Abstract
Many protein-coding genes in higher eukaryotes can produce circular RNAs (circRNAs) through back-splicing of exons. CircRNAs differ from mRNAs in their production, structure and turnover and thereby have unique cellular functions and potential biomedical applications. In this Review, I discuss recent progress in our understanding of the biogenesis of circRNAs and the regulation of their abundance and of their biological functions, including in transcription and splicing, sequestering or scaffolding of macromolecules to interfere with microRNA activities or signalling pathways, and serving as templates for translation. I further discuss the emerging roles of circRNAs in regulating immune responses and cell proliferation, and the possibilities of applying circRNA technologies in biomedical research.
Similar content being viewed by others
References
Black, D. L. Mechanisms of alternative pre-messenger RNA splicing. Annu. Rev. Biochem. 72, 291–336 (2003).
Nilsen, T. W. & Graveley, B. R. Expansion of the eukaryotic proteome by alternative splicing. Nature 463, 457–463 (2010).
Zhang, Y. et al. Circular intronic long noncoding RNAs. Mol. Cell 51, 792–806 (2013).
Lasda, E. & Parker, R. Circular RNAs: diversity of form and function. RNA 20, 1829–1842 (2014).
Li, X., Yang, L. & Chen, L. L. The biogenesis, functions, and challenges of circular RNAs. Mol. Cell 71, 428–442 (2018).
Sanger, H. L., Klotz, G., Riesner, D., Gross, H. J. & Kleinschmidt, A. K. Viroids are single-stranded covalently closed circular RNA molecules existing as highly base-paired rod-like structures. Proc. Natl Acad. Sci. USA 73, 3852–3856 (1976).
Capel, B. et al. Circular transcripts of the testis-determining gene Sry in adult mouse testis. Cell 73, 1019–1030 (1993).
Cocquerelle, C., Daubersies, P., Majerus, M. A., Kerckaert, J. P. & Bailleul, B. Splicing with inverted order of exons occurs proximal to large introns. EMBO J. 11, 1095–1098 (1992).
Cocquerelle, C., Mascrez, B., Hetuin, D. & Bailleul, B. Mis-splicing yields circular RNA molecules. FASEB J. 7, 155–160 (1993).
Nigro, J. M. et al. Scrambled exons. Cell 64, 607–613 (1991).
Pasman, Z., Been, M. D. & Garcia-Blanco, M. A. Exon circularization in mammalian nuclear extracts. RNA 2, 603–610 (1996).
Yang, L., Duff, M. O., Graveley, B. R., Carmichael, G. G. & Chen, L. L. Genomewide characterization of non-polyadenylated RNAs. Genome Biol. 12, R16 (2011).
Salzman, J., Gawad, C., Wang, P. L., Lacayo, N. & Brown, P. O. Circular RNAs are the predominant transcript isoform from hundreds of human genes in diverse cell types. PLoS One 7, e30733 (2012). This study suggests that circRNA production can be a general feature of gene expression in human cells.
Jeck, W. R. et al. Circular RNAs are abundant, conserved, and associated with ALU repeats. RNA 19, 141–157 (2013). This study uncovers circRNAs as a large class of RNA molecules in human cells.
Ivanov, A. et al. Analysis of intron sequences reveals hallmarks of circular RNA biogenesis in animals. Cell Rep. 10, 170–177 (2015).
Shen, Y., Guo, X. & Wang, W. Identification and characterization of circular RNAs in zebrafish. FEBS Lett. 591, 213–220 (2017).
Westholm, J. O. et al. Genome-wide analysis of Drosophila circular RNAs reveals their structural and sequence properties and age-dependent neural accumulation. Cell Rep. 9, 1966–1980 (2014).
Guo, J. U., Agarwal, V., Guo, H. & Bartel, D. P. Expanded identification and characterization of mammalian circular RNAs. Genome Biol. 15, 409 (2014).
Memczak, S. et al. Circular RNAs are a large class of animal RNAs with regulatory potency. Nature 495, 333–338 (2013). This study reports that circRNAs are a large class of RNA molecules with functional potential.
