulation of Dnmt2-dependent tRNA methylation by the micronutrient queuine. Nucleic Acids Res. 43, 10952–10962 (2015).
  Article CAS PubMed PubMed Central Google Scholar
• Vilfan, I. D. et al. Analysis of RNA base modification and structural rearrangement by single-molecule real-time detection of reverse transcription. J. Nanobiotechnol. 11, 8 (2013).
  Article CAS Google Scholar
• Wescoe, Z. L., Schreiber, J. & Akeson, M. Nanopores discriminate among five C5-cytosine variants in DNA. J. Am. Chem. Soc. 136, 16582–16587 (2014).
  Article CAS PubMed PubMed Central Google Scholar
• Kellner, S. et al. Profiling of RNA modifications by multiplexed stable isotope labelling. Chem. Commun. (Camb.) 50, 3516–3518 (2014).
  Article CAS Google Scholar
• Wasserstein, R. L. & Lazar, N. A. The ASA's statement on p-values: context, process, and purpose. Am. Stat. 70, 129–133 (2016).
  Article Google Scholar
• Rieder, D., Amort, T., Kugler, E., Lusser, A. & Trajanoski, Z. meRanTK: methylated RNA analysis ToolKit. Bioinformatics 32, 782–785 (2016).
  Article CAS PubMed Google Scholar
• Alon, S. et al. The majority of transcripts in the squid nervous system are extensively recoded by A-to-I RNA editing. eLife 4, e05198 (2015).
  Article CAS PubMed Central Google Scholar
• Hauenschild, R. et al. CoverageAnalyzer (CAn): a tool for inspection of modification signatures in RNA sequencing profiles. Biomolecules 6, 42 (2016).
  Article CAS PubMed Central Google Scholar
• Durairaj, A. & Limbach, P. A. Mass spectrometry of the fifth nucleoside: a review of the identification of pseudouridine in nucleic acids. Anal. Chim. Acta 623, 117–125 (2008).
  Article CAS PubMed PubMed Central Google Scholar
• Duffy, E. E. et al. Tracking distinct RNA populations using efficient and reversible covalent chemistry. Mol. Cell 59, 858–866 (2015).
  Article CAS PubMed PubMed Central Google Scholar
• Ladner, J. E. & Schweizer, M. P. Effects of dilute HCl on yeast tRNAPhe and E. coli tRNA1fMet. Nucleic Acids Res. 1, 183–192 (1974).
  Article CAS PubMed PubMed Central Google Scholar
• Schleich, H. G., Wintermeyer, W. & Zachau, H. G. Replacement of wybutine by hydrazines and its effect on the active conformation of yeast tRNAPhe. Nucleic Acids Res. 5, 1701–1713 (1978).
  Article CAS PubMed PubMed Central Google Scholar
• Peattie, D. A. Direct chemical method for sequencing RNA. Proc. Natl Acad. Sci. USA 76, 1760–1764 (1979).
  Article CAS PubMed PubMed Central Google Scholar
• Wintermeyer, W. & Zachau, H. G. Tertiary structure interactions of 7-methylguanosine in yeast tRNA Phe as studied by borohydride reduction. FEBS Lett. 58, 306–309 (1975).
  Article CAS PubMed Google Scholar
• Xing, F., Hiley, S. L., Hughes, T. R. & Phizicky, E. M. The specificities of four yeast dihydrouridine synthases for cytoplasmic tRNAs. J. Biol. Chem. 279, 17850–17860 (2004).
  Article CAS PubMed Google Scholar
• Plumbridge, J. A., Baumert, H. G., Ehrenberg, M. & Rigler, R. Characterisation of a new, fully active fluorescent derivative of E. coli tRNA Phe. Nucleic Acids Res. 8, 827–843 (1980).
  CAS PubMed PubMed Central Google Scholar
• Watson, B. S. et al. Macromolecular arrangement in the aminoacyl-tRNA.elongation factor Tu.GTP ternary complex. A fluorescence energy transfer study. Biochemistry 34, 7904–7912 (1995).
  Article CAS PubMed Google Scholar
• Igloi, G. L. & Kossel, H. Affinity electrophoresis for monitoring terminal phosphorylation and the presence of queuosine in RNA. Application of polyacrylamide containing a covalently bound boronicacid. Nucleic Acids Res. 13, 6881–6898 (1985).
  Article CAS PubMed PubMed Central Google Scholar
• Igloi, G. L. Interaction of tRNAs and of phosphorothioate-substituted nucleic acids with an organomercurial. Probing the chemical environment of thiolated residues by affinity electrophoresis. Biochemistry 27, 3842–3849 (1988).
  Article CAS PubMed Google Scholar