Skip to main content
PMC home page
Elsevier - PMC COVID-19 Collection logoLink to Elsevier - PMC COVID-19 Collection
. 2004 Feb 9;169(1):142–151. doi: 10.1016/0042-6822(89)90050-0

Sequence analysis of the nucleocapsid protein gene of human coronavirus 229E☆

Steven S Schreiber 1,1, Toshio Kamahora 1,2, Michael MC Lai 1
PMCID: PMC7131124  PMID: 2922924

Abstract

Human coronaviruses are important human pathogens and have also been implicated in multiple sclerosis. To further understand the molecular biology of human coronavirus 229E (HCV-229E), molecular cloning and sequence analysis of the viral RNA have been initiated. Following established protocols, the 3′-terminal 1732 nucleotides of the genome were sequenced. A large open reading frame encodes a 389 amino acid protein of 43,366 Da, which is presumably the nucleocapsid protein. The predicted protein is similar in size, chemical properties, and amino acid sequence to the nucleocapsid proteins of other coronaviruses. This is especially evident when the sequence is compared with that of the antigenically related porcine transmissible gastroenteritis virus (TGEV), with which a region of 46% amino acid sequence homology was found. Hydropathy profiles revealed the existence of several conserved domains which could have functional significance. An intergenic consensus sequence precedes the 5′-end of the proposed nucleocapsid protein gene. The consensus sequence is present in other coronaviruses and has been proposed as the site of binding of the leader sequence for mRNA transcriptional start. This region was also examined by primer extension analysis of mRNAs, which identified a 60-nucleotide leader sequence. The 3′-noncoding region of the genome contains an 11-nucleotide sequence, which is relatively conserved throughout the Coronavirus family and lends support to the theory that this region is important for the replication of negative-strand RNA.

Footnotes

☆

Sequence data from this article have been deposited with the EMBL/GenBank Data Libraries under Accession No. 104419.

