Production of infectious human respiratory syncytial virus from cloned cDNA confirms an essential role for the transcription elongation factor from the 5' proximal open reading frame of the M2 mRNA in gene expression and provides a capability for vaccine development (original) (raw)

Abstract

Infectious human respiratory syncytial virus (RSV) was produced by the intracellular coexpression of five plasmid-borne cDNAs. One cDNA encoded a complete positive-sense version of the RSV genome (corresponding to the replicative intermediate RNA or antigenome), and each of the other four encoded a separate RSV protein, namely, the major nucleocapsid N protein, the nucleocapsid P phosphoprotein, the major polymerase L protein, or the protein from the 5' proximal open reading frame of the M2 mRNA [M2(ORF1)]. RSV was not produced if any of the five plasmids was omitted. The requirement for the M2(ORF1) protein is consistent with its recent identification as a transcription elongation factor and confirms its importance for RSV gene expression. It should thus be possible to introduce defined changes into infectious RSV. This should be useful for basic studies of RSV molecular biology and pathogenesis; in addition, there are immediate applications to the development of live attenuated vaccine strains bearing predetermined defined attenuating mutations.

11563

Images in this article

Selected References

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

  1. Calain P., Roux L. The rule of six, a basic feature for efficient replication of Sendai virus defective interfering RNA. J Virol. 1993 Aug;67(8):4822–4830. doi: 10.1128/jvi.67.8.4822-4830.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Collins P. L., Hill M. G., Johnson P. R. The two open reading frames of the 22K mRNA of human respiratory syncytial virus: sequence comparison of antigenic subgroups A and B and expression in vitro. J Gen Virol. 1990 Dec;71(Pt 12):3015–3020. doi: 10.1099/0022-1317-71-12-3015. [DOI] [PubMed] [Google Scholar]
  3. Connors M., Crowe J. E., Jr, Firestone C. Y., Murphy B. R., Collins P. L. A cold-passaged, attenuated strain of human respiratory syncytial virus contains mutations in the F and L genes. Virology. 1995 Apr 20;208(2):478–484. doi: 10.1006/viro.1995.1178. [DOI] [PubMed] [Google Scholar]
  4. Conzelmann K. K., Schnell M. Rescue of synthetic genomic RNA analogs of rabies virus by plasmid-encoded proteins. J Virol. 1994 Feb;68(2):713–719. doi: 10.1128/jvi.68.2.713-719.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Crowe J. E., Jr, Bui P. T., Davis A. R., Chanock R. M., Murphy B. R. A further attenuated derivative of a cold-passaged temperature-sensitive mutant of human respiratory syncytial virus retains immunogenicity and protective efficacy against wild-type challenge in seronegative chimpanzees. Vaccine. 1994 Jul;12(9):783–790. doi: 10.1016/0264-410x(94)90286-0. [DOI] [PubMed] [Google Scholar]
  6. Grosfeld H., Hill M. G., Collins P. L. RNA replication by respiratory syncytial virus (RSV) is directed by the N, P, and L proteins; transcription also occurs under these conditions but requires RSV superinfection for efficient synthesis of full-length mRNA. J Virol. 1995 Sep;69(9):5677–5686. doi: 10.1128/jvi.69.9.5677-5686.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hall C. B. Prospects for a respiratory syncytial virus vaccine. Science. 1994 Sep 2;265(5177):1393–1394. doi: 10.1126/science.7915433. [DOI] [PubMed] [Google Scholar]
  8. Johnson P. R., Collins P. L. The A and B subgroups of human respiratory syncytial virus: comparison of intergenic and gene-overlap sequences. J Gen Virol. 1988 Nov;69(Pt 11):2901–2906. doi: 10.1099/0022-1317-69-11-2901. [DOI] [PubMed] [Google Scholar]
  9. Lawson N. D., Stillman E. A., Whitt M. A., Rose J. K. Recombinant vesicular stomatitis viruses from DNA. Proc Natl Acad Sci U S A. 1995 May 9;92(10):4477–4481. doi: 10.1073/pnas.92.10.4477. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Murphy B. R., Hall S. L., Kulkarni A. B., Crowe J. E., Jr, Collins P. L., Connors M., Karron R. A., Chanock R. M. An update on approaches to the development of respiratory syncytial virus (RSV) and parainfluenza virus type 3 (PIV3) vaccines. Virus Res. 1994 Apr;32(1):13–36. doi: 10.1016/0168-1702(94)90059-0. [DOI] [PubMed] [Google Scholar]
  11. Murphy B. R., Sotnikov A. V., Lawrence L. A., Banks S. M., Prince G. A. Enhanced pulmonary histopathology is observed in cotton rats immunized with formalin-inactivated respiratory syncytial virus (RSV) or purified F glycoprotein and challenged with RSV 3-6 months after immunization. Vaccine. 1990 Oct;8(5):497–502. doi: 10.1016/0264-410x(90)90253-i. [DOI] [PubMed] [Google Scholar]
  12. Pattnaik A. K., Ball L. A., LeGrone A. W., Wertz G. W. Infectious defective interfering particles of VSV from transcripts of a cDNA clone. Cell. 1992 Jun 12;69(6):1011–1020. doi: 10.1016/0092-8674(92)90619-n. [DOI] [PubMed] [Google Scholar]
  13. Randhawa J. S., Chambers P., Pringle C. R., Easton A. J. Nucleotide sequences of the genes encoding the putative attachment glycoprotein (G) of mouse and tissue culture-passaged strains of pneumonia virus of mice. Virology. 1995 Feb 20;207(1):240–245. doi: 10.1006/viro.1995.1071. [DOI] [PubMed] [Google Scholar]
  14. Schnell M. J., Mebatsion T., Conzelmann K. K. Infectious rabies viruses from cloned cDNA. EMBO J. 1994 Sep 15;13(18):4195–4203. doi: 10.1002/j.1460-2075.1994.tb06739.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Sidhu M. S., Chan J., Kaelin K., Spielhofer P., Radecke F., Schneider H., Masurekar M., Dowling P. C., Billeter M. A., Udem S. A. Rescue of synthetic measles virus minireplicons: measles genomic termini direct efficient expression and propagation of a reporter gene. Virology. 1995 Apr 20;208(2):800–807. doi: 10.1006/viro.1995.1215. [DOI] [PubMed] [Google Scholar]
  16. Wyatt L. S., Moss B., Rozenblatt S. Replication-deficient vaccinia virus encoding bacteriophage T7 RNA polymerase for transient gene expression in mammalian cells. Virology. 1995 Jun 20;210(1):202–205. doi: 10.1006/viro.1995.1332. [DOI] [PubMed] [Google Scholar]
  17. Yu Q., Hardy R. W., Wertz G. W. Functional cDNA clones of the human respiratory syncytial (RS) virus N, P, and L proteins support replication of RS virus genomic RNA analogs and define minimal trans-acting requirements for RNA replication. J Virol. 1995 Apr;69(4):2412–2419. doi: 10.1128/jvi.69.4.2412-2419.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Zoller M. J., Smith M. Oligonucleotide-directed mutagenesis: a simple method using two oligonucleotide primers and a single-stranded DNA template. DNA. 1984 Dec;3(6):479–488. doi: 10.1089/dna.1.1984.3.479. [DOI] [PubMed] [Google Scholar]