Brief Explanation of Ariadne

  1. Introduction
  2. Data and Parameters for Ariadne Search
  3. References

1. Introduction

Accumulating evidence shows that diverse types of RNAs generated from non-coding (nc) regions of the genome play pivotal roles in a variety of cellular processes, such as chromatin remodeling, transcriptional regulation, precursor mRNA processing, gene silencing, centromere function and translational regulation. Many RNAs suffer chemical modifications including methylations after their transcription. Some of these post-transcriptional modifications (PTMs) play important roles in RNA structure / function.


A work flow similar to bottom-up proteomics is used to identify / characterize RNAs and their PTMs by mass spectrometry, which includes (i) RNA separation and purification with various methods, (ii) nucleolytic fragmentation of sample RNA with a sequence specific RNase such as RNase T1, (iii) liquid charomatogarphy (LC)-tandem mass spectrometry (MS/MS) of the resulting fragments and (iv) analysis of the MS/MS dataset by manual and/or software-aided procedures.


Ariadne, a unique web-based database search engine avaiable freely here, is designed to help researchers analyze their MS/MS data of RNAs (the 4th step above). The software correlates tandem mass spectra of sample RNA nucleolytic fragments with an RNA nucleotide sequence in a DNA/RNA sequence database, thereby allowing MS/MS-based identification of RNA in biological samples. Ariadne identifies RNA by two probability-based evaluation steps of MS/MS data. In the first step, the software evaluates the matches between the masses of product ions generated by MS/MS of an RNase digest of sample RNA and those calculated from a candidate nucleotide sequence in a DNA/RNA sequence database, which then predicts the nucleotide sequences of these RNase fragments. In the second step, the candidate sequences are mapped for all RNA entries in the database, and each entry is scored for a function of occurrences of the candidate sequences to identify a particular RNA. Ariadne also predicts post-transcriptional modifications of RNA, such as methylation of nucleotide bases and/or ribose, by estimating mass shifts from the theoretical mass values.


We examined the performance of the LC-MS/MS-Ariadne method with a number of applications to identify mRNA synthesized in vitro as well as small and large non-coding RNAs isolated from biological samples. Ariadne is the first genome-oriented search engine for RNA analysis that could be equivalent to the many sequence search engines that are widely used for proteomics, such as SEQUEST or Mascot.


2. Data and Paramerters for Search

Ariadne receives search parameters including uninterpreted MS/MS data, a sequence database, and other parameters in text format from the search form. See details in 'Tutorials' section on the top page.


3. References

Our papers in which Ariadne is reported or used are listed below.

