RNA Editing pp 213-227 | Cite as

Detection of A-to-I Hyper-edited RNA Sequences

Part of the Methods in Molecular Biology book series (MIMB, volume 2181)


Following A-to-I editing of double-stranded RNA (dsRNA) molecules, sequencing reactions interpret the edited inosine (I) as guanosine (G). For this reason, current methods to detect A-to-I editing sites work to align RNA sequences to their reference DNA sequence in order to reveal A-to-G mismatches. However, areas with heavily edited reads produce dense clusters of A-to-G mismatches that hinder alignment, and complicate correct identification of the sites. The presented approach employs prudent alignment and examination of excessive mismatch events, enabling high-accuracy detection of hyper-edited reads and sites.

Key words

RNA editing Hyper-editing ADAR 



The authors thank H.T. Porath and the Levanon laboratory members for fruitful discussions. EYL was supported by the International Collaboration Grant from the Jacki and Bruce Barron Cancer Research Scholars’ Program, a partnership of the Israel Cancer Research Fund and City of Hope, as supported by The Harvey L. Miller Family Foundation [grant number 205467].


  1. 1.
    Hogg M, Paro S, Keegan LP, O’Connell MA (2011) RNA editing by mammalian ADARs. Adv Genet 73:87–120PubMedGoogle Scholar
  2. 2.
    Nishikura K (2016) A-to-I editing of coding and non-coding RNAs by ADARs. Nat Rev Mol Cell Biol 17:83–96Google Scholar
  3. 3.
    Savva YA, Rieder LE, Reenan RA (2012) The ADAR protein family. Genome Biol 13:252PubMedPubMedCentralGoogle Scholar
  4. 4.
    Eisenberg E, Levanon EY (2018) A-to-I RNA editing—immune protector and transcriptome diversifier. Nat Rev Genet 19:473–490Google Scholar
  5. 5.
    Lonsdale J et al (2013) The genotype-tissue expression (GTEx) project. Nat Genet 45:580–585Google Scholar
  6. 6.
    Tan MH et al (2017) Dynamic landscape and regulation of RNA editing in mammals. Nature 550:249–254PubMedPubMedCentralGoogle Scholar
  7. 7.
    Jain M et al (2018) RNA editing of Filamin A pre-mRNA regulates vascular contraction and diastolic blood pressure. EMBO J 37:e94813. Scholar
  8. 8.
    Cho DSC et al (2003) Requirement of dimerization for RNA editing activity of adenosine deaminases acting on RNA. J Biol Chem 278:17093–17102PubMedGoogle Scholar
  9. 9.
    Tomaselli S, Locatelli F, Gallo A (2014) The RNA editing enzymes ADARs: mechanism of action and human disease. Cell Tissue Res 356:527–532PubMedGoogle Scholar
  10. 10.
    Chen CX et al (2000) A third member of the RNA-specific adenosine deaminase gene family, ADAR3, contains both single- and double-stranded RNA binding domains. RNA 6:755–767PubMedPubMedCentralGoogle Scholar
  11. 11.
    Nishikura K (2010) Functions and regulation of RNA editing by ADAR deaminases. Annu Rev Biochem 79:321–349PubMedPubMedCentralGoogle Scholar
  12. 12.
    Palavicini JP, O’Connell MA, Rosenthal JJC (2009) An extra double-stranded RNA binding domain confers high activity to a squid RNA editing enzyme. RNA 15:1208–1218PubMedPubMedCentralGoogle Scholar
  13. 13.
    Keegan LP et al (2011) Functional conservation in human and Drosophila of Metazoan ADAR2 involved in RNA editing: loss of ADAR1 in insects. Nucleic Acids Res 39:7249–7262PubMedPubMedCentralGoogle Scholar
  14. 14.
    Lehmann KA, Bass BL (1999) The importance of internal loops within RNA substrates of ADAR1. J Mol Biol 291:1–13PubMedGoogle Scholar
  15. 15.
    Ramaswami G et al (2012) Accurate identification of human Alu and non-Alu RNA editing sites. Nat Methods 9:579–581PubMedPubMedCentralGoogle Scholar
  16. 16.
    Neeman Y, Dahary D, Levanon E, Sorek R, Eisenberg E (2005) Is there any sense in antisense editing?. Trends in Genetics 21 (10):544–547Google Scholar
  17. 17.
    Picardi E, Pesole G (2013) REDItools: high-throughput RNA editing detection made easy. Bioinformatics 29:1813–1814PubMedGoogle Scholar
  18. 18.