Fan, X. et al. Single-cell RNA-seq transcriptome analysis of linear and circular RNAs in mouse preimplantation embryos. Genome Biol. 16, 148 (2015).
Dong, R., Ma, X. K., Chen, L. L. & Yang, L. Increased complexity of circRNA expression during species evolution. RNA Biol. 14, 1064–1074 (2017).
Zhang, X. O. et al. Complementary sequence-mediated exon circularization. Cell 159, 134–147 (2014). This study reports that exon circularization often requires flanking ICSs and also uncovers alternative circularization.
Veno, M. T. et al. Spatio-temporal regulation of circular RNA expression during porcine embryonic brain development. Genome Biol. 16, 245 (2015).
Barrett, S. P., Wang, P. L. & Salzman, J. Circular RNA biogenesis can proceed through an exon-containing lariat precursor. eLife 4, e07540 (2015).
Broadbent, K. M. et al. Strand-specific RNA sequencing in Plasmodium falciparum malaria identifies developmentally regulated long non-coding RNA and circular RNA. BMC Genomics 16, 454 (2015).
Lu, T. et al. Transcriptome-wide investigation of circular RNAs in rice. RNA 21, 2076–2087 (2015).
Wang, P. L. et al. Circular RNA is expressed across the eukaryotic tree of life. PLoS One 9, e90859 (2014).
Dong, R., Ma, X. K., Li, G. W. & Yang, L. CIRCpedia v2: an updated database for comprehensive circular RNA annotation and expression comparison. Genomics Proteomics Bioinformatics 16, 226–233 (2018).
Zheng, Y., Ji, P., Chen, S., Hou, L. & Zhao, F. Reconstruction of full-length circular RNAs enables isoform-level quantification. Genome Med. 11, 2 (2019).
Ji, P. et al. Expanded expression landscape and prioritization of circular RNAs in mammals. Cell Rep. 26, 3444–3460 (2019).
Rybak-Wolf, A. et al. Circular RNAs in the mammalian brain are highly abundant, conserved, and dynamically expressed. Mol. Cell 58, 870–885 (2015). This study reports enrichment of circRNA expression in brains and provides an atlas of circRNA expression in mammalian brains.
You, X. et al. Neural circular RNAs are derived from synaptic genes and regulated by development and plasticity. Nat. Neurosci. 18, 603–610 (2015). This study shows enrichment of circRNA expression in brains and suggests circRNAs have a potential to regulate synaptic function.
Preusser, C. et al. Selective release of circRNAs in platelet-derived extracellular vesicles. J. Extracell. Vesicles 7, 1424473 (2018).
Conn, S. J. et al. The RNA binding protein quaking regulates formation of circRNAs. Cell 160, 1125–1134 (2015).
Nicolet, B. P. et al. Circular RNA expression in human hematopoietic cells is widespread and cell-type specific. Nucleic Acids Res. 46, 8168–8180 (2018).
Salzman, J., Chen, R. E., Olsen, M. N., Wang, P. L. & Brown, P. O. Cell-type specific features of circular RNA expression. PLoS Genet. 9, e1003777 (2013).
Zhang, X. O. et al. Diverse alternative back-splicing and alternative splicing landscape of circular RNAs. Genome Res. 26, 1277–1287 (2016). This study defines the diversity of alternative back-splicing and alternative splicing in circRNAs.
Errichelli, L. et al. FUS affects circular RNA expression in murine embryonic stem cell-derived motor neurons. Nat. Commun. 8, 14741 (2017).
Xia, P. et al. A circular RNA protects dormant hematopoietic stem cells from DNA sensor cGAS-mediated exhaustion. Immunity 48, 688–701 (2018). This study generates a circRNA-knockout mouse model that can exhibit phenotypes related to hematopoietic stem cell homeostasis.
Li, Q. et al. CircACC1 regulates assembly and activation of AMPK complex under metabolic stress. Cell Metab. 30, 157–173 e157 (2019).
Moore, M. J. & Proudfoot, N. J. Pre-mRNA processing reaches back to transcription and ahead to translation. Cell 136, 688–700 (2009).