  1. Nakayama H, Akiyama M, Taoka M, Yamauchi Y, Nobe Y, Ishikawa H, Takahashi N, Isobe T. Ariadne: a database search engine for identification and chemical analysis of RNA using tandem mass spectrometry data. Nucleic Acids Res. 2009 Apr;37(6):e47. Epub 2009 Mar 6.
  2. Taoka M, Yamauchi Y, Nobe Y, Masaki S, Nakayama H, Ishikawa H, Takahashi N, Isobe T. An analytical platform for mass spectrometry-based identification and chemical analysis of RNA in ribonucleoprotein complexes. Nucleic Acids Res. 2009 Nov;37(21):e140.
  3. Taoka M, Ikumi M, Nakayama H, Masaki S, Matsuda R, Nobe Y, Yamauchi Y, Takeda J, Takahashi N, Isobe T. In-gel digestion for mass spectrometric characterization of RNA from fluorescently stained polyacrylamide gels. Anal Chem. 2010 Sep 15;82(18):7795-803. doi: 10.1021/ac101623j. PubMed PMID: 20795640.
  4. Nakayama H, Takahashi N, Isobe T. Informatics for mass spectrometry-based RNA analysis. Mass Spectrom Rev. 2011 Nov-Dec;30(6):1000-12. doi: 10.1002/mas.20325. Epub 2011 Feb 16. Review. PubMed PMID: 21328601.
  5. Yamauchi Y, Taoka M, Nobe Y, Izumikawa K, Takahashi N, Nakayama H, Isobe T. Denaturing reversed phase liquid chromatographic separation of non-coding ribonucleic acids on macro-porous polystyrene-divinylbenzene resins. J Chromatogr A. 2013 Oct 18;1312:87-92. doi: 10.1016/j.chroma.2013.09.021. Epub 2013 Sep 9. PubMed PMID: 24044980.
  6. Ishikawa H, Nobe Y, Izumikawa K, Yoshikawa H, Miyazawa N, Terukina G, Kurokawa N, Taoka M, Yamauchi Y, Nakayama H, Isobe T, Takahashi N. Identification of truncated forms of U1 snRNA reveals a novel RNA degradation pathway during snRNP biogenesis. Nucleic Acids Res. 2014 Feb;42(4):2708-24. doi: 10.1093/nar/gkt1271. Epub 2013 Dec 5. PubMed PMID: 24311566; PubMed Central PMCID: PMC3936765.
  7. Taoka M, Ishikawa D, Nobe Y, Ishikawa H, Yamauchi Y, Terukina G, Nakayama H, Hirota K, Takahashi N, Isobe T. RNA cytidine acetyltransferase of small-subunit ribosomal RNA: identification of acetylation sites and the responsible acetyltransferase in fission yeast, Schizosaccharomyces pombe. PLoS One. 2014 Nov 17;9(11):e112156. doi: 10.1371/journal.pone.0112156. eCollection 2014. PubMed PMID: 25402480; PubMed Central PMCID: PMC4234376.
  8. Nakayama H, Yamauchi Y, Taoka M, Isobe T. Direct identification of human cellular microRNAs by nanoflow liquid chromatography-high-resolution tandem mass spectrometry and database searching. Anal Chem. 2015 Mar 3;87(5):2884-91. doi: 10.1021/ac504378s. Epub 2015 Feb 18. PubMed PMID: 25662820.
  9. Taoka M, Nobe Y, Hori M, Takeuchi A, Masaki S, Yamauchi Y, Nakayama H, Takahashi N, Isobe T. A mass spectrometry-based method for comprehensive quantitative determination of post-transcriptional RNA modifications: the complete chemical structure of Schizosaccharomyces pombe ribosomal RNAs. Nucleic Acids Res. 2015 Oct 15;43(18):e115. doi: 10.1093/nar/gkv560. Epub 2015 May 26. PubMed PMID: 26013808; PubMed Central PMCID: PMC4605285.
  10. Yamauchi Y, Nobe Y, Izumikawa K, Higo D, Yamagishi Y, Takahashi N, Nakayama H, Isobe T, Taoka M. A mass spectrometry-based method for direct determination of pseudouridine in RNA. Nucleic Acids Res. 2016 Apr 7;44(6):e59. doi: 10.1093/nar/gkv1462. Epub 2015 Dec 15. PubMed PMID: 26673725; PubMed Central PMCID: PMC4824092.
  11. Taoka M, Nobe Y, Yamaki Y, Yamauchi Y, Ishikawa H, Takahashi N, Nakayama H, Isobe T. The complete chemical structure of Saccharomyces cerevisiae rRNA: partial pseudouridylation of U2345 in 25S rRNA by snoRNA snR9. Nucleic Acids Res. 2016 Oct 14;44(18):8951-8961. Epub 2016 Jun 20. PubMed PMID: 27325748; PubMed Central PMCID: PMC5062969.
