1887

Abstract

Transgenic potato plants were generated which express a dicistronic mRNA corresponding to the 8 kDa protein and coat protein (CP) genes of potato virus X (PVX). Northern blot analysis of RNA isolated from these plants indicated that both the 8 kDa protein and the CP are translated from this dicistronic mRNA. Expression of both of these proteins in transgenic plants was demonstrated by Western blot analysis, and suggests that translation of CP takes place either by initiation by internal entry of ribosomes or by a termination/reinitiation mechanism. CP was detected in protoplasts electroporated with RNA transcripts of the dicistronic construct in the presence or absence of a stable hairpin structure at the 5′ terminus, suggesting that the former model is more likely. Similar results were obtained when a DNA fragment containing the PVX CP leader sequence was placed between two reporter genes in the transient assay system. These results suggest that potexvirus CP expression may take place from the genomic RNA as well as from the subgenomic RNA.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/0022-1317-78-11-3051
1997-11-01
2024-05-05
Loading full text...

Full text loading...

/deliver/fulltext/jgv/78/11/9367393.html?itemId=/content/journal/jgv/10.1099/0022-1317-78-11-3051&mimeType=html&fmt=ahah

References

  1. Basso J., Dallaire P., Charest P. J., Devantier Y., Laliberte J.-F. 1994; Evidence for an internal ribosome entry site within the 5ʹ non- translated region of turnip mosaic potyvirus RNA. Journal of General Virology 75:1–7
    [Google Scholar]
  2. Bendena W. G., Mackie G. A. 1986; Translational strategies in potexviruses: products encoded by clover yellow mosaic virus, foxtail mosaic virus and viola mottle mosaic virus RNAs in vitro. Virology 153:220–229
    [Google Scholar]
  3. Bendena W. G., AbouHaidar M., Mackie G. A. 1985; Synthesis in vitro of the coat protein of papaya mosaic virus. Virology 140:257–268
    [Google Scholar]
  4. Bendena W. G., Bancroft J. B., Mackie G. A. 1987; Molecular cloning of clover yellow mosaic virus RNA: identification of coat protein coding sequences in vivo and in vitro. Virology 157:276–284
    [Google Scholar]
  5. Borman A., Jackson R. J. 1992; Initiation of translation of human rhinovirus RNA: mapping the internal ribosome entry site. Virology 188:685–696
    [Google Scholar]
  6. Brown E. A., Day P. S., Jansen R. W., Lenon S. M. 1991; The 5′ -nontranslated region of hepatitis A virus RNA: secondary structure and elements required for translation in vitro. Journal of Virology 65:5828–5838
    [Google Scholar]
  7. Carrington J. C., Freed D. D. 1990; Cap-independent enhancement of translation by a plant potyvirus 5′ nontranslated region. Journal of Virology 64:1590–1597
    [Google Scholar]
  8. Dolja V. V., Glamu D. P., Morozov S. Yu., Atabekov J. G. 1987; Potato virus X-related single and double stranded RNAs: characterization and identification of terminal structure. FEBS Letters 241:398–412
    [Google Scholar]
  9. Feinberg A. P., Vogelstein B. 1983; A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Analytical Biochemistry 132:6–13
    [Google Scholar]
  10. Futterer J., Kiss-Lazlo Z., Hohn T. 1993; Nonlinear ribosome migration on cauliflower mosaic virus 35S RNA. Cell 73:789–802
    [Google Scholar]
  11. Glass M. J., Jia X.-Y., Summers D. F. 1993; Identifcation of the hepatitis A internal ribosome entry site: in vivo and in vitro analysis of bicistronic RNAs containing the HAV 5ʹ noncoding region. Virology 193:842–852
    [Google Scholar]
  12. Gorman C. M., Moffat L. F., Howard B. H. 1982; Recombinant genomes which express chloramphenicol acetyltransferase in mammalian cells. Molecular and Cellular Biology 2:1044–1051
    [Google Scholar]
  13. Gubler U., Hoffman B. J. 1983; A simple and very efficient method for generating cDNA libraries. Gene 25:263–269
    [Google Scholar]
  14. Hemenway C., Fang R., Kaniewski W., Chua N., Turner N. 1988; Analysis of the mechanism of protection in transgenic plants expressing the potato virus X coat protein or its antisense RNA. EMBO Journal 7:1273–1280
    [Google Scholar]
  15. Herman R. 1986; Internal initiation of translation on the vesicular stomatitis virus phosphoprotein mRNA yields a second protein. Journal of Virology 58:794–804
    [Google Scholar]
  16. Horsh R. B., Fry J. E., Hoffman N. L., Eichholtz D., Rogers S. G., Fraley R. T. 1985; A simple and general method for transferring genes into plants. Science 227:1229–1231
    [Google Scholar]
  17. Huisman M. J., Linthorst H. J. M., Bol J. F., Cornelissen B. J. C. 1988; The complete nucleotide sequence of potato virus X and its homologies at the amino acid level with various plus-stranded RNA viruses. Journal of General Virology 69:1789–1798
