Circular RNA Splicing

  • Nicole Eger
  • Laura Schoppe
  • Susanne Schuster
  • Ulrich Laufs
  • Jes-Niels BoeckelEmail author
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 1087)


Circular RNAs (circRNAs) are covalently closed single-stranded RNA molecules derived from exons by alternative mRNA splicing. Circularization of single-stranded RNA molecules was already described in 1976 for viroids in plants. Since then several additional types of circular RNAs in many species have been described such as the circular single-stranded RNA genome of the hepatitis delta virus (HDV) or circular RNAs as products or intermediates of tRNA and rRNA maturation in archaea. CircRNAs are generally formed by covalent binding of the 5′ site of an upstream exon with the 3′ of the same or a downstream exon. Meanwhile, two different models of circRNA biogenesis have been described, the lariat or exon skipping model and the direct backsplicing model. In the lariat model, canonical splicing occurs before backsplicing, whereas in the direct backsplicing model, the circRNA is generated first. In this chapter, we will review the formation of circular RNAs and highlight the derivation of different types of circular RNAs.


circRNA RNA splicing Circular RNA Backsplicing 



This work was supported by the German Cardiac Society (Deutsche Gesellschaft für Kardiologie (DGK)) to Jes-Niels Boeckel.

Competing Financial Interests

The authors declare no competing financial interests.


