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. 1992 Jun;66(6):3339–3346. doi: 10.1128/jvi.66.6.3339-3346.1992

Mechanism of coronavirus transcription: duration of primary transcription initiation activity and effects of subgenomic RNA transcription on RNA replication.

Y S Jeong 1, S Makino 1
PMCID: PMC241112  PMID: 1583719

Abstract

Previously, we established a system whereby an intergenic region from mouse hepatitis virus (MHV) inserted into an MHV defective interfering (DI) RNA led to transcription of a subgenomic DI RNA in helper virus-infected cells. By using this system, the duration of a primary transcription initiation activity which transcribes subgenomic-size RNAs from the genomic-size RNA template in MHV-infected cells was examined. Efficient DI genomic and subgenomic RNA synthesis was observed when the DI RNA was transfected at 1, 3, 3.5, 5, and 6 h postinfection, indicating that all activities which are necessary for MHV RNA synthesis are present continuously during the first 6 h of infection. The effect of subgenomic DI RNA synthesis on DI genomic RNA replication was then examined. Replication efficiency of the DI genomic RNA which synthesized the subgenomic RNA was approximately 70% lower than that of DI genomic RNA which did not synthesize the subgenomic DI RNA in MHV-infected cells. Cotransfection of two different-size DI RNAs demonstrated that replication of the larger DI RNA was strongly inhibited by replication of the smaller genomic DI RNA. Cotransfection of two DI RNA species of the same length into MHV-infected cells demonstrated that reduced replication of the genomic DI RNA which synthesizes the subgenomic RNA did not affect the replication of cotransfected DI RNA, demonstrating that the reduction in DI genomic RNA replication works only in cis, not in trans. Therefore, the previously proposed hypothesis that coronavirus, subgenomic RNA synthesis may inhibit the replication of genomic RNA by competing for a limited amount of virus-derived factors seems unlikely. Possible mechanisms of coronavirus transcription are discussed.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Baker S. C., Lai M. M. An in vitro system for the leader-primed transcription of coronavirus mRNAs. EMBO J. 1990 Dec;9(12):4173–4179. doi: 10.1002/j.1460-2075.1990.tb07641.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Baric R. S., Stohlman S. A., Razavi M. K., Lai M. M. Characterization of leader-related small RNAs in coronavirus-infected cells: further evidence for leader-primed mechanism of transcription. Virus Res. 1985 Jul;3(1):19–33. doi: 10.1016/0168-1702(85)90038-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Brayton P. R., Lai M. M., Patton C. D., Stohlman S. A. Characterization of two RNA polymerase activities induced by mouse hepatitis virus. J Virol. 1982 Jun;42(3):847–853. doi: 10.1128/jvi.42.3.847-853.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Brayton P. R., Stohlman S. A., Lai M. M. Further characterization of mouse hepatitis virus RNA-dependent RNA polymerases. Virology. 1984 Feb;133(1):197–201. doi: 10.1016/0042-6822(84)90439-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Compton S. R., Rogers D. B., Holmes K. V., Fertsch D., Remenick J., McGowan J. J. In vitro replication of mouse hepatitis virus strain A59. J Virol. 1987 Jun;61(6):1814–1820. doi: 10.1128/jvi.61.6.1814-1820.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Dennis D. E., Brian D. A. RNA-dependent RNA polymerase activity in coronavirus- infected cells. J Virol. 1982 Apr;42(1):153–164. doi: 10.1128/jvi.42.1.153-164.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Felgner P. L., Gadek T. R., Holm M., Roman R., Chan H. W., Wenz M., Northrop J. P., Ringold G. M., Danielsen M. Lipofection: a highly efficient, lipid-mediated DNA-transfection procedure. Proc Natl Acad Sci U S A. 1987 Nov;84(21):7413–7417. doi: 10.1073/pnas.84.21.7413. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Lai M. M., Baric R. S., Brayton P. R., Stohlman S. A. Characterization of leader RNA sequences on the virion and mRNAs of mouse hepatitis virus, a cytoplasmic RNA virus. Proc Natl Acad Sci U S A. 1984 Jun;81(12):3626–3630. doi: 10.1073/pnas.81.12.3626. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Lai M. M., Brayton P. R., Armen R. C., Patton C. D., Pugh C., Stohlman S. A. Mouse hepatitis virus A59: mRNA structure and genetic localization of the sequence divergence from hepatotropic strain MHV-3. J Virol. 1981 Sep;39(3):823–834. doi: 10.1128/jvi.39.3.823-834.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Lai M. M. Coronavirus: organization, replication and expression of genome. Annu Rev Microbiol. 1990;44:303–333. doi: 10.1146/annurev.mi.44.100190.001511. [DOI] [PubMed] [Google Scholar]
