The 5′-cap structures of eukaryotic mRNAs are important for RNA stability, pre-mRNA splicing, mRNA export, and protein translation. Many viruses have evolved mechanisms for generating their own cap structures with methylation at the N7 position of the capped guanine and the ribose 2′-Oposition of the first nucleotide, which help viral RNAs escape recognition by the host innate immune system. The RNA genomes of coronavirus were identified to have 5′-caps in the early 1980s. However, for decades the RNA capping mechanisms of coronaviruses remained unknown. Since 2003, the outbreak of severe acute respiratory syndrome coronavirus has drawn increased attention and stimulated numerous studies on the molecular virology of coronaviruses. Here, we review the current understanding of the mechanisms adopted by coronaviruses to produce the 5′-cap structure and methylation modification of viral genomic RNAs.
Abbas YM, Pichlmair A, Gorna MW, Superti-Furga G, Nagar B. 2013. Structural basis for viral 5′-PPP-RNA recognition by human IFIT proteins. Nature, 494: 60–64.
Adams MJ, Carstens EB. 2012. Ratification vote on taxonomic proposals to the International Committee on Taxonomy of Viruses. Arch Virol, 157: 1411–1422.
Ahola T, Ahlquist P. 1999. Putative RNA capping activities encoded by brome mosaic virus: methylation and covalent binding of guanylate by replicase protein 1a. J Virol, 73: 10061–10069.
Ahola T, Kaariainen L. 1995. Reaction in alphavirus mRNA capping: formation of a covalent complex of nonstructural protein nsP1 with 7-methyl-GMP. Proc Natl Acad Sci USA, 92: 507–511.
Banerjee AK. 1980. 5′-terminal cap structure in eucaryotic messenger ribonucleic acids. Microbiol Rev, 44: 175–205.
Bhardwaj K, Guarino L, Kao CC. 2004. The severe acute respiratory syndrome coronavirus Nsp15 protein is an endoribonuclease that prefers manganese as a cofactor. J Virol, 78: 12218–12224.
Bouvet M, Debarnot C, Imbert I, Selisko B, Snijder EJ, Canard B, Decroly E. 2010. In vitro reconstitution of SARS-coronavirus mRNA cap methylation. PLoS Pathog, 6: e1000863.
Bouvet M, Imbert I, Subissi L, Gluais L, Canard B, Decroly E. 2012. RNA 3′-end mismatch excision by the severe acute respiratory syndrome coronavirus nonstructural protein nsp10/nsp14 exoribonuclease complex. Proc Natl Acad Sci USA, 109: 9372–9377.
Bowzard JB, Ranjan P, Sambhara S. 2013. RIG-I goes beyond naked recognition. Cell Host Microbe, 13: 247–249.
Brierley I, Digard P, Inglis SC. 1989. Characterization of an efficient coronavirus ribosomal frameshifting signal: requirement for an RNA pseudoknot. Cell, 57: 537–547.
Cauchemez S, van Kerkhove MD, Riley S, Donnelly CA, Fraser C, Ferguson NM. 2013. Transmission scenarios for Middle East Respiratory Syndrome Coronavirus (MERS-CoV) and how to tell them apart. Euro Surveill, 18: 20503.
Chen P, Jiang M, Hu T, Liu Q, Chen XS, Guo D. 2007. Biochemical characterization of exoribonuclease encoded by SARS coronavirus. J Biochem Mol Biol, 40: 649–655.
Chen Y, Cai H, Pan J, Xiang N, Tien P, Ahola T, Guo D. 2009. Functional screen reveals SARS coronavirus nonstructural protein nsp14 as a novel cap N7 methyltransferase. Proc Natl Acad Sci USA, 106: 3484–3489.
Chen Y, Su C, Ke M, Jin X, Xu L, Zhang Z, Wu A, Sun Y, Yang Z, Tien P, Ahola T, Liang Y, Liu X, Guo D. 2011. Biochemical and structural insights into the mechanisms of SARS coronavirus RNA ribose 2′-O-methylation by nsp16/nsp10 protein complex. PLoS Pathog, 7: e1002294.
Chen Y, Tao J, Sun Y, Wu A, Su C, Gao G, Cai H, Qiu S, Wu Y, Ahola T, Guo D. 2013. Structure-function analysis of severe acute respiratory syndrome coronavirus RNA cap guanine-N7- methyltransferase. J Virol, 87: 6296–6305.