  12. Ishikawa H, Yoshikawa H, Izumikawa K, Miura Y, Taoka M, Nobe Y, Yamauchi Y, Nakayama H, Simpson RJ, Isobe T, Takahashi N. Poly(A)-specific ribonuclease regulates the processing of small-subunit rRNAs in human cells. Nucleic Acids Res. 2017 Apr 7;45(6):3437-3447. doi: 10.1093/nar/gkw1047. PubMed PMID: 27899605; PubMed Central PMCID: PMC5389690.
  13. Izumikawa K, Nobe Y, Yoshikawa H, Ishikawa H, Miura Y, Nakayama H, Nonaka T, Hasegawa M, Egawa N, Inoue H, Nishikawa K, Yamano K, Simpson RJ, Taoka M, Yamauchi Y, Isobe T, Takahashi N. TDP-43 stabilises the processing intermediates of mitochondrial transcripts. Sci Rep. 2017 Aug 9;7(1):7709. doi: 10.1038/s41598-017-06953-y. PubMed PMID: 28794432; PubMed Central PMCID: PMC5550480.
  14. Shalev-Benami M, Zhang Y, Rozenberg H, Nobe Y, Taoka M, Matzov D, Zimmerman E, Bashan A, Isobe T, Jaffe CL, Yonath A, Skiniotis G. Atomic resolution snapshot of Leishmania ribosome inhibition by the aminoglycoside paromomycin. Nat Commun. 2017 Nov 17;8(1):1589. doi: 10.1038/s41467-017-01664-4. PMID: 29150609.
  15. Ishikawa H, Nobe Y, Izumikawa K, Taoka M, Yamauchi Y, Nakayama H, Simpson RJ, Isobe T, Takahash N. Truncated forms of U2 snRNA (U2-tfs) are shunted toward a novel uridylylation pathway that differs from the degradation pathway for U1-tfs. RNA Biol. 2018 Feb 1;15(2):261-268. doi: 10.1080/15476286.2017.1408766. Epub 2017 Dec 15. PubMed PMID: 29168419; PubMed Central PMCID: PMC5798949.
  16. Taoka M, Nobe Y, Yamaki Y, Sato K, Ishikawa H, Izumikawa K, Yamauchi Y, Hirota K, Nakayama H, Takahashi N, Isobe T. Landscape of the complete RNA chemical modifications in the human 80S ribosome. Nucleic Acids Res. 2018 Oct 12;46(18):9289-9298. doi: 10.1093/nar/gky811. PubMed PMID: 30202881; PubMed Central PMCID: PMC6182160.
  17. Izumikawa K, Nobe Y, Ishikawa H, Yamauchi Y, Taoka M, Sato K, Nakayama H, Simpson RJ, Isobe T, Takahashi N. TDP-43 regulates site-specific 2'-O-methylation of U1 and U2 snRNAs via controlling the Cajal body localization of a subset of C/D scaRNAs. Nucleic Acids Res. 2019 Mar 18;47(5):2487-2505. doi: 10.1093/nar/gkz086. PubMed PMID: 30759234; PubMed Central PMCID: PMC6412121.
  18. Nakayama H, Yamauchi Y, Nobe Y, Sato K, Takahashi N, Shalev-Benami M, Isobe T, Taoka M. Method for Direct Mass-Spectrometry-Based Identification of Monomethylated RNA Nucleoside Positional Isomers and Its Application to the Analysis of Leishmania rRNA. Anal Chem. 2019 Dec 17;91(24):15634-15643. doi: 10.1021/acs.analchem.9b03735. Epub 2019 Dec 5. PMID: 31725277
  19. Sas-Chen A, Thomas JM, Matzov D, Taoka M, Nance KD, Nir R, Bryson KM, Shachar R, Liman GLS, Burkhart BW, Gamage ST, Nobe Y, Briney CA, Levy MJ, Fuchs RT, Robb GB, Hartmann J, Sharma S, Lin Q, Florens L, Washburn MP, Isobe T, Santangelo TJ, Shalev-Benami M, Meier JL, Schwartz S. Dynamic RNA acetylation revealed by quantitative cross-evolutionary mapping. Nature. 2020 Jul;583(7817):638-643. doi: 10.1038/s41586-020-2418-2. Epub 2020 Jun 17. PMID: 32555463.
  20. Yamaki Y, Nobe Y, Koike M, Yamauchi Y, Hirota K, Takahashi N, Nakayama H, Isobe T, Taoka M. Direct Determination of Pseudouridine in RNA by Mass Spectrometry Coupled with Stable Isotope Labeling. Anal Chem. 2020 Aug 18;92(16):11349-11356. doi: 10.1021/acs.analchem.0c02122. Epub 2020 Jul 28. PMID: 32662983.
  21. Nakayama H, Nobe Y, Koike M, Taoka M. Liquid Chromatography-Mass Spectrometry-Based Qualitative Profiling of mRNA Therapeutic Reagents Using Stable Isotope-Labeled Standards Followed by the Automatic Quantitation Software Ariadne. Anal Chem. 2023 Jan 17;95(2):1366-1375. doi: 10.1021/acs.analchem.2c04323. Epub 2022 Dec 27. PMID: 36574727

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