  1. 1.
    Hsu MT, Coca-Prados M (1979) Electron microscopic evidence for the circular form of RNA in the cytoplasm of eukaryotic cells. Nature 280:339–340CrossRefGoogle Scholar
  2. 2.
    Nigro JM, Cho KR, Fearon ER et al (1991) Scrambled exons. Cell 64:607–613CrossRefGoogle Scholar
  3. 3.
    Zaphiropoulos PG (1997) Exon skipping and circular RNA formation in transcripts of the human cytochrome P-450 2C18 gene in epidermis and of the rat androgen binding protein gene in testis. Mol Cell Biol 17:2985–2993CrossRefGoogle Scholar
  4. 4.
    Cocquerelle C, Mascrez B, Hétuin D et al (1993) Mis-splicing yields circular RNA molecules. FASEB J 7:155–160CrossRefGoogle Scholar
  5. 5.
    Cocquerelle C, Daubersies P, Majérus MA et al (1992) Splicing with inverted order of exons occurs proximal to large introns. EMBO J 11:1095–1098CrossRefGoogle Scholar
  6. 6.
    Burd CE, Jeck WR, Liu Y et al (2010) Expression of linear and novel circular forms of an INK4/ARF-associated non-coding RNA correlates with atherosclerosis risk. PLoS Genet 6:e1001233CrossRefGoogle Scholar
  7. 7.
    Aebi M, Weissman C (1987) Precision and orderliness in splicing. Trends Genet 3:102–107CrossRefGoogle Scholar
  8. 8.
    Zhang Y, Zhang XO, Chen T et al (2013) Circular intronic long noncoding RNAs. Mol Cell 51:792–806CrossRefGoogle Scholar
  9. 9.
    Dubin RA, Kazmi MA, Ostrer H (1995) Inverted repeats are necessary for circularization of the mouse testis Sry transcript. Gene 167:245–248CrossRefGoogle Scholar
  10. 10.
    Sanger HL, Klotz G, Riesner D et al (1976) Viroids are single-stranded covalently closed circular RNA molecules existing as highly base-paired rod-like structures. Proc Natl Acad Sci U S A 73:3852–3856CrossRefGoogle Scholar
  11. 11.
    Schultz ES, Folsom D (1923) A “spindling-tuber disease” of Irish potatoes. Science 57:149. CrossRefPubMedGoogle Scholar
  12. 12.
    Diener TO (1971) Potato spindle tuber “virus”. IV. A replicating, low molecular weight RNA. Virology 45:411–428CrossRefGoogle Scholar
  13. 13.
    Stoler MH, Broker TR (1986) In situ hybridization detection of human papillomavirus DNAs and messenger RNAs in genital condylomas and a cervical carcinoma. Hum Pathol 17:1250–1258CrossRefGoogle Scholar
  14. 14.
    Summers J, Jones SE, Anderson MJ (1983) Characterization of the genome of the agent of erythrocyte aplasia permits its classification as a human parvovirus. J Gen Virol 64(Pt 11):2527–2532CrossRefGoogle Scholar
  15. 15.
    Kos A, Dijkema R, Arnberg AC et al (1986) The hepatitis delta (delta) virus possesses a circular RNA. Nature 323:558–560CrossRefGoogle Scholar
  16. 16.
    Soma A, Onodera A, Sugahara J et al (2007) Permuted tRNA genes expressed via a circular RNA intermediate in Cyanidioschyzon merolae. Science 318:450–453CrossRefGoogle Scholar
  17. 17.
    Lykke-Andersen J, Aagaard C, Semionenkov M et al (1997) Archaeal introns: splicing, intercellular mobility and evolution. Trends Biochem Sci 22:326–331CrossRefGoogle Scholar
  18. 18.
    Salgia SR, Singh SK, Gurha P et al (2003) Two reactions of Haloferax volcanii RNA splicing enzymes: joining of exons and circularization of introns. RNA 9:319–330CrossRefGoogle Scholar
  19. 19.
    Danan M, Schwartz S, Edelheit S et al (2012) Transcriptome-wide discovery of circular RNAs in Archaea. Nucleic Acids Res 40:3131–3142CrossRefGoogle Scholar
  20. 20.
    Tang TH, Rozhdestvensky TS, d’Orval BC et al (2002) RNomics in Archaea reveals a further link between splicing of archaeal introns and rRNA processing. Nucleic Acids Res 30:921–930CrossRefGoogle Scholar