Chrebet GL, Wisniewski D, Perkins AL, Deng Q, Kurtz MB, Marcy A, Parent SA. 2005. Cell-based assays to detect inhibitors of fungal mRNA capping enzymes and characterization of sinefungin as a cap methyltransferase inhibitor. J Biomol Screen, 10: 355–364.
Cowling VH. 2010. Regulation of mRNA cap methylation. Biochem J, 425: 295–302.
Daffis S, Szretter KJ, Schriewer J, Li J, Youn S, Errett J, Lin TY, Schneller S, Zust R, Dong H, Thiel V, Sen GC, Fensterl V, Klimstra WB, Pierson TC, Buller RM, Gale M, Jr., Shi PY, Diamond MS. 2010. 2′-O methylation of the viral mRNA cap evades host restriction by IFIT family members. Nature, 468: 452–456.
Darnell JE, Jr. 1979. Transcription units for mRNA production in eukaryotic cells and their DNA viruses. Prog Nucleic Acid Res Mol Biol, 22: 327–353.
De la Pena M, Kyrieleis OJ, Cusack S. 2007. Structural insights into the mechanism and evolution of the vaccinia virus mRNA cap N7 methyl-transferase. EMBO J, 26: 4913–4925.
Decroly E, Debarnot C, Ferron F, Bouvet M, Coutard B, Imbert I, Gluais L, Papageorgiou N, Sharff A, Bricogne G, Ortiz-Lombardia M, Lescar J, Canard B. 2011. Crystal Structure and Functional Analysis of the SARS-Coronavirus RNA Cap 2′-OMethyltransferase nsp10/nsp16 Complex. PLoS Pathog, 7: e1002059.
Decroly E, Ferron F, Lescar J, Canard B. 2012. Conventional and unconventional mechanisms for capping viral mRNA. Nat Rev Microbiol, 10: 51–65.
Decroly E, Imbert I, Coutard B, Bouvet M, Selisko B, Alvarez K, Gorbalenya AE, Snijder EJ, Canard B. 2008. Coronavirus nonstructural protein 16 is a cap-0 binding enzyme possessing (nucleoside- 2′O)-methyltransferase activity. J Virol, 82: 8071–8084.
Eckerle LD, Becker MM, Halpin RA, Li K, Venter E, Lu X, Scherbakova S, Graham RL, Baric RS, Stockwell TB, Spiro DJ, Denison MR. 2010. Infidelity of SARS-CoV Nsp14-exonuclease mutant virus replication is revealed by complete genome sequencing. PLoS Pathog, 6: e1000896.
Eckerle LD, Lu X, Sperry SM, Choi L, Denison MR. 2007. High fidelity of murine hepatitis virus replication is decreased in nsp14 exoribonuclease mutants. J Virol, 81: 12135–12144.
Enserink M. 2003. Infectious diseases. Clues to the animal origins of SARS. Science, 300: 1351.
Ferron F, Decroly E, Selisko B, Canard B. 2012. The viral RNA capping machinery as a target for antiviral drugs. Antiviral Res, 96: 21–31.
Furuichi Y. 2015. Discovery of m(7)G-cap in eukaryotic mRNAs. Proc Jpn Acad Ser B Phys Biol Sci, 91: 394–409.
Furuichi Y, Shatkin AJ. 2000. Viral and cellular mRNA capping: past and prospects. Adv Virus Res, 55: 135–184.
Ge XY, Li JL, Yang XL, Chmura AA, Zhu G, Epstein JH, Mazet JK, Hu B, Zhang W, Peng C, Zhang YJ, Luo CM, Tan B, Wang N, Zhu Y, Crameri G, Zhang SY, Wang LF, Daszak P, Shi ZL. 2013. Isolation and characterization of a bat SARS-like coronavirus that uses the ACE2 receptor. Nature, 503: 535–538.
Gonzalez JM, Gomez-Puertas P, Cavanagh D, Gorbalenya AE, Enjuanes L. 2003. A comparative sequence analysis to revise the current taxonomy of the family Coronaviridae. Arch Virol, 148: 2207–2235.
Gorbalenya AE, Enjuanes L, Ziebuhr J, Snijder EJ. 2006. Nidovirales: evolving the largest RNA virus genome. Virus Res, 117: 17–37.
Ho CK, Martins A, Shuman S. 2000. A yeast-based genetic system for functional analysis of viral mRNA capping enzymes. J Virol, 74: 5486–5494.
