1887

Abstract

Transgenic tobacco plants expressing an altered form of the 2a replicase gene from the Fny strain of (CMV) exhibit suppressed virus replication and restricted virus movement when inoculated mechanically or by aphid vectors. Additional transformants have been generated which contain replicase gene constructs designed to determine the role(s) of transgene mRNA and/or protein in resistance. Resistance to systemic disease caused by CMV, as well as delayed infection, was observed in several lines of transgenic plants which were capable of expressing either full-length or truncated replicase proteins. In contrast, among plants which contained nontranslatable transgene constructs, only one of 61 lines examined exhibited delays or resistance. Once infected, plants never recovered, regardless of transgene translatability. Transgenic plants exhibiting a range of resistance levels were examined for transgene copy number, mRNA and protein levels. Although ribonuclease protection assays demonstrated that transgene mRNA levels were very low, resistant lines had consistently more steady-state transgene mRNA than susceptible lines. Furthermore, chlorotic or necrotic local lesions developed on the inoculated leaves of transgenic lines containing translatable transgenes, but not on inoculated leaves of lines containing nontranslatable transgenes. These results demonstrate that translatability of the transgene and possibly expression of the transgene protein itself facilitates replicase-mediated resistance to CMV in tobacco.

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2000-03-01
2019-10-20
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References