    Morse DP, Aruscavage PJ, Bass BL (2002) RNA hairpins in noncoding regions of human brain and Caenorhabditis elegans mRNA are edited by adenosine deaminases that act on RNA. Proc Natl Acad Sci U S A 99:7906–7911PubMedPubMedCentralGoogle Scholar
  19. 19.
    Levanon EY et al (2004) Systematic identification of abundant A-to-I editing sites in the human transcriptome. Nat Biotechnol 22:1001–1005PubMedGoogle Scholar
  20. 20.
    Porath HT, Carmi S, Levanon EY (2014) A genome-wide map of hyper-edited RNA reveals numerous new sites. Nat Commun 5:1–10Google Scholar
  21. 21.
    Liddicoat BJ et al (2015) RNA editing by ADAR1 prevents MDA5 sensing of endogenous dsRNA as nonself. Science 349:1–9Google Scholar
  22. 22.
    Pestal K et al (2015) Isoforms of RNA-editing enzyme ADAR1 independently control nucleic acid sensor MDA5-driven autoimmunity and multi-organ development. Immunity 43:933–944PubMedPubMedCentralGoogle Scholar
  23. 23.
    Mannion NM et al (2014) The RNA-editing enzyme ADAR1 controls innate immune responses to RNA. Cell Rep 9:1482–1494PubMedPubMedCentralGoogle Scholar
  24. 24.
    Samuel CE (2001) Antiviral actions of interferons. Clin Microbiol Rev 14:778–809, table of contentsPubMedPubMedCentralGoogle Scholar
  25. 25.
    Patterson JB, Samuel CE (1995) Expression and regulation by interferon of a double-stranded-RNA-specific adenosine deaminase from human cells: evidence for two forms of the deaminase. Mol Cell Biol 15:5376–5388PubMedPubMedCentralGoogle Scholar
  26. 26.
    Paz-Yaacov N et al (2015) Elevated RNA editing activity is a major contributor to transcriptomic diversity in tumors. Cell Rep 13:267–276PubMedGoogle Scholar
  27. 27.
    Fumagalli D et al (2015) Principles governing A-to-I RNA editing in the breast cancer transcriptome article principles governing A-to-I RNA editing in the breast cancer transcriptome. Cell Rep 13:277–289PubMedPubMedCentralGoogle Scholar
  28. 28.
    Han L et al (2015) The genomic landscape and clinical relevance of A-to-I RNA editing in human cancers. Cancer Cell 28:515–528PubMedPubMedCentralGoogle Scholar
  29. 29.
    Galeano F, Tomaselli S, Locatelli F, Gallo A (2012) A-to-I RNA editing: the ‘ADAR’ side of human cancer. Semin Cell Dev Biol 23:244–250PubMedGoogle Scholar
  30. 30.
    Porath HT et al (2019) RNA editing is abundant and correlates with task performance in a social bumblebee. Nat Commun 10:1605PubMedPubMedCentralGoogle Scholar
  31. 31.
    Li Q et al (2014) Caste-specific RNA editomes in the leaf-cutting ant Acromyrmex echinatior. Nat Commun 5:4943PubMedPubMedCentralGoogle Scholar
  32. 32.
    Buchumenski I et al (2017) Dynamic hyper-editing underlies temperature adaptation in Drosophila. PLoS Genet 13:e1006931PubMedPubMedCentralGoogle Scholar
  33. 33.
    Rieder LE et al (2015) Dynamic response of RNA editing to temperature in Drosophila. BMC Biol 13:1PubMedPubMedCentralGoogle Scholar
  34. 34.
    Robinson JE, Paluch J, Dickman DK, Joiner WJ (2016) ADAR-mediated RNA editing suppresses sleep by acting as a brake on glutamatergic synaptic plasticity. Nat Commun 7:10512PubMedPubMedCentralGoogle Scholar
  35. 35.
    Basilio C, Wahba AJ, Lengyel P, Speyer JF, Ochoa S (1962) Synthetic polynucleotides and the amino acid code. Proc Natl Acad Sci U S A 48:613–616PubMedPubMedCentralGoogle Scholar
  36. 36.
    Bass BL (2002) RNA editing by adenosine deaminases that act on RNA. Annu Rev Biochem 71:817–846PubMedGoogle Scholar
  37. 37.
    Athanasiadis A, Rich A, Maas S (2004) Widespread A-to-I RNA editing of Alu-containing mRNAs in the human transcriptome. PLoS Biol 2:e391PubMedPubMedCentralGoogle Scholar
  38. 38.
    Li JB et al (2009) Genome-wide identification of human RNA editing sites by parallel DNA capturing and sequencing. Science 324:1210–1213PubMedGoogle Scholar
  39. 39.
    Lee J-H, Ang JK, Xiao X (2013) Analysis and design of RNA sequencing experiments for identifying RNA editing and other single-nucleotide variants. RNA 19:725–732PubMedPubMedCentralGoogle Scholar
  40. 40.