Vo, J. N. et al. The landscape of circular RNA in cancer. Cell 176, 869–881 e813 (2019).
Ashwal-Fluss, R. et al. circRNA biogenesis competes with pre-mRNA splicing. Mol. Cell 56, 55–66 (2014).
Starke, S. et al. Exon circularization requires canonical splice signals. Cell Rep. 10, 103–111 (2015).
Wang, Y. & Wang, Z. Efficient backsplicing produces translatable circular mRNAs. RNA 21, 172–179 (2015).
Li, X. et al. A unified mechanism for intron and exon definition and back-splicing. Nature 573, 375–380 (2019). This study provides cryo-electron microscopy structures of the yeast spliceosomal E complex and demonstrates back-splicing is catalysed by the spliceosome.
Zhang, Y. et al. The biogenesis of nascent circular RNAs. Cell Rep. 15, 611–624 (2016).
Liang, D. & Wilusz, J. E. Short intronic repeat sequences facilitate circular RNA production. Genes Dev. 28, 2233–2247 (2014).
Liang, D. et al. The output of protein-coding genes shifts to circular RNAs when the pre-mRNA processing machinery is limiting. Mol. Cell 68, 940–954 e943 (2017). This study suggests differential use of spliceosome components between back-splicing and canonical splicing.
Zaphiropoulos, P. G. Circular RNAs from transcripts of the rat cytochrome P450 2C24 gene: correlation with exon skipping. Proc. Natl Acad. Sci. USA 93, 6536–6541 (1996).
Kelly, S., Greenman, C., Cook, P. R. & Papantonis, A. Exon skipping is correlated with exon circularization. J. Mol. Biol. 427, 2414–2417 (2015).
Chen, L. L. & Yang, L. Regulation of circRNA biogenesis. RNA Biol. 12, 381–388 (2015).
Jeck, W. R. & Sharpless, N. E. Detecting and characterizing circular RNAs. Nat. Biotechnol. 32, 453–461 (2014).
Gao, Y. et al. Comprehensive identification of internal structure and alternative splicing events in circular RNAs. Nat. Commun. 7, 12060 (2016).
Ottesen, E. W., Luo, D., Seo, J., Singh, N. N. & Singh, R. N. Human survival motor neuron genes generate a vast repertoire of circular RNAs. Nucleic Acids Res. 47, 2884–2905 (2019).
Braunschweig, U., Gueroussov, S., Plocik, A. M., Graveley, B. R. & Blencowe, B. J. Dynamic integration of splicing within gene regulatory pathways. Cell 152, 1252–1269 (2013).
Fong, N. et al. Pre-mRNA splicing is facilitated by an optimal RNA polymerase II elongation rate. Genes Dev. 28, 2663–2676 (2014).
Dubin, R. A., Kazmi, M. A. & Ostrer, H. Inverted repeats are necessary for circularization of the mouse testis Sry transcript. Gene 167, 245–248 (1995).
Kramer, M. C. et al. Combinatorial control of Drosophila circular RNA expression by intronic repeats, hnRNPs, and SR proteins. Genes Dev. 29, 2168–2182 (2015).
Guarnerio, J. et al. Oncogenic role of fusion-circRNAs derived from cancer-associated chromosomal translocations. Cell 165, 289–302 (2016).
Wang, M., Hou, J., Muller-McNicoll, M., Chen, W. & Schuman, E. M. Long and repeat-rich intronic sequences favor circular RNA formation under conditions of reduced spliceosome activity. iScience 20, 237–247 (2019).
Khan, M. A. et al. RBM20 regulates circular RNA production from the titin gene. Circ. Res. 119, 996–1003 (2016).
Fei, T. et al. Genome-wide CRISPR screen identifies HNRNPL as a prostate cancer dependency regulating RNA splicing. Proc. Natl Acad. Sci. USA 114, E5207–E5215 (2017).
Patino, C., Haenni, A. L. & Urcuqui-Inchima, S. NF90 isoforms, a new family of cellular proteins involved in viral replication? Biochimie 108, 20–24 (2015).