  21. 21.
    Jeck WR, Sorrentino JA, Wang K et al (2013) Circular RNAs are abundant, conserved, and associated with ALU repeats. RNA 19:141–157CrossRefGoogle Scholar
  22. 22.
    Barrett SP, Wang PL, Salzman J (2015) Circular RNA biogenesis can proceed through an exon-containing lariat precursor. Elife 4:e07540CrossRefGoogle Scholar
  23. 23.
    Diener TO, Raymer WB (1967) Potato spindle tuber virus: a plant virus with properties of a free nucleic acid. Science 158:378–381CrossRefGoogle Scholar
  24. 24.
    Flores R, Hernández C, Martínez de Alba AE et al (2005) Viroids and viroid-host interactions. Annu Rev Phytopathol 43:117–139CrossRefGoogle Scholar
  25. 25.
    Bonfiglioli, McFadden, Symons RH (1994) In situ hybridization localizes avocado sunblotch viroid on chloroplast thylakoid membranes and coconut cadang cadang viroid in the nucleus. Plant J 6(1):99–103CrossRefGoogle Scholar
  26. 26.
    Przybilski R, Gräf S, Lescoute A et al (2005) Functional hammerhead ribozymes naturally encoded in the genome of Arabidopsis thaliana. Plant Cell 17:1877–1885CrossRefGoogle Scholar
  27. 27.
    Steger G, Perreault JP (2016) Structure and associated biological functions of viroids. Adv Virus Res 94:141–172CrossRefGoogle Scholar
  28. 28.
    Flores R, Grubb D, Elleuch A et al (2011) Rolling-circle replication of viroids, viroid-like satellite RNAs and hepatitis delta virus: variations on a theme. RNA Biol 8(2):200–206CrossRefGoogle Scholar
  29. 29.
    Flores R, Gas ME, Molina-Serrano D et al (2009) Viroid replication: rolling-circles, enzymes and ribozymes. Viruses 1:317–334CrossRefGoogle Scholar
  30. 30.
    Branch AD, Robertson HD (1984) A replication cycle for viroids and other small infectious RNA’s. Science 223:450–455CrossRefGoogle Scholar
  31. 31.
    Flores (1999) Viroids with hammerhead ribozymes: some unique structural and functional aspects with respect to other members of the group. Biol Chem 380(7-8):849–854CrossRefGoogle Scholar
  32. 32.
    Macnaughton TB, Shi ST, Modahl LE et al (2002) Rolling circle replication of hepatitis delta virus RNA is carried out by two different cellular RNA polymerases. J Virol 76:3920–3927CrossRefGoogle Scholar
  33. 33.
    Reid CE, Lazinski DW (2000) A host-specific function is required for ligation of a wide variety of ribozyme-processed RNAs. Proc Natl Acad Sci U S A 97:424–429CrossRefGoogle Scholar
  34. 34.
    Petkovic S, Müller S (2015) RNA circularization strategies in vivo and in vitro. Nucleic Acids Res 43:2454–2465CrossRefGoogle Scholar
  35. 35.
    Barral PM, Sarkar D, Su ZZ et al (2009) Functions of the cytoplasmic RNA sensors RIG-I and MDA-5: key regulators of innate immunity. Pharmacol Ther 124:219–234CrossRefGoogle Scholar
  36. 36.
    Yoneyama M, Onomoto K, Jogi M et al (2015) Viral RNA detection by RIG-I-like receptors. Curr Opin Immunol 32:48–53CrossRefGoogle Scholar
  37. 37.
    Abelson J, Trotta CR, Li H (1998) tRNA splicing. J Biol Chem 273:12685–12688CrossRefGoogle Scholar
  38. 38.
    Knapp G, Ogden RC, Peebles CL et al (1979) Splicing of yeast tRNA precursors: structure of the reaction intermediates. Cell 18:37–45CrossRefGoogle Scholar
  39. 39.
    Lu Z, Filonov GS, Noto JJ, Schmidt CA, Hatkevich TL, Wen Y, Jaffrey SR, Matera AG (2015) Metazoan tRNA introns generate stable circular RNAs in vivo. RNA 21:1554–1565CrossRefGoogle Scholar
  40. 40.
    Suzuki H, Zuo Y, Wang J et al (2006) Characterization of RNase R-digested cellular RNA source that consists of lariat and circular RNAs from pre-mRNA splicing. Nucleic Acids Res 34:e63CrossRefGoogle Scholar