    Eisenberg E, Li JB, Levanon EY (2010) Sequence based identification of RNA editing sites. RNA Biol 7:248–252PubMedGoogle Scholar
  41. 41.
    Schrider DR, Gout J-F, Hahn MW (2011) Very few RNA and DNA sequence differences in the human transcriptome. PLoS One 6:e25842PubMedPubMedCentralGoogle Scholar
  42. 42.
    Diroma MA, Ciaccia L, Pesole G, Picardi E (2017) Elucidating the editome: bioinformatics approaches for RNA editing detection. Brief Bioinform 20(2):436–447. Scholar
  43. 43.
    Reich DP, Bass BL (2019) Mapping the dsRNA world. Cold Spring Harb Perspect Biol 11:a035352PubMedPubMedCentralGoogle Scholar
  44. 44.
    Carmi S, Borukhov I, Levanon EY (2011) Identification of widespread ultra-edited human RNAs. PLoS Genet 7:e1002317PubMedPubMedCentralGoogle Scholar
  45. 45.
    Li H, Durbin R (2009) Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics 25:1754–1760PubMedPubMedCentralGoogle Scholar
  46. 46.
    Zaranek AW, Levanon EY, Zecharia T, Clegg T, Church GM (2010) A survey of genomic traces reveals a common sequencing error, RNA editing, and DNA editing. PLoS Genet 6:8Google Scholar
  47. 47.
    Dobin A et al (2013) STAR: ultrafast universal RNA-seq aligner. Bioinformatics 29:15–21PubMedPubMedCentralGoogle Scholar
  48. 48.
    Kim D, Langmead B, Salzberg SL (2015) HISAT: a fast spliced aligner with low memory requirements. Nat Methods 12:357–360PubMedPubMedCentralGoogle Scholar
  49. 49.
    Eggington JM, Greene T, Bass BL (2011) Predicting sites of ADAR editing in double-stranded RNA. Nat Commun 2:319PubMedPubMedCentralGoogle Scholar
  50. 50.
    Porath HT, Knisbacher BA, Eisenberg E, Levanon EY (2017) Massive A-to-I RNA editing is common across the Metazoa and correlates with dsRNA abundance. Genome Biol 18:185PubMedPubMedCentralGoogle Scholar
  51. 51.
    Wu TD, Nacu S (2010) Fast and SNP-tolerant detection of complex variants and splicing in short reads. Bioinformatics 26:873–881PubMedPubMedCentralGoogle Scholar
  52. 52.
    Baruzzo G et al (2017) Simulation-based comprehensive benchmarking of RNA-seq aligners. Nat Methods 14:135–139PubMedGoogle Scholar
  53. 53.
    Williams AG, Thomas S, Wyman SK, Holloway AK (2014) RNA-seq data: challenges in and recommendations for experimental design and analysis. Curr Protoc Hum Genet 83:11.13.1–11.13.20Google Scholar
  54. 54.
    Bass BL (1997) RNA editing and hypermutation by adenosine deamination. Trends Biochem Sci 22:157–162PubMedGoogle Scholar
  55. 55.
    Sommer B, Kohler M, Sprengel R, Seeburg PH (1991) RNA editing in brain controls a determinant of ion flow in glutamate-gated channels. Cell 67:11–19PubMedGoogle Scholar
  56. 56.
    Pinto Y, Cohen HY, Levanon EY (2014) Mammalian conserved ADAR targets comprise only a small fragment of the human editosome. Genome Biol 15:R5PubMedPubMedCentralGoogle Scholar
  57. 57.
    Ramaswami G, Li JB (2014) RADAR: a rigorously annotated database of A-to-I RNA editing. Nucleic Acids Res 42:D109–D113PubMedGoogle Scholar
  58. 58.
    Hartner JC, Walkley CR, Lu J, Orkin SH (2009) ADAR1 is essential for the maintenance of hematopoiesis and suppression of interferon signaling. Nat Immunol 10:109–115PubMedGoogle Scholar
  59. 59.
    Liddicoat BJ, Chalk AM, Walkley CR (2015) ADAR1, inosine and the immune sensing system: distinguishing self from non-self. Wiley Interdiscip Rev RNA 7:157–172PubMedGoogle Scholar
  60. 60.
    Porath HT et al (2017) A-to-I RNA editing in the earliest-diverging eumetazoan phyla. Mol Biol Evol 34:1890–1901PubMedPubMedCentralGoogle Scholar
  61. 61.
    Liscovitch-Brauer N et al (2017) Trade-off between transcriptome plasticity and genome evolution in cephalopods. Cell 169:191–202.e11PubMedPubMedCentralGoogle Scholar

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© Springer Science+Business Media, LLC, part of Springer Nature 2021

Authors and Affiliations

  1. 1.Mina and Everard Goodman Faculty of Life SciencesBar-Ilan UniversityRamat-GanIsrael

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