Li, X. et al. Coordinated circRNA biogenesis and function with NF90/NF110 in viral infection. Mol. Cell 67, 214–227 (2017).
Aktas, T. et al. DHX9 suppresses RNA processing defects originating from the Alu invasion of the human genome. Nature 544, 115–119 (2017).
Braunschweig, U. et al. Widespread intron retention in mammals functionally tunes transcriptomes. Genome Res. 24, 1774–1786 (2014).
Li, Z. et al. Exon-intron circular RNAs regulate transcription in the nucleus. Nat. Struct. Mol. Biol. 22, 256–264 (2015).
Conn, V. M. et al. A circRNA from SEPALLATA3 regulates splicing of its cognate mRNA through R-loop formation. Nat. Plants 3, 17053 (2017).
Huang, C., Liang, D., Tatomer, D. C. & Wilusz, J. E. A length-dependent evolutionarily conserved pathway controls nuclear export of circular RNAs. Genes Dev. 32, 639–644 (2018). This study reveals that circRNA nuclear export occurs in a length-dependent manner.
Gatfield, D. et al. The DExH/D box protein HEL/UAP56 is essential for mRNA nuclear export in Drosophila. Curr. Biol. 11, 1716–1721 (2001).
Liu, C. X. et al. Structure and degradation of circular RNAs regulate PKR activation in innate immunity. Cell 177, 865–880 e821 (2019). This study shows that circRNAs can form unique structures and regulate PKR activity, and reports that circRNA misregulation is related to an autoimmune disease.
Zhou, C. et al. Genome-wide maps of m6A circRNAs identify widespread and cell-type-specific methylation patterns that are distinct from mRNAs. Cell Rep. 20, 2262–2276 (2017).
Roundtree, I. A. et al. YTHDC1 mediates nuclear export of N6-methyladenosine methylated mRNAs. Elife 6, e31311 (2017).
Enuka, Y. et al. Circular RNAs are long-lived and display only minimal early alterations in response to a growth factor. Nucleic Acids Res. 44, 1370–1383 (2016).
Hansen, T. B. et al. miRNA-dependent gene silencing involving Ago2-mediated cleavage of a circular antisense RNA. EMBO J. 30, 4414–4422 (2011).
Kleaveland, B., Shi, C. Y., Stefano, J. & Bartel, D. P. A network of noncoding regulatory RNAs acts in the mammalian brain. Cell 174, 350–362 (2018).
Hansen, T. B. et al. Natural RNA circles function as efficient microRNA sponges. Nature 495, 384–388 (2013). This study functionally characterizes naturally expressed circRNAs by their acting as miRNA sponges.
Piwecka, M. et al. Loss of a mammalian circular RNA locus causes miRNA deregulation and affects brain function. Science 357, eaam8526 (2017). This study generates a circRNA-knockout mouse model that can exhibit neuronal phenotypes.
Han, Y. et al. Structure of human RNase L reveals the basis for regulated RNA decay in the IFN response. Science 343, 1244–1248 (2014).
Park, O. H. et al. Endoribonucleolytic cleavage of m6A-containing RNAs by RNase P/MRP complex. Mol. Cell 74, 494–507 (2019).
Jarrous, N. Roles of RNase P and its subunits. Trends Genet. 33, 594–603 (2017).
Fischer, J. W., Busa, V. F., Shao, Y. & Leung, A. K. L. Structure-mediated RNA decay by UPF1 and G3BP1. Mol. Cell 78, 70–84 (2020).
Kim, Y. K. & Maquat, L. E. UPFront and center in RNA decay: UPF1 in nonsense-mediated mRNA decay and beyond. RNA 25, 407–422 (2019).
Dong, R. et al. CircRNA-derived pseudogenes. Cell Res. 26, 747–750 (2016).
Guarnerio, J. et al. Intragenic antagonistic roles of protein and circRNA in tumorigenesis. Cell Res. 29, 628–640 (2019).
Liu, Y. et al. Back-spliced RNA from retrotransposon binds to centromere and regulates centromeric chromatin loops in maize. PLoS Biol. 18, e3000582 (2020).
Salmena, L., Poliseno, L., Tay, Y., Kats, L. & Pandolfi, P. P. A ceRNA hypothesis: the Rosetta Stone of a hidden RNA language? Cell 146, 353–358 (2011).
Poliseno, L. et al. A coding-independent function of gene and pseudogene mRNAs regulates tumour biology. Nature 465, 1033–1038 (2010).
Bosson, A. D., Zamudio, J. R. & Sharp, P. A. Endogenous miRNA and target concentrations determine susceptibility to potential ceRNA competition. Mol. Cell 56, 347–359 (2014).
Denzler, R., Agarwal, V., Stefano, J., Bartel, D. P. & Stoffel, M. Assessing the ceRNA hypothesis with quantitative measurements of miRNA and target abundance. Mol. Cell 54, 766–776 (2014).
Huang, R. et al. Circular RNA HIPK2 regulates astrocyte activation via cooperation of autophagy and ER stress by targeting MIR124–2HG. Autophagy 13, 1722–1741 (2017).
Zheng, Q. et al. Circular RNA profiling reveals an abundant circHIPK3 that regulates cell growth by sponging multiple miRNAs. Nat. Commun. 7, 11215 (2016).
Stoll, L. et al. Circular RNAs as novel regulators of beta-cell functions in normal and disease conditions. Mol. Metab. 9, 69–83 (2018).
Kristensen, L. S., Okholm, T. L. H., Veno, M. T. & Kjems, J. Circular RNAs are abundantly expressed and upregulated during human epidermal stem cell differentiation. RNA Biol. 15, 280–291 (2018).
Yu, C. Y. et al. The circular RNA circBIRC6 participates in the molecular circuitry controlling human pluripotency. Nat. Commun. 8, 1149 (2017).
Li, Q. et al. Circular RNA MAT2B promotes glycolysis and malignancy of hepatocellular carcinoma through the miR-338-3p/PKM2 Axis under hypoxic stress. Hepatology 70, 1298–1316 (2019).
Hu, Z. Q. et al. Circular RNA sequencing identifies CircASAP1 as a key regulator in hepatocellular carcinoma metastasis. Hepatology https://doi.org/10.1002/hep.31068 (2019).
Li, Y. et al. Circular RNA is enriched and stable in exosomes: a promising biomarker for cancer diagnosis. Cell Res. 25, 981–984 (2015).
Du, W. W. et al. Foxo3 circular RNA retards cell cycle progression via forming ternary complexes with p21 and CDK2. Nucleic Acids Res. 44, 2846–2858 (2016).
Du, W. W. et al. Foxo3 circular RNA promotes cardiac senescence by modulating multiple factors associated with stress and senescence responses. Eur. Heart J. 38, 1402–1412 (2017).
Zeng, Y. et al. A circular RNA binds to and activates AKT phosphorylation and nuclear localization reducing apoptosis and enhancing cardiac repair. Theranostics 7, 3842–3855 (2017).
Huang, S. et al. Loss of super-enhancer-regulated circRNA Nfix induces cardiac regeneration after myocardial infarction in adult mice. Circulation 139, 2857–2876 (2019).
Burd, C. E. et al. Expression of linear and novel circular forms of an INK4/ARF-associated non-coding RNA correlates with atherosclerosis risk. PLoS Genet. 6, e1001233 (2010).
Holdt, L. M. et al. Circular non-coding RNA ANRIL modulates ribosomal RNA maturation and atherosclerosis in humans. Nat. Commun. 7, 12429 (2016).
Abdelmohsen, K. et al. Identification of HuR target circular RNAs uncovers suppression of PABPN1 translation by CircPABPN1. RNA Biol. 14, 361–369 (2017).
Grammatikakis, I., Abdelmohsen, K. & Gorospe, M. Posttranslational control of HuR function. Wiley Interdiscip. Rev. RNA 8, e1372 (2017).
Hein, M. Y. et al. A human interactome in three quantitative dimensions organized by stoichiometries and abundances. Cell 163, 712–723 (2015).
Armakola, M. et al. Inhibition of RNA lariat debranching enzyme suppresses TDP-43 toxicity in ALS disease models. Nat. Genet. 44, 1302–1309 (2012).
Zhang, S. Y. et al. Inborn errors of RNA lariat metabolism in humans with brainstem viral infection. Cell 172, 952–965 (2018).
Harashima, A., Guettouche, T. & Barber, G. N. Phosphorylation of the NFAR proteins by the dsRNA-dependent protein kinase PKR constitutes a novel mechanism of translational regulation and cellular defense. Genes Dev. 24, 2640–2653 (2010).
Isken, O. et al. Members of the NF90/NFAR protein group are involved in the life cycle of a positive-strand RNA virus. EMBO J. 22, 5655–5665 (2003).
Smola, M. J., Rice, G. M., Busan, S., Siegfried, N. A. & Weeks, K. M. Selective 2’-hydroxyl acylation analyzed by primer extension and mutational profiling (SHAPE-MaP) for direct, versatile and accurate RNA structure analysis. Nat. Protoc. 10, 1643–1669 (2015).
Moldovan, L. I. et al. High-throughput RNA sequencing from paired lesional- and non-lesional skin reveals major alterations in the psoriasis circRNAome. BMC Med. Genomics 12, 174 (2019).
Zhu, P. et al. IL-13 secreted by ILC2s promotes the self-renewal of intestinal stem cells through circular RNA circPan3. Nat. Immunol. 20, 183–194 (2019).
Chen, Y. G. et al. Sensing self and foreign circular RNAs by intron identity. Mol. Cell 67, 228–238 e225 (2017).
Wesselhoeft, R. A. et al. RNA circularization diminishes immunogenicity and can extend translation duration in vivo. Mol. Cell 74, 508–520 e504 (2019).
Chen, Y. G. et al. N6-methyladenosine modification controls circular RNA immunity. Mol. Cell 76, 96–109 (2019).
Toptan, T. et al. Circular DNA tumor viruses make circular RNAs. Proc. Natl Acad. Sci. USA 115, E8737–E8745 (2018).
Huang, J. T. et al. Identification of virus-encoded circular RNA. Virology 529, 144–151 (2019).
Tagawa, T. et al. Discovery of Kaposi’s sarcoma herpesvirus-encoded circular RNAs and a human antiviral circular RNA. Proc. Natl Acad. Sci. USA 115, 12805–12810 (2018).
Ungerleider, N. et al. The Epstein Barr virus circRNAome. PLoS Pathog. 14, e1007206 (2018).
Zhao, J. et al. Transforming activity of an oncoprotein-encoding circular RNA from human papillomavirus. Nat. Commun. 10, 2300 (2019).
Chen, S. et al. Widespread and functional RNA circularization in localized prostate cancer. Cell 176, 831–843 (2019).
Bachmayr-Heyda, A. et al. Correlation of circular RNA abundance with proliferation — exemplified with colorectal and ovarian cancer, idiopathic lung fibrosis, and normal human tissues. Sci. Rep. 5, 8057 (2015).
Panda, A. C. et al. Identification of senescence-associated circular RNAs (SAC-RNAs) reveals senescence suppressor CircPVT1. Nucleic Acids Res. 45, 4021–4035 (2017).
Yu, J. et al. Circular RNA cSMARCA5 inhibits growth and metastasis in hepatocellular carcinoma. J. Hepatol. 68, 1214–1227 (2018).
Yang, W., Du, W. W., Li, X., Yee, A. J. & Yang, B. B. Foxo3 activity promoted by non-coding effects of circular RNA and Foxo3 pseudogene in the inhibition of tumor growth and angiogenesis. Oncogene 35, 3919–3931 (2016).
Lukiw, W. J. Circular RNA (circRNA) in Alzheimer’s disease (AD). Front. Genet. 4, 307 (2013).
Dube, U. et al. An atlas of cortical circular RNA expression in Alzheimer disease brains demonstrates clinical and pathological associations. Nat. Neurosci. 22, 1903–1912 (2019).
Chen, Y. J. et al. Genome-wide, integrative analysis of circular RNA dysregulation and the corresponding circular RNA-microRNA-mRNA regulatory axes in autism. Genome Res. 30, 375–391 (2020).
Litke, J. L. & Jaffrey, S. R. Highly efficient expression of circular RNA aptamers in cells using autocatalytic transcripts. Nat. Biotechnol. 37, 667–675 (2019).
Jost, I. et al. Functional sequestration of microRNA-122 from hepatitis C virus by circular RNA sponges. RNA Biol. 15, 1032–1039 (2018).
Memczak, S., Papavasileiou, P., Peters, O. & Rajewsky, N. Identification and characterization of circular RNAs as a new class of putative biomarkers in human blood. PLoS One 10, e0141214 (2015).
Li, H. et al. Comprehensive circular RNA profiles in plasma reveals that circular RNAs can be used as novel biomarkers for systemic lupus erythematosus. Clin. Chim. Acta 480, 17–25 (2018).
Bahn, J. H. et al. The landscape of microRNA, Piwi-interacting RNA, and circular RNA in human saliva. Clin. Chem. 61, 221–230 (2015).
Rees, H. A. & Liu, D. R. Base editing: precision chemistry on the genome and transcriptome of living cells. Nat. Rev. Genet. 19, 770–788 (2018).
Abudayyeh, O. O. et al. RNA targeting with CRISPR-Cas13. Nature 550, 280–284 (2017).
Konermann, S. et al. Transcriptome engineering with RNA-targeting type VI-D CRISPR effectors. Cell 173, 665–676 e614 (2018).
Yang, L. Z. et al. Dynamic imaging of RNA in living cells by CRISPR-Cas13 systems. Mol. Cell 76, 981–997 e987 (2019).
Zhang, Y., Yang, L. & Chen, L. L. Characterization of circular RNAs. Methods Mol. Biol. 1402, 215–227 (2016).
Xiao, M. S. & Wilusz, J. E. An improved method for circular RNA purification using RNase R that efficiently removes linear RNAs containing G-quadruplexes or structured 3’ ends. Nucleic Acids Res. 47, 8755–8769 (2019).
Ma, X. K. et al. CIRCexplorer3: a CLEAR pipeline for direct comparison of circular and linear RNA expression. Genomics Proteomics Bioinformatics 17, 511–521 (2019).
Jakobi, T., Uvarovskii, A. & Dieterich, C. Circtools-a one-stop software solution for circular RNA research. Bioinformatics 35, 2326–2328 (2019).
Chuang, T. J. et al. Integrative transcriptome sequencing reveals extensive alternative trans-splicing and cis-backsplicing in human cells. Nucleic Acids Res. 46, 3671–3691 (2018).
Dahl, M. et al. Enzyme-free digital counting of endogenous circular RNA molecules in B-cell malignancies. Lab. Invest. 98, 1657–1669 (2018).
Li, T. et al. Plasma circular RNA profiling of patients with gastric cancer and their droplet digital RT-PCR detection. J. Mol. Med. 96, 85–96 (2018).
Legnini, I. et al. Circ-ZNF609 is a circular RNA that can be translated and functions in myogenesis. Mol Cell 66, 22–37 e29 (2017).
Pamudurti, N. R. et al. Translation of circRNAs. Mol. Cell 66, 9–21 (2017).
Yang, Y. et al. Extensive translation of circular RNAs driven by N6-methyladenosine. Cell Res. 27, 626–641 (2017).
Schneider, T. et al. CircRNA-protein complexes: IMP3 protein component defines subfamily of circRNPs. Sci. Rep. 6, 31313 (2016).
Wang, K. et al. A circular RNA protects the heart from pathological hypertrophy and heart failure by targeting miR-223. Eur. Heart J. 37, 2602–2611 (2016).
Chen, C. Y. & Sarnow, P. Initiation of protein synthesis by the eukaryotic translational apparatus on circular RNAs. Science 268, 415–417 (1995).
