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. 1986 Aug;5(8):1769–1774. doi: 10.1002/j.1460-2075.1986.tb04425.x

Modulation of replication, aminoacylation and adenylation in vitro and infectivity in vivo of BMV RNAs containing deletions within the multifunctional 3' end.

J J Bujarski, P Ahlquist, T C Hall, T W Dreher, P Kaesberg
PMCID: PMC1167039  PMID: 3758026

Abstract

The genome of brome mosaic virus (BMV) is comprised of three (+) strand RNAs, each containing a similar, highly structured, 200 base long sequence at its 3' end. A 134 base subset of this sequence contains signals directing interaction of the viral RNA with BMV RNA replicase, ATP,CTP:tRNA nucleotidyl transferase and aminoacyl tRNA synthetase. A series of mutants containing deletions within this region, previously constructed and tested in vitro for the effect on replication and aminoacylation activities, has now been assayed in vitro for adenylation function and in vivo for ability to replicate in isolated protoplasts and whole plants. These tests indicate that features of viral RNA recognized by BMV replicase overlap those directing adenylation, but are distinct from those directing aminoacylation. Consequently, the lethality of a deletion preferentially inhibiting aminoacylation suggests that this function may have an essential role contributing to viral replication in vivo. An RNA3 mutant bearing a 20-base deletion yielding normal levels of aminoacylation and enhanced levels of replicase template activity and adenylation in vitro was able to replicate in protoplasts and plants; however, its accumulation in protoplasts was reduced relative to wild-type. This suggests that additional functions affecting the replication and accumulation of viral RNA reside in the conserved 3' sequence.

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Selected References

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  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., 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]
  6. 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]
  7. Bujarski J. J., Kaesberg P. Genetic recombination between RNA components of a multipartite plant virus. 1986 May 29-Jun 4Nature. 321(6069):528–531. doi: 10.1038/321528a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Donis-Keller H. Phy M: an RNase activity specific for U and A residues useful in RNA sequence analysis. Nucleic Acids Res. 1980 Jul 25;8(14):3133–3142. doi: 10.1093/nar/8.14.3133. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dreher T. W., Bujarski J. J., Hall T. C. Mutant viral RNAs synthesized in vitro show altered aminoacylation and replicase template activities. Nature. 1984 Sep 13;311(5982):171–175. doi: 10.1038/311171a0. [DOI] [PubMed] [Google Scholar]
  10. England T. E., Bruce A. G., Uhlenbeck O. C. Specific labeling of 3' termini of RNA with T4 RNA ligase. Methods Enzymol. 1980;65(1):65–74. doi: 10.1016/s0076-6879(80)65011-3. [DOI] [PubMed] [Google Scholar]
  11. 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]
  12. Hall T. C. Transfer RNA-like structures in viral genomes. Int Rev Cytol. 1979;60:1–26. doi: 10.1016/s0074-7696(08)61257-7. [DOI] [PubMed] [Google Scholar]
  13. 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]
  14. Joshi R. L., Joshi S., Chapeville F., Haenni A. L. tRNA-like structures of plant viral RNAs: conformational requirements for adenylation and aminoacylation. EMBO J. 1983;2(7):1123–1127. doi: 10.1002/j.1460-2075.1983.tb01556.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kao T., Miller D. L., Abo M., Ofengand J. Formation and properties of a covalent complex between elongation factor Tu and Phe-tRNA bearing a photoaffinity probe on its 3-(3-amino-3-carboxypropyl)uridine residue. J Mol Biol. 1983 May 25;166(3):383–405. doi: 10.1016/s0022-2836(83)80091-6. [DOI] [PubMed] [Google Scholar]
  16. Lane L. C. The bromoviruses. Adv Virus Res. 1974;19:151–220. doi: 10.1016/s0065-3527(08)60660-0. [DOI] [PubMed] [Google Scholar]
  17. Miller W. A., Bujarski J. J., Dreher T. W., Hall T. C. Minus-strand initiation by brome mosaic virus replicase within the 3' tRNA-like structure of native and modified RNA templates. J Mol Biol. 1986 Feb 20;187(4):537–546. doi: 10.1016/0022-2836(86)90332-3. [DOI] [PubMed] [Google Scholar]
  18. Miller W. A., Dreher T. W., Hall T. C. Synthesis of brome mosaic virus subgenomic RNA in vitro by internal initiation on (-)-sense genomic RNA. Nature. 1985 Jan 3;313(5997):68–70. doi: 10.1038/313068a0. [DOI] [PubMed] [Google Scholar]
  19. Rietveld K., Linschooten K., Pleij C. W., Bosch L. The three-dimensional folding of the tRNA-like structure of tobacco mosaic virus RNA. A new building principle applied twice. EMBO J. 1984 Nov;3(11):2613–2619. doi: 10.1002/j.1460-2075.1984.tb02182.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Rietveld K., Pleij C. W., Bosch L. Three-dimensional models of the tRNA-like 3' termini of some plant viral RNAs. EMBO J. 1983;2(7):1079–1085. doi: 10.1002/j.1460-2075.1983.tb01549.x. [DOI] [PMC free article] [PubMed] [Google Scholar]

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