Skip to main content
PMC home page
Journal of Virology logoLink to Journal of Virology
. 1988 Oct;62(10):3581–3588. doi: 10.1128/jvi.62.10.3581-3588.1988

Infectious in vitro transcripts from cowpea chlorotic mottle virus cDNA clones and exchange of individual RNA components with brome mosaic virus.

R F Allison 1, M Janda 1, P Ahlquist 1
PMCID: PMC253497  PMID: 3418781

Abstract

Complete cDNA copies of genomic RNA1, RNA2, and RNA3 of cowpea chlorotic mottle virus (CCMV) were cloned 1 base downstream from a T7 RNA polymerase promoter. The mixture of capped in vitro transcripts from all three clones produced normal CCMV infections in barley protoplasts and cowpea plants. By using transcripts from these clones and from a similar set of biologically active clones of the related brome mosaic virus (BMV), all possible single component exchanges between the BMV and CCMV tripartite genomes were tested. Viral RNA replication was not observed with any heterologous combination of RNA1 and RNA2, which encode trans-acting viral RNA replication factors. However, substitution of the heterologous RNA3 into either genome produced viable hybrid viruses, both of which replicated in barley protoplasts and produced lesions on Chenopodium hybridum, a local lesion host for both parent viruses. In hybrid infections, BMV and CCMV coat proteins each readily packaged RNAs from the heterologous virus, but BMV RNAs were replicated to a higher level than CCMV RNAs, even when trans-acting RNA replication factors were provided by CCMV genes. Neither hybrid systemically infected the natural host of either parent virus, suggesting that host specificity determinants in BMV and CCMV are encoded by RNA3 and at least one other genomic RNA.

Full text

PDF
3583

Images in this article

3583Image
on p.3583
3583Image
on p.3583
3584Image
on p.3584
3585Image
on p.3585
3585Image
on p.3585