References

  1. Armstrong J., Smeekens S., Rottier P. Sequence of the nucleocapsid gene from murine coronavirus MHV-A59. Nucleic Acids Res. 1983;11:883–891. doi: 10.1093/nar/11.3.883. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Baric R.S., Stohlman S.A., Razavi M.K., Lai M.M.C. Characterization of leader-related small RNAs in coronavirus-infected cells: Further evidence for leader-primed mechanism of transcription. Virus Res. 1985;3:19–33. doi: 10.1016/0168-1702(85)90038-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Boursnell M.E.G., Binns M.M., Foulds I.J., Brown T.D.K. Sequences of the nucleocapsid genes from two strains of avian infectious bronchitis virus. J. Gen. Virol. 1985;66:573–580. doi: 10.1099/0022-1317-66-3-573. [DOI] [PubMed] [Google Scholar]
  4. Boursnell M.E.G., Brown T.D.K., Foulds I.J., Green P.F., Tomley F.M., Binns M.M. Completion of the sequence of the genome of the coronavirus avian infectious bronchitis virus. J. Gen. Virol. 1987;68:57–77. doi: 10.1099/0022-1317-68-1-57. [DOI] [PubMed] [Google Scholar]
  5. Brayton P.R., Lai M.M.C., Patton C.D., Stohlman S.A. Characterization of two RNA polymerase activities induced by mouse hepatitis virus. J. Virol. 1982;42:847–853. doi: 10.1128/jvi.42.3.847-853.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Budzilowicz C.J., Wilczynski S.P., Weiss S.R. Three intergenic regions of coronavirus mouse hepatitis virus strain A59 genome RNA contain a common nucleotide sequence that is homologous to the Tend of the viral mRNA leader sequence. J. Virol. 1985;53:834–840. doi: 10.1128/jvi.53.3.834-840.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Burks J., Devald B.L., Jankovsky L.D., Gerdes J.C. Two coronaviruses isolated from central nervous system tissue of two multiple sclerosis patients. Science. 1980;209:933–934. doi: 10.1126/science.7403860. [DOI] [PubMed] [Google Scholar]
  8. Dagert M., Ehrlich S.D. Prolonged incubation in calcium chloride improves the competence of Escherichia coli cells. Gene. 1979;6:23–29. doi: 10.1016/0378-1119(79)90082-9. [DOI] [PubMed] [Google Scholar]
  9. Grunstein M., Hogness D.S. Vol. 72. 1975. Colony hybridization: A method for the isolation of cloned DNAs that contain a specific gene; pp. 3961–3965. (Proc. Natl. Acad. Sci. USA). [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gubler U., Hoffman B.J. Simple and very efficient method for generating cDNA libraries. Gene. 1983;25:263–269. doi: 10.1016/0378-1119(83)90230-5. [DOI] [PubMed] [Google Scholar]
  11. Hansen J.N. Use of solubilizable acrylamide disulfide gels for isolation of DNA fragments suitable for sequence analysis. Anal. Biochem. 1981;116:146–151. doi: 10.1016/0003-2697(81)90337-7. [DOI] [PubMed] [Google Scholar]
  12. Hansen J.N., Pheiffer B.H., Boehnert J.A. Chemical and electrophoretic properties of solubilizable disulfide gels. Anal. Biochem. 1980;105:192–201. doi: 10.1016/0003-2697(80)90445-5. [DOI] [PubMed] [Google Scholar]
  13. Hierholzer J.C. Purification and biophysical properties of human coronavirus 229E. Virology. 1976;75:155–165. doi: 10.1016/0042-6822(76)90014-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hierholzer J.C., Kemp M.C., Tannock G.A. The RNA and proteins of human coronaviruses. In: ter Meulen V., Siddell S., Wege H., editors. Vol. 142. Plenum; New York/London: 1981. pp. 43–69. (Biochemistry and Biology of Coronaviruses). [Google Scholar]
  15. Kamahora T., Soe L.H., Lai M.M.C. Sequence analysis of nucleocapsid gene and leader RNA of human coronavirus OC43. Virus Res. 1988 doi: 10.1016/0168-1702(89)90048-8. in press. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kapke P.A., Brian D.A. Sequence analysis of the porcine transmissable gastroenteritis coronavirus nucleocapsid protein gene. Virology. 1986;151:41–49. doi: 10.1016/0042-6822(86)90102-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Keck J.G., Hogue B.G., Brian D.A., Lai M.M.C. Temporal regulation of bovine coronavirus RNA synthesis. Virus Res. 1988;9:343–356. doi: 10.1016/0168-1702(88)90093-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kemp M.C., Hierholzer J.C., Harrison A., Burks J.S. Characterization of viral proteins synthesized in 229E-infected cells and effect(s) of inhibition of glycosylation and glycoprotein transport. In: Rottier P.J.M., van der Zeijst B.A.M., Spaan W.J.M., Horzinek M.C., editors. Vol. 173. Plenum; New York/London: 1984. pp. 65–79. (Molecular Biology and Pathogenesis of Coronaviruses). [DOI] [PubMed] [Google Scholar]
  19. Kennedy D.A., Johnson-Lussenberg C.M. Isolation and morphology of the internal component of human coronavirus, strain 229E. Intervirology. 1975/76;6:197–206. doi: 10.1159/000149474. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. King B., Potts B.J., Brian D.A. Bovine coronavirus hemagglutinin protein. Virus Res. 1985;2:53–59. doi: 10.1016/0168-1702(85)90059-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kozak M. Comparison of initiation of protein synthesis in procaryotes, eucaryotes, and organelles. Microbiol. Rev. 1983;47:1–45. doi: 10.1128/mr.47.1.1-45.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Lai M.M.C. Replication of coronavirus RNA. In: Holland J., Ahlquist P., Domingo E., editors. Vol. 1. CRC Press; Boca Raton, FL: 1988. pp. 116–136. (RNA Genetics). [Google Scholar]
  23. Lai M.M.C., Baric R.S., Brayton P.R., Stohlman S.A. Vol. 81. 1984. Characterization of leader RNA sequences on the virion and mRNAs of mouse hepatitis virus, a cytoplasmic virus; pp. 3626–3630. (Proc. Natl. Acad. Sci. USA). [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Lai M.M.C., Brayton P.R., Armen R.C., Patton C.D., Pugh C., Stohlman S.A. Mouse hepatitis virus A59: Messenger RNA structure and genetic localization of the sequence divergence from the hepatotropic strain MHV-3. J. Virol. 1981;39:823–834. doi: 10.1128/jvi.39.3.823-834.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lai M.M.C., Making S., Soe L.H., Shieh C.-K., Keck J.G., Fleming J.O. Coronavirus: A jumping RNA transcription. Cold Spring Harbor Symp. Quant. Biol. 1987;52:359–365. doi: 10.1101/sqb.1987.052.01.041. [DOI] [PubMed] [Google Scholar]