Hornung V, Ellegast J, Kim S, Brzozka K, Jung A, Kato H, Poeck H, Akira S, Conzelmann KK, Schlee M, Endres S, Hartmann G. 2006. 5′-Triphosphate RNA is the ligand for RIG-I. Science, 314: 994–997.
Hussain S, Pan J, Chen Y, Yang Y, Xu J, Peng Y, Wu Y, Li Z, Zhu Y, Tien P, Guo D. 2005. Identification of novel subgenomic RNAs and noncanonical transcription initiation signals of severe acute respiratory syndrome coronavirus. J Virol, 79: 5288–5295.
Imbert I, Guillemot JC, Bourhis JM, Bussetta C, Coutard B, Egloff MP, Ferron F, Gorbalenya AE, Canard B. 2006. A second, noncanonical RNA-dependent RNA polymerase in SARS coronavirus. Embo J, 25: 4933–4942.
Ivanov KA, Thiel V, Dobbe JC, van der Meer Y, Snijder EJ, Ziebuhr J. 2004. Multiple enzymatic activities associated with severe acute respiratory syndrome coronavirus helicase. J Virol, 78: 5619–5632.
Ivanov KA, Ziebuhr J. 2004. Human coronavirus 229E nonstructural protein 13: characterization of duplex-unwinding, nucleoside triphosphatase, and RNA 5′-triphosphatase activities. J Virol, 78: 7833–7838.
Jin H, Elliott RM. 1993. Characterization of Bunyamwera virus S RNA that is transcribed and replicated by the L protein expressed from recombinant vaccinia virus. J Virol, 67: 1396–1404.
Jin X, Chen Y, Sun Y, Zeng C, Wang Y, Tao J, Wu A, Yu X, Zhang Z, Tian J, Guo D. 2013. Characterization of the guanine- N7 methyltransferase activity of coronavirus nsp14 on nucleotide GTP. Virus Res, 176: 45–52.
Joseph JS, Saikatendu KS, Subramanian V, Neuman BW, Buchmeier MJ, Stevens RC, Kuhn P. 2007. Crystal structure of a monomeric form of severe acute respiratory syndrome coronavirus endonuclease nsp15 suggests a role for hexamerization as an allosteric switch. J Virol, 81: 6700–6708.
Ke M, Chen Y, Wu A, Sun Y, Su C, Wu H, Jin X, Tao J, Wang Y, Ma X, Pan JA, Guo D. 2012. Short peptides derived from the interaction domain of SARS coronavirus nonstructural protein nsp10 can suppress the 2′-O-methyltransferase activity of nsp10/nsp16 complex. Virus Res, 167: 322–328.
Lai MM, Patton CD, Stohlman SA. 1982. Further characterization of mRNA′s of mouse hepatitis virus: presence of common 5′- end nucleotides. J Virol, 41: 557–565.
Lai MM, Stohlman SA. 1981. Comparative analysis of RNA genomes of mouse hepatitis viruses. J Virol, 38: 661–670.
Leahy MB, Dessens JT, Nuttall PA. 1997. In vitro polymerase activity of Thogoto virus: evidence for a unique cap-snatching mechanism in a tick-borne orthomyxovirus. J Virol, 71: 8347–8351.
Li W, Shi Z, Yu M, Ren W, Smith C, Epstein JH, Wang H, Crameri G, Hu Z, Zhang H, Zhang J, McEachern J, Field H, Daszak P, Eaton BT, Zhang S, Wang LF. 2005. Bats are natural reservoirs of SARS-like coronaviruses. Science, 310: 676–679.
Liu H, Kiledjian M. 2006. Decapping the message: a beginning or an end. Biochem Soc Trans, 34: 35–38.
Ma Y, Wu L, Shaw N, Gao Y, Wang J, Sun Y, Lou Z, Yan L, Zhang R, Rao Z. 2015. Structural basis and functional analysis of the SARS coronavirus nsp14-nsp10 complex. Proc Natl Acad Sci USA, 112: 9436–9441.
Mao X, Shuman S. 1994. Intrinsic RNA (guanine-7) methyltransferase activity of the vaccinia virus capping enzyme D1 subunit is stimulated by the D12 subunit. Identification of amino acid residues in the D1 protein required for subunit association and methyl group transfer. J Biol Chem, 269: 24472–24479.
Martina BE, Haagmans BL, Kuiken T, Fouchier RA, Rimmelzwaan GF, van Amerongen G, Peiris JS, Lim W, Osterhaus AD. 2003. Virology: SARS virus infection of cats and ferrets. Nature, 425: 915.
Martinez-Salas E, Pineiro D, Fernandez N. 2012. Alternative Mechanisms to Initiate Translation in Eukaryotic mRNAs. Comp Funct Genomics, 2012: 391546.
Minskaia E, Hertzig T, Gorbalenya AE, Campanacci V, Cambillau C, Canard B, Ziebuhr J. 2006. Discovery of an RNA virus 3′→5′ exoribonuclease that is critically involved in coronavirus RNA synthesis. Proc Natl Acad Sci U S A, 103: 5108–5113.
Mizutani T. 2013. A novel coronavirus, MERS-CoV. Uirusu, 63: 1–6. (In Japanese)
Nallagatla SR, Toroney R, Bevilacqua PC. 2008. A brilliant disguise for self RNA: 5′-end and internal modifications of primary transcripts suppress elements of innate immunity. RNA Biol, 5: 140–144.
Ogino T, Banerjee AK. 2007. Unconventional mechanism of mRNA capping by the RNA-dependent RNA polymerase of vesicular stomatitis virus. Mol Cell, 25: 85–97.
Pan J, Peng X, Gao Y, Li Z, Lu X, Chen Y, Ishaq M, Liu D, Dediego ML, Enjuanes L, Guo D. 2008. Genome-wide analysis of protein-protein interactions and involvement of viral proteins in SARS-CoV replication. PLoS One, 3: e3299.
Pelletier J, Sonenberg N. 1988. Internal initiation of translation of eukaryotic mRNA directed by a sequence derived from poliovirus RNA. Nature, 334: 320–325.
Pichlmair A, Schulz O, Tan CP, Naslund TI, Liljestrom P, Weber F, Reis e Sousa C. 2006. RIG-I-mediated antiviral responses to single-stranded RNA bearing 5′-phosphates. Science, 314: 997–1001.
Prentice E, McAuliffe J, Lu X, Subbarao K, Denison MR. 2004. Identification and characterization of severe acute respiratory syndrome coronavirus replicase proteins. J Virol, 78: 9977–9986.
Ray D, Shah A, Tilgner M, Guo Y, Zhao Y, Dong H, Deas TS, Zhou Y, Li H, Shi PY. 2006. West Nile virus 5′-cap structure is formed by sequential guanine N-7 and ribose 2′-O methylations by nonstructural protein 5. J Virol, 80: 8362–8370.
Rehwinkel J, Tan CP, Goubau D, Schulz O, Pichlmair A, Bier K, Robb N, Vreede F, Barclay W, Fodor E, Reis e Sousa C. 2010. RIG-I detects viral genomic RNA during negative-strand RNA virus infection. Cell, 140: 397–408.
Saha N, Schwer B, Shuman S. 1999. Characterization of human, Schizosaccharomyces pombe, and Candida albicans mRNA cap methyltransferases and complete replacement of the yeast capping apparatus by mammalian enzymes. J Biol Chem, 274: 16553–16562.
Saha N, Shuman S, Schwer B. 2003. Yeast-based genetic system for functional analysis of poxvirus mRNA cap methyltransferase. J Virol, 77: 7300–7307.
Sawicki SG, Sawicki DL, Younker D, Meyer Y, Thiel V, Stokes H, Siddell SG. 2005. Functional and genetic analysis of coronavirus replicase-transcriptase proteins. PLoS Pathog, 1: e39.
Schwer B, Lehman K, Saha N, Shuman S. 2001. Characterization of the mRNA capping apparatus of Candida albicans. J Biol Chem, 276: 1857–1864.
Schwer B, Mao X, Shuman S. 1998. Accelerated mRNA decay in conditional mutants of yeast mRNA capping enzyme. Nucleic Acids Res, 26: 2050–2057.
Sevajol M, Subissi L, Decroly E, Canard B, Imbert I. 2014. Insights into RNA synthesis, capping, and proofreading mechanisms of SARS-coronavirus. Virus Res, 194: 90–99.
Shatkin AJ. 1976. Capping of eucaryotic mRNAs. Cell, 9: 645–653.
Shuman S. 2001. Structure, mechanism, and evolution of the mRNA capping apparatus. Prog Nucleic Acid Res Mol Biol, 66: 1–40.