Selected References

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

  1. Ahlquist P., Dasgupta R., Kaesberg P. Near identity of 3- RNA secondary structure in bromoviruses and cucumber mosaic virus. Cell. 1981 Jan;23(1):183–189. doi: 10.1016/0092-8674(81)90283-x. [DOI] [PubMed] [Google Scholar]
  2. Ahlquist P., Dasgupta R., Kaesberg P. Nucleotide sequence of the brome mosaic virus genome and its implications for viral replication. J Mol Biol. 1984 Feb 5;172(4):369–383. doi: 10.1016/s0022-2836(84)80012-1. [DOI] [PubMed] [Google Scholar]
  3. Ahlquist P., French R., Janda M., Loesch-Fries L. S. Multicomponent RNA plant virus infection derived from cloned viral cDNA. Proc Natl Acad Sci U S A. 1984 Nov;81(22):7066–7070. doi: 10.1073/pnas.81.22.7066. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Ahlquist P., Janda M. cDNA cloning and in vitro transcription of the complete brome mosaic virus genome. Mol Cell Biol. 1984 Dec;4(12):2876–2882. doi: 10.1128/mcb.4.12.2876. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Ahlquist P., Luckow V., Kaesberg P. Complete nucleotide sequence of brome mosaic virus RNA3. J Mol Biol. 1981 Nov 25;153(1):23–38. doi: 10.1016/0022-2836(81)90524-6. [DOI] [PubMed] [Google Scholar]
  6. Ahlquist P., Strauss E. G., Rice C. M., Strauss J. H., Haseloff J., Zimmern D. Sindbis virus proteins nsP1 and nsP2 contain homology to nonstructural proteins from several RNA plant viruses. J Virol. 1985 Feb;53(2):536–542. doi: 10.1128/jvi.53.2.536-542.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Atabekov J. G., Dorokhov YuL Plant virus-specific transport function and resistance of plants to viruses. Adv Virus Res. 1984;29:313–364. doi: 10.1016/s0065-3527(08)60412-1. [DOI] [PubMed] [Google Scholar]
  8. Biggin M. D., Gibson T. J., Hong G. F. Buffer gradient gels and 35S label as an aid to rapid DNA sequence determination. Proc Natl Acad Sci U S A. 1983 Jul;80(13):3963–3965. doi: 10.1073/pnas.80.13.3963. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Bujarski J. J., Ahlquist P., Hall T. C., Dreher T. W., Kaesberg P. Modulation of replication, aminoacylation and adenylation in vitro and infectivity in vivo of BMV RNAs containing deletions within the multifunctional 3' end. EMBO J. 1986 Aug;5(8):1769–1774. doi: 10.1002/j.1460-2075.1986.tb04425.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Bujarski J. J., Dreher T. W., Hall T. C. Deletions in the 3'-terminal tRNA-like structure of brome mosaic virus RNA differentially affect aminoacylation and replication in vitro. Proc Natl Acad Sci U S A. 1985 Sep;82(17):5636–5640. doi: 10.1073/pnas.82.17.5636. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Dasgupta R., Kaesberg P. Complete nucleotide sequences of the coat protein messenger RNAs of brome mosaic virus and cowpea chlorotic mottle virus. Nucleic Acids Res. 1982 Jan 22;10(2):703–713. doi: 10.1093/nar/10.2.703. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Dasmahapatra B., Dasgupta R., Saunders K., Selling B., Gallagher T., Kaesberg P. Infectious RNA derived by transcription from cloned cDNA copies of the genomic RNA of an insect virus. Proc Natl Acad Sci U S A. 1986 Jan;83(1):63–66. doi: 10.1073/pnas.83.1.63. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Dawson W. O., Beck D. L., Knorr D. A., Grantham G. L. cDNA cloning of the complete genome of tobacco mosaic virus and production of infectious transcripts. Proc Natl Acad Sci U S A. 1986 Mar;83(6):1832–1836. doi: 10.1073/pnas.83.6.1832. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Deom C. M., Oliver M. J., Beachy R. N. The 30-kilodalton gene product of tobacco mosaic virus potentiates virus movement. Science. 1987 Jul 24;237(4813):389–394. doi: 10.1126/science.237.4813.389. [DOI] [PubMed] [Google Scholar]
  15. French R., Ahlquist P. Characterization and engineering of sequences controlling in vivo synthesis of brome mosaic virus subgenomic RNA. J Virol. 1988 Jul;62(7):2411–2420. doi: 10.1128/jvi.62.7.2411-2420.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. French R., Ahlquist P. Intercistronic as well as terminal sequences are required for efficient amplification of brome mosaic virus RNA3. J Virol. 1987 May;61(5):1457–1465. doi: 10.1128/jvi.61.5.1457-1465.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. French R., Janda M., Ahlquist P. Bacterial gene inserted in an engineered RNA virus: efficient expression in monocotyledonous plant cells. Science. 1986 Mar 14;231(4743):1294–1297. doi: 10.1126/science.231.4743.1294. [DOI] [PubMed] [Google Scholar]
  18. Goldbach R. Genome similarities between plant and animal RNA viruses. Microbiol Sci. 1987 Jul;4(7):197–202. [PubMed] [Google Scholar]