  1. Anderson, J. M., Palukaitis, P. & Zaitlin, M. ( 1992; ). A defective replicase gene induces resistance to cucumber mosaic virus in transgenic tobacco plants. Proceedings of the National Academy of Sciences, USA 89, 8759-8763.[CrossRef]
    [Google Scholar]
  2. Argos, P., Kamer, G., Nicklin, M. J. H. & Wimmer, E. ( 1984; ). Similarity in gene organization and homology between proteins of animal picornaviruses and a plant comovirus suggest common ancestry of these virus families. Nucleic Acids Research 12, 7251-7267.[CrossRef]
    [Google Scholar]
  3. Baulcombe, D. ( 1996; ). Mechanisms of pathogen-derived resistance to viruses in transgenic plants. Plant Cell 8, 1833-1844.[CrossRef]
    [Google Scholar]
  4. Brederode, F. T., Taschner, P. E. M., Posthumus, E. & Bol, J. F. ( 1995; ). Replicase-mediated resistance to alfalfa mosaic virus. Virology 207, 467-474.[CrossRef]
    [Google Scholar]
  5. Bruenn, J. A. ( 1991; ). Relationships among the positive strand and double-strand RNA viruses as viewed through their RNA-dependent RNA polymerases. Nucleic Acids Research 19, 217-226.[CrossRef]
    [Google Scholar]
  6. Canto, T. & Palukaitis, P. ( 1999; ). Replicase-mediated resistance to cucumber mosaic virus does not inhibit localization and/or trafficking of the viral movement protein. Molecular Plant–Microbe Interactions 12, 743-747.[CrossRef]
    [Google Scholar]
  7. Carr, J. P., Marsh, L. E., Lomonossoff, G. P., Sekiya, M. E. & Zaitlin, M. ( 1992; ). Resistance to tobacco mosaic virus induced by the 54-kDa gene sequence requires expression of the 54-kDa protein. Molecular Plant–Microbe Interactions 5, 397-404.[CrossRef]
    [Google Scholar]
  8. Carr, J. P., Gal-On, A., Palukaitis, P. & Zaitlin, M. ( 1994; ). Replicase-mediated resistance to cucumber mosaic virus in transgenic plants involves suppression of both virus replication in the inoculated leaves and long-distance movement. Virology 199, 439-447.[CrossRef]
    [Google Scholar]
  9. Cassidy, B. G. & Nelson, R. S. ( 1995; ). Differences in protection phenotypes in tobacco plants expressing coat protein genes from peanut stripe potyvirus with or without an engineered ATG. Molecular Plant–Microbe Interactions 8, 357-365.[CrossRef]
    [Google Scholar]
  10. Flavell, R. B. ( 1994; ). Inactivation of gene expression in plants as a consequence of specific sequence duplication. Proceedings of the National Academy of Sciences, USA 91, 3490-3496.[CrossRef]
    [Google Scholar]
  11. Fulton, T. M., Chunwongse, J. & Tanksley, S. D. ( 1995; ). Microprep protocol for extraction of DNA from tomato and other herbaceous plants. Plant Molecular Biology Reporter 13, 207-209.[CrossRef]
    [Google Scholar]
  12. Gal-On, A., Wolf, D., Wang, Y., Faure, J.-E., Pilowsky, M. & Zelcer, A. ( 1998; ). Transgenic resistance to cucumber mosaic virus in tomato: blocking of long-distance movement of the virus in lines harboring a defective viral replicase gene. Phytopathology 88, 1101-1107.[CrossRef]
    [Google Scholar]
  13. Hellwald, K.-H. & Palukaitis, P. ( 1995; ). Viral RNA as a potential target for two independent mechanisms of replicase-mediated resistance against cucumber mosaic virus. Cell 83, 937-946.[CrossRef]
    [Google Scholar]
  14. Lindbo, J. A., Silva-Rosales, L., Proebsting, W. M. & Dougherty, W. G. ( 1993; ). Induction of a highly specific antiviral state in transgenic plants: implications for regulation of gene expression and virus resistance. Plant Cell 5, 1749-1759.[CrossRef]
    [Google Scholar]
  15. MacFarlane, S. A. & Davies, J. W. ( 1992; ). Plants transformed with a region of the 201-kilodalton replicase gene from pea early browning virus RNA 1 are resistant to virus infection. Proceedings of the National Academy of Sciences, USA 89, 5829-5833.[CrossRef]
    [Google Scholar]
  16. Marano, M. R. & Baulcombe, D. ( 1998; ). Pathogen-derived resistance targeted against the negative-strand RNA of tobacco mosaic virus: RNA strand-specific gene silencing? Plant Journal 13, 537-546.[CrossRef]
    [Google Scholar]
  17. Mueller, E., Gilbert, J., Davenport, G., Brigneti, G. & Baulcombe, D. C. ( 1995; ). Homology-dependent resistance: transgenic virus resistance in plants related to homology-dependent gene silencing. Plant Journal 7, 1001-1013.[CrossRef]
    [Google Scholar]
  18. Nguyen, L., Lucas, W. J., Ding, B. & Zaitlin, M. ( 1996; ). Replicase-mediated resistance in transgenic tobacco plants involves inhibition of viral cell-to-cell movement. Proceedings of the National Academy of Sciences, USA 93, 12643-12647.[CrossRef]
    [Google Scholar]
  19. Palukaitis, P. & Zaitlin, M. ( 1997; ). Replicase-mediated resistance to plant virus disease. Advances in Virus Research 48, 349-377.
    [Google Scholar]
  20. Pang, S.-Z., Bock, J. H., Gonsalves, C., Slightom, J. L. & Gonsalves, D. ( 1994; ). Resistance of transgenic Nicotiana benthamiana plants to tomato spotted wilt and impatiens necrotic spot tospoviruses: evidence of involvement of the N protein and N gene RNA in resistance. Phytopathology 84, 243-249.[CrossRef]
    [Google Scholar]
  21. Roberts, B. E. & Paterson, B. M. ( 1973; ). Efficient translation of TMV RNA and rabbit globin 9S RNA in a cell-free system from commercial wheat germ. Proceedings of the National Academy of Sciences, USA 70, 2330-2339.[CrossRef]
    [Google Scholar]
  22. Rodgers, S. G., Horsch, R. B. & Fraley, R. T. ( 1986; ). Gene transfer in plants: production of transformed plants using Ti plasmid vectors. Methods in Enzymology 118, 627-640.
    [Google Scholar]
  23. Roossinck, M. J. & Palukaitis, P. ( 1990; ). Rapid induction and severity of symptoms in zucchini squash (Cucurbita pepo) map to RNA 1 of cucumber mosaic virus. Molecular Plant–Microbe Interactions 3, 188-192.[CrossRef]
    [Google Scholar]
  24. Sambrook, J., Fritsch, E. F. & Maniatis, T. (1989). Molecular Cloning: A Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
  25. Sleat, D. E., Gallie, D. R., Watts, J. W., Deom, C. M., Turner, P. C., Beachy, R. N. & Wilson, T. M. A. ( 1988; ). Selective recovery of foreign gene transcripts as virus-like particles in TMV-infected transgenic tobaccos. Nucleic Acids Research 16, 3127-3140.[CrossRef]
    [Google Scholar]
  26. Smith, H. A., Swaney, S. L., Parks, T. D., Wernsman, E. A. & Dougherty, W. G. ( 1994; ). Transgenic plant virus resistance mediated by untranslatable sense RNAs: expression, regulation, and fate of nonessential RNAs. Plant Cell 6, 1441-1453.[CrossRef]
    [Google Scholar]
  27. Tanzer, M. M., Thompson, W. F., Law, M. D., Wernsman, E. A. & Uknes, S. ( 1997; ). Characterization of post-transcriptionally suppressed transgene expression that confers resistance to tobacco etch virus infection in tobacco. Plant Cell 9, 1411-1423.[CrossRef]
    [Google Scholar]
  28. Tenllado, F., Garcia-Luque, I., Serra, M. T. & Diaz-Ruiz, J. R. ( 1995; ). Nicotiana benthamiana plants transformed with the 54-kDa region of the pepper mild mottle tobamovirus replicase gene exhibit two types of resistance responses against viral infection. Virology 211, 170-183.[CrossRef]
    [Google Scholar]
  29. Tenllado, F., Garcia-Luque, I., Serra, M. T. & Diaz-Ruiz, J. R. ( 1996; ). Resistance to pepper mild mottle tobamovirus conferred by the 54-kDa gene sequence in transgenic plants does not require expression of the wild-type 54-kDa protein. Virology 219, 330-335.[CrossRef]
    [Google Scholar]
  30. Wintermantel, W. M., Banerjee, N., Oliver, J. C., Paolillo, D. J. & Zaitlin, M. ( 1997; ). Cucumber mosaic virus is restricted from entering minor veins in transgenic tobacco exhibiting replicase-mediated resistance. Virology 231, 248-257.[CrossRef]
    [Google Scholar]
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