  11. Lai M. M., Patton C. D., Baric R. S., Stohlman S. A. Presence of leader sequences in the mRNA of mouse hepatitis virus. J Virol. 1983 Jun;46(3):1027–1033. doi: 10.1128/jvi.46.3.1027-1033.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Lai M. M., Stohlman S. A. RNA of mouse hepatitis virus. J Virol. 1978 May;26(2):236–242. doi: 10.1128/jvi.26.2.236-242.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Lee H. J., Shieh C. K., Gorbalenya A. E., Koonin E. V., La Monica N., Tuler J., Bagdzhadzhyan A., Lai M. M. The complete sequence (22 kilobases) of murine coronavirus gene 1 encoding the putative proteases and RNA polymerase. Virology. 1991 Feb;180(2):567–582. doi: 10.1016/0042-6822(91)90071-I. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Leibowitz J. L., DeVries J. R. Synthesis of virus-specific RNA in permeabilized murine coronavirus-infected cells. Virology. 1988 Sep;166(1):66–75. doi: 10.1016/0042-6822(88)90147-X. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Leibowitz J. L., Wilhelmsen K. C., Bond C. W. The virus-specific intracellular RNA species of two murine coronaviruses: MHV-a59 and MHV-JHM. Virology. 1981 Oct 15;114(1):39–51. doi: 10.1016/0042-6822(81)90250-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Mahy B. W., Siddell S., Wege H., ter Meulen V. RNA-dependent RNA polymerase activity in murine coronavirus-infected cells. J Gen Virol. 1983 Jan;64(Pt 1):103–111. doi: 10.1099/0022-1317-64-1-103. [DOI] [PubMed] [Google Scholar]
  17. Makino S., Fujioka N., Fujiwara K. Structure of the intracellular defective viral RNAs of defective interfering particles of mouse hepatitis virus. J Virol. 1985 May;54(2):329–336. doi: 10.1128/jvi.54.2.329-336.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Makino S., Joo M., Makino J. K. A system for study of coronavirus mRNA synthesis: a regulated, expressed subgenomic defective interfering RNA results from intergenic site insertion. J Virol. 1991 Nov;65(11):6031–6041. doi: 10.1128/jvi.65.11.6031-6041.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Makino S., Lai M. M. High-frequency leader sequence switching during coronavirus defective interfering RNA replication. J Virol. 1989 Dec;63(12):5285–5292. doi: 10.1128/jvi.63.12.5285-5292.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Makino S., Shieh C. K., Keck J. G., Lai M. M. Defective-interfering particles of murine coronavirus: mechanism of synthesis of defective viral RNAs. Virology. 1988 Mar;163(1):104–111. doi: 10.1016/0042-6822(88)90237-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Makino S., Shieh C. K., Soe L. H., Baker S. C., Lai M. M. Primary structure and translation of a defective interfering RNA of murine coronavirus. Virology. 1988 Oct;166(2):550–560. doi: 10.1016/0042-6822(88)90526-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Makino S., Soe L. H., Shieh C. K., Lai M. M. Discontinuous transcription generates heterogeneity at the leader fusion sites of coronavirus mRNAs. J Virol. 1988 Oct;62(10):3870–3873. doi: 10.1128/jvi.62.10.3870-3873.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Makino S., Stohlman S. A., Lai M. M. Leader sequences of murine coronavirus mRNAs can be freely reassorted: evidence for the role of free leader RNA in transcription. Proc Natl Acad Sci U S A. 1986 Jun;83(12):4204–4208. doi: 10.1073/pnas.83.12.4204. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. 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 Nov;139(1):138–151. doi: 10.1016/0042-6822(84)90335-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Sawicki S. G., Sawicki D. L. Coronavirus minus-strand RNA synthesis and effect of cycloheximide on coronavirus RNA synthesis. J Virol. 1986 Jan;57(1):328–334. doi: 10.1128/jvi.57.1.328-334.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Sawicki S. G., Sawicki D. L. Coronavirus transcription: subgenomic mouse hepatitis virus replicative intermediates function in RNA synthesis. J Virol. 1990 Mar;64(3):1050–1056. doi: 10.1128/jvi.64.3.1050-1056.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Sethna P. B., Hofmann M. A., Brian D. A. Minus-strand copies of replicating coronavirus mRNAs contain antileaders. J Virol. 1991 Jan;65(1):320–325. doi: 10.1128/jvi.65.1.320-325.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Sethna P. B., Hung S. L., Brian D. A. Coronavirus subgenomic minus-strand RNAs and the potential for mRNA replicons. Proc Natl Acad Sci U S A. 1989 Jul;86(14):5626–5630. doi: 10.1073/pnas.86.14.5626. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Shieh C. K., Soe L. H., Makino S., Chang M. F., Stohlman S. A., Lai M. M. The 5'-end sequence of the murine coronavirus genome: implications for multiple fusion sites in leader-primed transcription. Virology. 1987 Feb;156(2):321–330. doi: 10.1016/0042-6822(87)90412-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. van der Most R. G., Bredenbeek P. J., Spaan W. J. A domain at the 3' end of the polymerase gene is essential for encapsidation of coronavirus defective interfering RNAs. J Virol. 1991 Jun;65(6):3219–3226. doi: 10.1128/jvi.65.6.3219-3226.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]

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