  19. Gubler U., Hoffman B. J. A simple and very efficient method for generating cDNA libraries. Gene. 1983 Nov;25(2-3):263–269. doi: 10.1016/0378-1119(83)90230-5. [DOI] [PubMed] [Google Scholar]
  20. Gupta K. C., Kingsbury D. W. Complete sequences of the intergenic and mRNA start signals in the Sendai virus genome: homologies with the genome of vesicular stomatitis virus. Nucleic Acids Res. 1984 May 11;12(9):3829–3841. doi: 10.1093/nar/12.9.3829. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Haseloff J., Goelet P., Zimmern D., Ahlquist P., Dasgupta R., Kaesberg P. Striking similarities in amino acid sequence among nonstructural proteins encoded by RNA viruses that have dissimilar genomic organization. Proc Natl Acad Sci U S A. 1984 Jul;81(14):4358–4362. doi: 10.1073/pnas.81.14.4358. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Janda M., French R., Ahlquist P. High efficiency T7 polymerase synthesis of infectious RNA from cloned brome mosaic virus cdna and effects of 5' extensions on transcript infectivity. Virology. 1987 May;158(1):259–262. doi: 10.1016/0042-6822(87)90265-0. [DOI] [PubMed] [Google Scholar]
  23. Kaplan G., Lubinski J., Dasgupta A., Racaniello V. R. In vitro synthesis of infectious poliovirus RNA. Proc Natl Acad Sci U S A. 1985 Dec;82(24):8424–8428. doi: 10.1073/pnas.82.24.8424. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kiberstis P. A., Loesch-Fries L. S., Hall T. C. Viral protein synthesis in barley protoplasts inoculated with native and fractionated brome mosaic virus RNA. Virology. 1981 Jul 30;112(2):804–808. doi: 10.1016/0042-6822(81)90331-7. [DOI] [PubMed] [Google Scholar]
  25. Knorr D. A., Dawson W. O. A point mutation in the tobacco mosaic virus capsid protein gene induces hypersensitivity in Nicotiana sylvestris. Proc Natl Acad Sci U S A. 1988 Jan;85(1):170–174. doi: 10.1073/pnas.85.1.170. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Meshi T., Ishikawa M., Motoyoshi F., Semba K., Okada Y. In vitro transcription of infectious RNAs from full-length cDNAs of tobacco mosaic virus. Proc Natl Acad Sci U S A. 1986 Jul;83(14):5043–5047. doi: 10.1073/pnas.83.14.5043. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Meshi T., Watanabe Y., Saito T., Sugimoto A., Maeda T., Okada Y. Function of the 30 kd protein of tobacco mosaic virus: involvement in cell-to-cell movement and dispensability for replication. EMBO J. 1987 Sep;6(9):2557–2563. doi: 10.1002/j.1460-2075.1987.tb02544.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Miller W. A., Hall T. C. RNA-dependent RNA polymerase isolated from cowpea chlorotic mottle virus-infected cowpeas is specific for bromoviral RNA. Virology. 1984 Jan 15;132(1):53–60. doi: 10.1016/0042-6822(84)90090-4. [DOI] [PubMed] [Google Scholar]
  29. Miller W. A., Hall T. C. Use of micrococcal nuclease in the purification of highly template dependent RNA-dependent RNA polymerase from brome mosaic virus-infected barley. Virology. 1983 Feb;125(1):236–241. doi: 10.1016/0042-6822(83)90077-6. [DOI] [PubMed] [Google Scholar]
  30. Mizutani S., Colonno R. J. In vitro synthesis of an infectious RNA from cDNA clones of human rhinovirus type 14. J Virol. 1985 Nov;56(2):628–632. doi: 10.1128/jvi.56.2.628-632.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Rice C. M., Levis R., Strauss J. H., Huang H. V. Production of infectious RNA transcripts from Sindbis virus cDNA clones: mapping of lethal mutations, rescue of a temperature-sensitive marker, and in vitro mutagenesis to generate defined mutants. J Virol. 1987 Dec;61(12):3809–3819. doi: 10.1128/jvi.61.12.3809-3819.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Rybicki E. P., Von Wechmar M. B. The serology of the bromoviruses. I. Serological interrelationships of the bromoviruses. Virology. 1981 Mar;109(2):391–402. doi: 10.1016/0042-6822(81)90509-2. [DOI] [PubMed] [Google Scholar]
  33. Vieira J., Messing J. Production of single-stranded plasmid DNA. Methods Enzymol. 1987;153:3–11. doi: 10.1016/0076-6879(87)53044-0. [DOI] [PubMed] [Google Scholar]
  34. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]
  35. Zimmern D., Hunter T. Point mutation in the 30-K open reading frame of TMV implicated in temperature-sensitive assembly and local lesion spreading of mutant Ni 2519. EMBO J. 1983;2(11):1893–1900. doi: 10.1002/j.1460-2075.1983.tb01676.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. van der Werf S., Bradley J., Wimmer E., Studier F. W., Dunn J. J. Synthesis of infectious poliovirus RNA by purified T7 RNA polymerase. Proc Natl Acad Sci U S A. 1986 Apr;83(8):2330–2334. doi: 10.1073/pnas.83.8.2330. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Virology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES