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. 1993 May;5(5):577–586. doi: 10.1105/tpc.5.5.577

Analysis of a tobacco mosaic virus strain capable of overcoming N gene-mediated resistance.

H S Padgett 1, R N Beachy 1
PMCID: PMC160295  PMID: 8518557

Abstract

The genome of Ob, a tobamovirus that overcomes the N gene-mediated hypersensitive response (HR), was cloned as a cDNA, and its nucleotide sequence was determined. The genomic organization of Ob is similar to that of other tobamoviruses, consisting of 6506 nucleotides and containing at least four open reading frames. These open reading frames encode a 126-kD polypeptide with a 183-kD readthrough product, a 30.6-kD movement protein, and an 18-kD coat protein. A bacteriophage T7 promoter sequence was fused to the full-length cDNA clone to obtain infectious RNA transcripts. These transcripts, when inoculated onto tobacco plants, induced disease symptoms indistinguishable from plants inoculated with Ob viral RNA. To determine which viral factor is responsible for the resistance-breaking character of Ob, a recombinant virus was constructed in which the movement protein gene of tobacco mosaic virus was replaced with that of Ob. Cultivar Xanthi NN tobacco plants infected with this virus responded with an HR, indicating that the Ob movement protein alone does not act to overcome the N gene-mediated response. Following mutagenesis of the infectious Ob cDNA clone with hydroxylamine, populations of transcripts from the mutagenized DNA were inoculated onto Xanthi NN tobacco, and a variant that induced the HR was identified. The mutant was analyzed and found to contain a single nucleotide change in the 126-kD gene. Recreating the mutation in the Ob cDNA clone by site-directed mutagenesis resulted in a virus that caused symptoms identical to the chemically induced mutant.