  26. Lai M.M.C., Patton C.D., Baric R.S., Stohlman S.A. Presence of leader sequences in the mRNA of mouse hepatitis virus. J. Virol. 1983;46:1027–1033. doi: 10.1128/jvi.46.3.1027-1033.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Lai M.M.C., Stohlman S.A. The RNA of mouse hepatitis virus. J. Virol. 1978;26:236–242. doi: 10.1128/jvi.26.2.236-242.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Lapps W., Hogue B.G., Brian D.A. Sequence analysis of the bovine coronavirus nucleocapsid and matrix protein genes. Virology. 1987;157:47–57. doi: 10.1016/0042-6822(87)90312-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. MacNaughton M.R. The polypeptides of human and mouse coronaviruses. Arch. Virol. 1980;63:75–80. doi: 10.1007/BF01320763. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. MacNaughton M.R. Structural and antigenic relationships between human, murine and avian coronaviruses. In: ter Meulen V., Siddell S., Wege H., editors. Vol. 142. Plenum; New York/London: 1981. pp. 19–29. (Biochemistry and Biology of Coronaviruses). [DOI] [PubMed] [Google Scholar]
  31. Makino S., Stohlman S.A., Lai M.M.C. Vol. 83. 1986. Leader sequences of murine coronavirus RNA can be freely reassorted: Evidence for the role of free leader RNA in transcription; pp. 4204–4208. (Proc. Natl. Acad. Sci. USA). [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Makino S., Taguchi F., Hirano N., Fujiwara K. Analysis of genomic and intracellular viral RNAs of small plaque mutants of mouse hepatitis virus, JHM strain. Virology. 1984;139:138–151. doi: 10.1016/0042-6822(84)90335-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Maxam A.M., Gilbert W. Vol. 74. 1977. A new method for sequencing DNA; pp. 560–564. (Proc. Natl. Acad. Sci. USA). [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Maxam A.M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. In: Grossman L., Moldave K., editors. Vol. 65. Academic Press; Orlando, FL: 1980. pp. 499–560. (Methods in Enzymology). [DOI] [PubMed] [Google Scholar]
  35. McIntosh K., Chao R.K., Krause H.E., Wasil R., Mocega H.E., Mufson M.A. Coronavirus infection in acute lower respiratory tract disease of infants. J. Infect. Dis. 1974;139:502–510. doi: 10.1093/infdis/130.5.502. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. McMaster G.K., Carmichael G.G. Vol. 74. 1977. Analysis of single and double-stranded nucleic acids on polyacrylamide and agarose gels by using glyoxal and acridine orange; pp. 4835–4838. (Proc. Natl. Acad. Sci. USA). [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Messing J., Vierira J. A new pair of M13 vectors for selecting either DNA strand of double-digest restriction fragments. Gene. 1982;19:269–276. doi: 10.1016/0378-1119(82)90016-6. [DOI] [PubMed] [Google Scholar]
  38. Pedersen F.S., Haseltine W.A. A micromethod for detailed characterization of high molecular weight RNA. In: Grossman L., Moldave K., editors. Vol. 65. Academic Press; Orlando, FL: 1980. pp. 680–687. (Methods in Enzymology). [DOI] [PubMed] [Google Scholar]
  39. Pedersen N.C., Ward J., Mengeling W.L. Antigenic relationship of the feline infectious peritonitis virus to coronaviruses of other species. Arch. Virol. 1978;58:45–53. doi: 10.1007/BF01315534. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Resta S., Luby J.P., Rosenfeld C.R., Siegel J.D. Isolation and propagation of a human enteric coronavirus. Science. 1985;229:978–981. doi: 10.1126/science.2992091. [DOI] [PubMed] [Google Scholar]
  41. Rottier P.J.M., Spaan W.J.M., Horzinek N.C., Van Der Zeijst B.A.M. Translation of three mouse hepatitis virus strain A59 subgenomic RNAs in Xenopus laevis oocytes. J. Virol. 1981;38:20–26. doi: 10.1128/jvi.38.1.20-26.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Sanger F., Nicklen S., Coulson A.R. Vol. 74. 1977. DNA sequencing with chain-terminating inhibitors; pp. 5463–5467. (Proc. Natl. Acad. Sci. USA). [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Shieh C.-K., Soe L.H., Making S., Chang M.-F., Stohlman S.A., Lai M.M.C. The 5′-end sequence of the murine coronavirus genome: Implications for multiple fusion sites in leader-primed transcription. Virology. 1987;156:321–330. doi: 10.1016/0042-6822(87)90412-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Skinner M., Siddell S. Nucleotide sequencing of mouse hepatitis virus strain JHM messenger RNA 7. In: Rottier P.J.M., van de Zeijst B.A.M., Spaan W.J.M., Horzinek M.C., editors. Vol. 173. Plenum; New York/London: 1984. pp. 163–173. (Molecular Biology and Pathogenesis of Coronaviruses). [DOI] [PubMed] [Google Scholar]
  45. Stohlman S.A., Lai M.M.C. Phosphoproteins of murine hepatitis virus. J. Virol. 1979;32:672–675. doi: 10.1128/jvi.32.2.672-675.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Tabor S., Richardson C.C. Vol. 82. 1985. A bacteriophage T7 RNA polymerase/promoter system for controlled exclusive expression of specific genes; pp. 1074–1078. (Proc. Natl. Acad. Sci. USA). [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Thomas P.S. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc. Natl. Acad. Sci. USA. 1980;77:5201–5205. doi: 10.1073/pnas.77.9.5201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Wege H., Siddell S., Ter Mellen V. The biology and pathogenesis of coronaviruses. Curr. Top. Microbiol. Immunol. 1982;99:165–200. doi: 10.1007/978-3-642-68528-6_5. [DOI] [PubMed] [Google Scholar]
  49. Weiner L.P., Stohlman S.A. Viral models of demyelination. Neurology. 1978;28:111–114. doi: 10.1212/wnl.28.9_part_2.111. [DOI] [PubMed] [Google Scholar]
  50. Weiss S.R., Leibowitz J.L. Comparison of the RNAs of murine and human coronaviruses. In: ter Meulen V., Siddell S., Wage H., editors. Vol. 142. Plenum; New York/London: 1981. pp. 43–69. (Biochemistry and Biology of Coronaviruses). [Google Scholar]

Articles from Virology are provided here courtesy of Elsevier

RESOURCES