  41. 41.
    Salzman J, Gawad C, Wang PL, Lacayo N, Brown PO (2012) Circular RNAs are the predominant transcript isoform from hundreds of human genes in diverse cell types. PLoS One 7:e30733CrossRefGoogle Scholar
  42. 42.
    Lasda E, Parker R, Parker ROY (2014) Circular RNAs : diversity of form and function. RNA 20:1829–1842CrossRefGoogle Scholar
  43. 43.
    Grabowski PJ, Zaug AJ, Cech TR (1981) The intervening sequence of the ribosomal RNA precursor is converted to a circular RNA in isolated nuclei of Tetrahymena. Cell 23:467–476CrossRefGoogle Scholar
  44. 44.
    Durovic P, Dennis PP (1994) Separate pathways for excision and processing of 16S and 23S rRNA from the primary rRNA operon transcript from the hyperthermophilic archaebacterium Sulfolobus acidocaldarius: similarities to eukaryotic rRNA processing. Mol Microbiol 13:229–242CrossRefGoogle Scholar
  45. 45.
    Dennis PP, Ziesche S, Mylvaganam S (1998) Transcription analysis of two disparate rRNA operons in the halophilic archaeon Haloarcula marismortui. J Bacteriol 180:4804–4813PubMedPubMedCentralGoogle Scholar
  46. 46.
    Kruger K, Grabowski PJ, Zaug AJ et al (1982) Self-splicing RNA: autoexcision and autocyclization of the ribosomal RNA intervening sequence of Tetrahymena. Cell 31:147–157CrossRefGoogle Scholar
  47. 47.
    Grabowski PJ, Seiler SR, Sharp PA (1985) A multicomponent complex is involved in the splicing of messenger RNA precursors. Cell 42:345–353CrossRefGoogle Scholar
  48. 48.
    Qian L, Vu MN, Carter M, Wilkinson MF (1992) A spliced intron accumulates as a lariat in the nucleus of T cells. Nucleic Acids Res 20:5345–5350CrossRefGoogle Scholar
  49. 49.
    Tabak HF, Van der Horst G, Smit J et al (1988) Discrimination between RNA circles, interlocked RNA circles and lariats using two-dimensional polyacrylamide gel electrophoresis. Nucleic Acids Res 16:6597–6605CrossRefGoogle Scholar
  50. 50.
    Hedberg A, Johansen SD (2013) Nuclear group I introns in self-splicing and beyond. Mob DNA 4:1CrossRefGoogle Scholar
  51. 51.
    Lambowitz AM, Zimmerly S (2011) Group II introns: mobile ribozymes that invade DNA. Cold Spring Harb Perspect Biol 3:1–19CrossRefGoogle Scholar
  52. 52.
    Lehmann K, Schmidt U (2003) Group II introns: structure and catalytic versatility of large natural ribozymes. Crit Rev Biochem Mol Biol 38:249–303CrossRefGoogle Scholar
  53. 53.
    Li-Pook-Than J, Bonen L (2006) Multiple physical forms of excised group II intron RNAs in wheat mitochondria. Nucleic Acids Res 34:2782–2790CrossRefGoogle Scholar
  54. 54.
    Soesanto W, Lin HY, Hu E et al (2009) Mammalian target of rapamycin is a critical regulator of cardiac hypertrophy in spontaneously hypertensive rats. Hypertension 54:1321–1327CrossRefGoogle Scholar
  55. 55.
    Capel B, Swain A, Nicolis S et al (1993) Circular transcripts of the testis-determining gene Sry in adult mouse testis. Cell 73:1019–1030CrossRefGoogle Scholar
  56. 56.
    Pasman Z, Been MD, Garcia-Blanco MA (1996) Exon circularization in mammalian nuclear extracts. RNA 2:603–610PubMedPubMedCentralGoogle Scholar
  57. 57.
    Caldas C, So CW, MacGregor A et al (1998) Exon scrambling of MLL transcripts occur commonly and mimic partial genomic duplication of the gene. Gene 208:167–176CrossRefGoogle Scholar
  58. 58.
    Zaphiropoulos PG (1996) Circular RNAs from transcripts of the rat cytochrome P450 2C24 gene: correlation with exon skipping. Proc Natl Acad Sci U S A 93:6536–6541CrossRefGoogle Scholar
  59. 59.
    Baralle FE, Giudice J (2017) Alternative splicing as a regulator of development and tissue identity. Nat Rev Mol Cell Biol 18:437–451CrossRefGoogle Scholar