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Selected References

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  1. Bruening G., Beachy R. N., Scalla R., Zaitlin M. In vitro and in vivo translation of the ribonucleic acids of a cowpea strain of tobacco mosaic virus. Virology. 1976 Jun;71(2):498–517. doi: 10.1016/0042-6822(76)90377-9. [DOI] [PubMed] [Google Scholar]
  2. Calder V. L., Palukaitis P. Nucleotide sequence analysis of the movement genes of resistance breaking strains of tomato mosaic virus. J Gen Virol. 1992 Jan;73(Pt 1):165–168. doi: 10.1099/0022-1317-73-1-165. [DOI] [PubMed] [Google Scholar]
  3. DeBorde D. C., Naeve C. W., Herlocher M. L., Maassab H. F. Resolution of a common RNA sequencing ambiguity by terminal deoxynucleotidyl transferase. Anal Biochem. 1986 Sep;157(2):275–282. doi: 10.1016/0003-2697(86)90626-3. [DOI] [PubMed] [Google Scholar]
  4. Deom C. M., Schubert K. R., Wolf S., Holt C. A., Lucas W. J., Beachy R. N. Molecular characterization and biological function of the movement protein of tobacco mosaic virus in transgenic plants. Proc Natl Acad Sci U S A. 1990 May;87(9):3284–3288. doi: 10.1073/pnas.87.9.3284. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Deom C. M., Wolf S., Holt C. A., Lucas W. J., Beachy R. N. Altered function of the tobacco mosaic virus movement protein in a hypersensitive host. Virology. 1991 Jan;180(1):251–256. doi: 10.1016/0042-6822(91)90029-b. [DOI] [PubMed] [Google Scholar]
  6. Fichot O., Girard M. An improved method for sequencing of RNA templates. Nucleic Acids Res. 1990 Oct 25;18(20):6162–6162. doi: 10.1093/nar/18.20.6162. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Goelet P., Lomonossoff G. P., Butler P. J., Akam M. E., Gait M. J., Karn J. Nucleotide sequence of tobacco mosaic virus RNA. Proc Natl Acad Sci U S A. 1982 Oct;79(19):5818–5822. doi: 10.1073/pnas.79.19.5818. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. 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]
  9. Holt C. A., Beachy R. N. In vivo complementation of infectious transcripts from mutant tobacco mosaic virus cDNAs in transgenic plants. Virology. 1991 Mar;181(1):109–117. doi: 10.1016/0042-6822(91)90475-q. [DOI] [PubMed] [Google Scholar]
  10. Holt C. A., Hodgson R. A., Coker F. A., Beachy R. N., Nelson R. S. Characterization of the masked strain of tobacco mosaic virus: identification of the region responsible for symptom attenuation by analysis of an infectious cDNA clone. Mol Plant Microbe Interact. 1990 Nov-Dec;3(6):417–423. doi: 10.1094/mpmi-3-417. [DOI] [PubMed] [Google Scholar]
  11. 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]
  12. Meshi T., Motoyoshi F., Adachi A., Watanabe Y., Takamatsu N., Okada Y. Two concomitant base substitutions in the putative replicase genes of tobacco mosaic virus confer the ability to overcome the effects of a tomato resistance gene, Tm-1. EMBO J. 1988 Jun;7(6):1575–1581. doi: 10.1002/j.1460-2075.1988.tb02982.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Nejidat A., Cellier F., Holt C. A., Gafny R., Eggenberger A. L., Beachy R. N. Transfer of the movement protein gene between two tobamoviruses: influence on local lesion development. Virology. 1991 Jan;180(1):318–326. doi: 10.1016/0042-6822(91)90036-b. [DOI] [PubMed] [Google Scholar]
  14. Ohno T., Aoyagi M., Yamanashi Y., Saito H., Ikawa S., Meshi T., Okada Y. Nucleotide sequence of the tobacco mosaic virus (tomato strain) genome and comparison with the common strain genome. J Biochem. 1984 Dec;96(6):1915–1923. doi: 10.1093/oxfordjournals.jbchem.a135026. [DOI] [PubMed] [Google Scholar]
  15. Otsuki Y., Shimomura T., Takebe I. Tobacco mosaic virus multiplication and expression of the N gene in necrotic responding tobacco varieties. Virology. 1972 Oct;50(1):45–50. doi: 10.1016/0042-6822(72)90344-3. [DOI] [PubMed] [Google Scholar]
  16. Putney S. D., Benkovic S. J., Schimmel P. R. A DNA fragment with an alpha-phosphorothioate nucleotide at one end is asymmetrically blocked from digestion by exonuclease III and can be replicated in vivo. Proc Natl Acad Sci U S A. 1981 Dec;78(12):7350–7354. doi: 10.1073/pnas.78.12.7350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Saito T., Meshi T., Takamatsu N., Okada Y. Coat protein gene sequence of tobacco mosaic virus encodes a host response determinant. Proc Natl Acad Sci U S A. 1987 Sep;84(17):6074–6077. doi: 10.1073/pnas.84.17.6074. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Solis I., Garcia-Arenal F. The complete nucleotide sequence of the genomic RNA of the tobamovirus tobacco mild green mosaic virus. Virology. 1990 Aug;177(2):553–558. doi: 10.1016/0042-6822(90)90520-2. [DOI] [PubMed] [Google Scholar]
  19. Takamatsu N., Ishikawa M., Meshi T., Okada Y. Expression of bacterial chloramphenicol acetyltransferase gene in tobacco plants mediated by TMV-RNA. EMBO J. 1987 Feb;6(2):307–311. doi: 10.1002/j.1460-2075.1987.tb04755.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Ugaki M., Tomiyama M., Kakutani T., Hidaka S., Kiguchi T., Nagata R., Sato T., Motoyoshi F., Nishiguchi M. The complete nucleotide sequence of cucumber green mottle mosaic virus (SH strain) genomic RNA. J Gen Virol. 1991 Jul;72(Pt 7):1487–1495. doi: 10.1099/0022-1317-72-7-1487. [DOI] [PubMed] [Google Scholar]

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