Smith EC, Blanc H, Surdel MC, Vignuzzi M, Denison MR. 2013. Coronaviruses lacking exoribonuclease activity are susceptible to lethal mutagenesis: evidence for proofreading and potential therapeutics. PLoS Pathog, 9: e1003565.
Smith EC, Case JB, Blanc H, Isakov O, Shomron N, Vignuzzi M, Denison MR. 2015. Mutations in coronavirus nonstructural protein 10 decrease virus replication fidelity. J Virol, 89: 6418–6426.
Snijder EJ, Bredenbeek PJ, Dobbe JC, Thiel V, Ziebuhr J, Poon LL, Guan Y, Rozanov M, Spaan WJ, Gorbalenya AE. 2003. Unique and conserved features of genome and proteome of SARS-coronavirus, an early split-off from the coronavirus group 2 lineage. J Mol Biol, 331: 991–1004.
Sun Y, Wang Z, Tao J, Wang Y, Wu A, Yang Z, Wang K, Shi L, Chen Y, Guo D. 2014. Yeast-based assays for the high-throughput screening of inhibitors of coronavirus RNA cap guanine- N7-methyltransferase. Antiviral Res, 104: 156–164.
Tanner JA, Watt RM, Chai YB, Lu LY, Lin MC, Peiris JS, Poon LL, Kung HF, Huang JD. 2003. The severe acute respiratory syndrome (SARS) coronavirus NTPase/helicase belongs to a distinct class of 5′ to 3′ viral helicases. J Biol Chem, 278: 39578–39582.
te Velthuis AJ, van den Worm SH, Snijder EJ. 2012. The SARScoronavirus nsp7+nsp8 complex is a unique multimeric RNA polymerase capable of both de novo initiation and primer extension. Nucleic Acids Res, 40: 1737–1747.
Thiel V, Ivanov KA, Putics A, Hertzig T, Schelle B, Bayer S, Weissbrich B, Snijder EJ, Rabenau H, Doerr HW, Gorbalenya AE, Ziebuhr J. 2003. Mechanisms and enzymes involved in SARS coronavirus genome expression. J Gen Virol, 84: 2305–2315.
Tsukiyama-Kohara K, Iizuka N, Kohara M, Nomoto A. 1992. Internal ribosome entry site within hepatitis C virus RNA. J Virol, 66: 1476–1483.
van Vliet AL, Smits SL, Rottier PJ, de Groot RJ. 2002. Discontinuous and non-discontinuous subgenomic RNA transcription in a nidovirus. Embo J, 21: 6571–6580.
von Grotthuss M, Wyrwicz LS, Rychlewski L. 2003. mRNA cap-1 methyltransferase in the SARS genome. Cell, 113: 701–702.
Wang LF, Shi Z, Zhang S, Field H, Daszak P, Eaton BT. 2006. Review of bats and SARS. Emerg Infect Dis, 12: 1834–1840.
Wang Y, Sun Y, Wu A, Xu S, Pan R, Zeng C, Jin X, Ge X, Shi Z, Ahola T, Chen Y, Guo D. 2015. Coronavirus nsp10/nsp16 Methyltransferase Can Be Targeted by nsp10-Derived Peptide In Vitro and In Vivo To Reduce Replication and Pathogenesis. J Virol, 89: 8416–8427.
Woyciniuk P, Linder M, Scholtissek C. 1995. The methyltransferase inhibitor Neplanocin A interferes with influenza virus replication by a mechanism different from that of 3-deazaadenosine. Virus Res, 35: 91–99.
Ziebuhr J. 2004. Molecular biology of severe acute respiratory syndrome coronavirus. Curr Opin Microbiol, 7: 412–419.
Zuniga S, Sola I, Alonso S, Enjuanes L. 2004. Sequence motifs involved in the regulation of discontinuous coronavirus subgenomic RNA synthesis. J Virol, 78: 980–994.
Zust R, Cervantes-Barragan L, Habjan M, Maier R, Neuman BW, Ziebuhr J, Szretter KJ, Baker SC, Barchet W, Diamond MS, Siddell SG, Ludewig B, Thiel V. 2011. Ribose 2′-O-methylation provides a molecular signature for the distinction of self and non-self mRNA dependent on the RNA sensor Mda5. Nat Immunol, 12: 137–143.
About this article
Cite this article
Chen, Y., Guo, D. Molecular mechanisms of coronavirus RNA capping and methylation. Virol. Sin. 31, 3–11 (2016). https://doi.org/10.1007/s12250-016-3726-4
- RNA capping
- cap structure