  60. 60.
    Salzman J, Chen RE, Olsen MN et al (2013) Cell-type specific features of circular RNA expression. PLoS Genet 9:e1003777CrossRefGoogle Scholar
  61. 61.
    Zhang XO, Dong R, Zhang Y et al (2016) Diverse alternative back-splicing and alternative splicing landscape of circular RNAs. Genome Res 26:1277–1287CrossRefGoogle Scholar
  62. 62.
    Zhang XO, Wang HB, Zhang Y et al (2014) Complementary sequence-mediated exon circularization. Cell 159:134–147CrossRefGoogle Scholar
  63. 63.
    Chen LL (2016) The biogenesis and emerging roles of circular RNAs. Nat Rev Mol Cell Biol 17:205–211CrossRefGoogle Scholar
  64. 64.
    1000 Genomes Project Consortium, Auton A, Brooks LD et al (2015) A global reference for human genetic variation. Nature 526:68–74CrossRefGoogle Scholar
  65. 65.
    Lynch M, Conery JS (2003) The origins of genome complexity. Science 302:1401–1404CrossRefGoogle Scholar
  66. 66.
    Khanduja JS, Calvo IA, Joh RI, Hill IT, Motamedi M (2016) Nuclear noncoding RNAs and genome stability. Mol Cell 63:7–20CrossRefGoogle Scholar
  67. 67.
    Böhmdorfer G, Wierzbicki AT (2015) Control of chromatin structure by long noncoding RNA. Trends Cell Biol 25:623–663CrossRefGoogle Scholar
  68. 68.
    Magistri M, Faghihi MA, St Laurent G, Wahlestedt C (2012) Regulation of chromatin structure by long noncoding RNAs: focus on natural antisense transcripts. Trends Genet 28:389–396CrossRefGoogle Scholar
  69. 69.
    Vance KW, Ponting CP (2014) Transcriptional regulatory functions of nuclear long noncoding RNAs. Trends Genet 30:348–355CrossRefGoogle Scholar
  70. 70.
    Wilusz JE, Sunwoo H, Spector DL (2009) Long noncoding RNAs: functional surprises from the RNA world. Genes Dev 23:1494–1504CrossRefGoogle Scholar
  71. 71.
    Bonasio R, Shiekhattar R (2014) Regulation of transcription by long noncoding RNAs. Annu Rev Genet 48:433–455CrossRefGoogle Scholar
  72. 72.
    Lee TI, Young RA (2000) Transcription of eukaryotic protein-coding genes. Annu Rev Genet 34:77–137CrossRefGoogle Scholar
  73. 73.
    Nogales E, Louder RK, He Y (2017) Structural insights into the eukaryotic transcription initiation machinery. Annu Rev Biophys 46:59–83CrossRefGoogle Scholar
  74. 74.
    Jurado AR, Tan D, Jiao X et al (2014) Structure and function of pre-mRNA 5′-end capping quality control and 3′-end processing. Biochemistry 53:1882–1898CrossRefGoogle Scholar
  75. 75.
    Moore MJ, Proudfoot NJ (2009) Pre-mRNA processing reaches back to transcription and ahead to translation. Cell 136:688–700CrossRefGoogle Scholar
  76. 76.
    Furuichi Y, Shatkin AJ (2000) Viral and cellular mRNA capping: past and prospects. Adv Virus Res 55:135–184CrossRefGoogle Scholar
  77. 77.
    Shandilya J, Roberts SGE (2012) The transcription cycle in eukaryotes: from productive initiation to RNA polymerase II recycling. Biochim Biophys Acta 1819:391–400CrossRefGoogle Scholar
  78. 78.
    Wahle E, Rüegsegger U (1999) 3′-End processing of pre-mRNA in eukaryotes. FEMS Microbiol Rev 23:277–295CrossRefGoogle Scholar
  79. 79.
    Frendewey D, Keller W (1985) Stepwise assembly of a pre-mRNA splicing complex requires U-snRNPs and specific intron sequences. Cell 42:355–367CrossRefGoogle Scholar
  80. 80.
    Burset M, Seledtsov IA, Solovyev VV (2000) Analysis of canonical and non-canonical splice sites in mammalian genomes. Nucleic Acids Res 28:4364–4375CrossRefGoogle Scholar
  81. 81.