Wesselhoeft, R. A., Kowalski, P. S. & Anderson, D. G. Engineering circular RNA for potent and stable translation in eukaryotic cells. Nat. Commun. 9, 2629 (2018).
van Heesch, S. et al. The translational landscape of the human heart. Cell 178, 242–260 (2019).
Fan, X. et al. Pervasive translation of circular RNAs driven by short IRES-like elements. bioRxiv https://doi.org/10.1101/473207 (2019).
Tang, C. et al. m6A-dependent biogenesis of circular RNAs in male germ cells. Cell Res. 30, 211–228 (2020).
Mankan, A. K. et al. Cytosolic RNA:DNA hybrids activate the cGAS-STING axis. EMBO J. 33, 2937–2946 (2014).
Acknowledgements
The author apologizes to colleagues whose work is not discussed here owing to space limitations. The author thanks L. Yang, C.-X. Liu, X. Li and S.-K. Guo for discussions. This work was supported by grants from the Chinese Academy of Sciences (XDB19020104), the National Natural Science Foundation of China (91940303, 31725009, 31821004, 31861143025) and the HHMI International Research Scholar Program (55008728).
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
The author declares no competing interests.
Additional information
Peer review information
Nature Reviews Molecular Cell Biology thanks Albrecht Bindereif, Howard Chang and Jeremy Wilusz for their contribution to the peer review of this work.
Publisher’s note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Box 1
Glossary
- RNase R
-
A 3′-to-5′ exonuclease that preferentially digests linear RNAs, thereby allowing the enrichment of circular RNAs.
- Internal ribosome entry site
-
(IRES). A structural RNA element that makes possible the initiation of cap-independent translation.
- Spliceosomal E complex
-
Formation of this complex initiates the splicing cycle and is crucial for the accurate definition of introns and exons by the splicing machinery.
- Cassette exons
-
Exons present in one RNA transcript but absent in an isoform of the transcript.
- Alu elements
-
Primate-specific retrotransposons that constitute almost 11% of the human genome.
- Exon definition complexes
-
Protein complexes that initially recognize splice sites and direct prespliceosome assembly on exons. They further interact across long introns to form the catalytic spliceosome.
- A-to-I editing
-
In higher eukaryotes, the predominant form of RNA modification, in which adenosine is modified to inosine within imperfect double-stranded RNAs.
- Nonsense-mediated mRNA decay
-
A mechanism of selective degradation of mRNAs; a means of post-transcriptional gene regulation in mammals.
- R-loops
-
Triple-stranded nucleic acid structures that form during transcription; they consist of a DNA–RNA hybrid and the single-stranded non-template DNA.
- Group I introns
-
Large autocatalytic ribozymes that catalyse their own excision from mRNA, tRNA and ribosomal RNA precursors.
- Pattern recognition receptor
-
Cellular protein that recognizes pathogenic molecules and confers protection against viral infection by eliciting innate immunity responses.
Rights and permissions
About this article
Cite this article
Chen, LL. The expanding regulatory mechanisms and cellular functions of circular RNAs. Nat Rev Mol Cell Biol 21, 475–490 (2020). https://doi.org/10.1038/s41580-020-0243-y
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/s41580-020-0243-y
- Springer Nature Limited
This article is cited by
-
EIF4A3-mediated biogenesis of circSTX6 promotes bladder cancer metastasis and cisplatin resistance
Journal of Experimental & Clinical Cancer Research (2024)
-
Cancer therapy resistance mediated by cancer-associated fibroblast-derived extracellular vesicles: biological mechanisms to clinical significance and implications
Molecular Cancer (2024)
-
Circular RNAs and their roles in idiopathic pulmonary fibrosis
Respiratory Research (2024)
-
Exosomes secreted by Fusobacterium nucleatum-infected colon cancer cells transmit resistance to oxaliplatin and 5-FU by delivering hsa_circ_0004085
Journal of Nanobiotechnology (2024)
-
Regulatory mechanisms of PD-1/PD-L1 in cancers
Molecular Cancer (2024)