    Hwang DY, Cohen JB (1996) Base pairing at the 5′ splice site with U1 small nuclear RNA promotes splicing of the upstream intron but may be dispensable for slicing of the downstream intron. Mol Cell Biol 16:3012–3022CrossRefGoogle Scholar
  82. 82.
    Liu J, Liu T, Wang X, He A (2017) Circles reshaping the RNA world: from waste to treasure. Mol Cancer 16:58CrossRefGoogle Scholar
  83. 83.
    Huang S, Yang B, Chen BJ et al (2017) The emerging role of circular RNAs in transcriptome regulation. Genomics 109:401–407CrossRefGoogle Scholar
  84. 84.
    Starke S, Jost I, Rossbach O, Schneider T et al (2015) Exon circularization requires canonical splice signals. Cell Rep 10:103–111CrossRefGoogle Scholar
  85. 85.
    Gilbert W (1978) Why genes in pieces? Nature 271:501CrossRefGoogle Scholar
  86. 86.
    Valdivia HH (2007) One gene, many proteins: alternative splicing of the ryanodine receptor gene adds novel functions to an already complex channel protein. Circ Res 100:761–763CrossRefGoogle Scholar
  87. 87.
    George CH, Rogers SA, Bertrand BMA et al (2007) Alternative splicing of ryanodine receptors modulates cardiomyocyte Ca2+ signaling and susceptibility to apoptosis. Circ Res 100:874–883CrossRefGoogle Scholar
  88. 88.
    Lanner JT, Georgiou DK, Joshi AD, Hamilton SL (2010) Ryanodine receptors: structure, expression, molecular details, and function in calcium release. Cold Spring Harb Perspect Biol 2:a003996CrossRefGoogle Scholar
  89. 89.
    Mabon SA, Misteli T (2005) Differential recruitment of pre-mRNA splicing factors to alternatively spliced transcripts in vivo. PLoS Biol 3:1893–1901CrossRefGoogle Scholar
  90. 90.
    Boeckel J-N, Jaé N, Heumüller AW et al (2015) Identification and characterization of hypoxia-regulated endothelial circular RNA. Circ Res 117(10):884–890CrossRefGoogle Scholar
  91. 91.
    Konarska MM, Padgett RA, Sharp PA (1985) Trans splicing of mRNA precursors in vitro. Cell 42:165–171CrossRefGoogle Scholar
  92. 92.
    Liang D, Wilusz JE (2014) Short intronic repeat sequences facilitate circular RNA production. Genes Dev 28:2233–2247CrossRefGoogle Scholar
  93. 93.
    Lev-Maor G, Ram O, Kim E et al (2008) Intronic Alus influence alternative splicing. PLoS Genet 4:1–12CrossRefGoogle Scholar
  94. 94.
    Hu S, Wang X, Shan G (2016) Insertion of an Alu element in a lncRNA leads to primate-specific modulation of alternative splicing. Nat Struct Mol Biol 23:1011–1019CrossRefGoogle Scholar
  95. 95.
    Barrett SP, Salzman J (2016) Circular RNAs: analysis, expression and potential functions. Development 143:1838–1847CrossRefGoogle Scholar
  96. 96.
    Braun S, Domdey H, Wiebauer K (1996) Inverse splicing of a discontinuous pre-mRNA intron generates a circular exon in a HeLa cell nuclear extract. Nucleic Acids Res 24:4152–4157CrossRefGoogle Scholar
  97. 97.
    Chen LL, Yang L (2015) Regulation of circRNA biogenesis. RNA Biol 12:381–388CrossRefGoogle Scholar
  98. 98.
    Schindewolf C, Braun S, Domdey H (1996) In vitro generation of a circular exon from a linear pre-mRNA transcript. Nucleic Acids Res 24:1260–1266CrossRefGoogle Scholar
  99. 99.
    Hesselberth JR (2013) Lives that introns lead after splicing. Wiley Interdiscip Rev RNA 4:677–691CrossRefGoogle Scholar

Copyright information

© Springer Nature Singapore Pte Ltd. 2018

Authors and Affiliations

  • Nicole Eger
    • 1
  • Laura Schoppe
    • 1
  • Susanne Schuster
    • 2
  • Ulrich Laufs
    • 2
  • Jes-Niels Boeckel
    • 2
    Email author
  1. 1.University of HeidelbergHeidelbergGermany
  2. 2.Clinic and Polyclinic for CardiologyUniversity Hospital LeipzigLeipzigGermany

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