    [Google Scholar]
  18. Ivanov I., Tom M., Wishart P., May E. 1987; Chemical synthesis and expression of the human calcitonin gene. Gene 59:223–230
    [Google Scholar]
  19. Jang S. K., Wimmer E. 1990; Cap-independent translation of encephalomyocarditis virus RNA: structural elements of the internal ribosomal entry site and involvement of a cellular 5ʹ-kD RNA-binding protein. Genes & Development 4:1560–1572
    [Google Scholar]
  20. Jang S. K., Pestova T. V., Hellen C. U. T., Witherell G. W., Wimmer E. 1990; Cap-independent translation of picornavirus RNAs: structure and function of the internal ribosome entry site. Enzyme 44:292–309
    [Google Scholar]
  21. Karasev A. V., Aleksandrova N. M., Miroshnichenko N. A., Nesterova I. V., Dorokhov Y. L. 1988; Translational activity of messenger RNAs from plants infected with potato virus X. Molecular Biology (Moscow) 21:1132–1140
    [Google Scholar]
  22. Kozak M. 1986; Bifunctional messenger RNAs in eukaryotes. Cell 47:481–483
    [Google Scholar]
  23. Kozak M. 1989; The scanning model for translation: an update. Journal of Cell Biology 108:229–241
    [Google Scholar]
  24. Kozak M. 1990; Downstream secondary structure facilitates recognition of initiator codons by eukaryotic ribosomes. Proceedings of the National Academy of Sciences, USA 87:8301–8305
    [Google Scholar]
  25. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685
    [Google Scholar]
  26. Le S.-Y., Chen J.-H., Sonenberg N., Maizel J. V. 1992; Conserved tertiary structure elements in the 5′ untranslated region of human enteroviruses and rhinoviruses. Virology 191:858–866
    [Google Scholar]
  27. Luciano C. S., Rhoads R. E., Shaw J. G. 1987; Synthesis of potyviral RNA and proteins in tobacco mesophyll protoplasts inoculated by electroporation. Plant Science 51:295–303
    [Google Scholar]
  28. Macejak D. G., Sarnow P. 1991; Internal initiation of translation mediated by the 5ʹ leader of a cellular mRNA. Nature 353:90–94
    [Google Scholar]
  29. Mackie G. A., Bancroft J. B. 1986; The longer RNA species in narcissus mosaic virus encodes all viral functions. Virology 153:215–219
    [Google Scholar]
  30. Meerovitch K., Svitkin Y. V., Lee H. S., Lejebkowicz F., Kenan D. J., Chan K. L., Agol V. I., Keene J. D., Sonenberg N. 1993; La autoantigen enhances and corrects aberrant translation of poliovirus RNA in reticulocyte lysate. Journal of Virology 67:3798–3807
    [Google Scholar]
  31. Merrick W. C. 1992; Mechanism and regulation of eukaryotic protein synthesis. Microbiological Reviews 56:291–315
    [Google Scholar]
  32. Miller J. H. 1972 Experiments in Molecular Genetics pp. 352–355 Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  33. Morozov S. Y., Miroshnichenko N. A., Zelenina D. A., Fedorkin O. N., Solovijev A. G., Lukasheva L. I., Atabekov J. C. 1990; Expression of RNA transcripts of potato virus X full-length and subgenomic cDNAs. Biochimie 72:677–684
    [Google Scholar]
  34. Morozov S. Y., Miroshnichenko N. A., Solovyev A. G., Fedorkin O. N., Zelenina D. A., Lukasheva L. I., Karasev A. V., Dolja V. V., Atabekov J. G. 1991; Expression strategy of the potato virus X triple gene block. Journal of General Virology 72:2039–2042
    [Google Scholar]
  35. Oh S. K., Scott M. P., Sarnow P. 1992; Homeotic gene anten- napedia messenger RNA contains 5ʹ-noncoding sequences that confer translational initiation by internal ribosome binding. Genes & Development 1:1643–1653
    [Google Scholar]
  36. Otsuki Y., Takebe I. 1972; Ultrastructure of infection of tobacco mesophyll protoplasts by tobacco mosaic virus. Virology 49:188–194
    [Google Scholar]
  37. Pelletier J., Sonenberg N. 1985; Internal initiation of translation of eukaryotic mRNA directed by a sequence derived from poliovirus RNA. Nature 334:320–322
    [Google Scholar]
  38. Pelletier J., Kaplan G., Racaniello V. R., Sonenberg N. 1988; Translational efficiency of poliovirus mRNA: mapping inhibitory cis- acting elements within the 5ʹ noncoding region. Journal of Virology 62:2219–2227
    [Google Scholar]
  39. Ricciardi R. P., Goodman R. M., Gottlieb D. 1978; Translation of PVX RNA in vitro by wheat germ. Virology 85:310–314
    [Google Scholar]
  40. Sarnow P. 1995 Cap-Independent Translation Berlin, New York: Springer;
    [Google Scholar]
  41. Short M. N., Davies J. W. 1983; Narcissus mosaic virus : a potexvirus with an encapsidated subgenomic messenger RNA for coat protein. Bioscience Reports 3:837–846
    [Google Scholar]
  42. Solovyev A. G., Novikov V. K., Merits A., Savenkov E. I., Zeleninu D. A., Tyulkina L. G., Morozov S. Y. 1994; Genome characterization and taxonomy of Plantago asiatica mosaic potexvirus. Journal of General Virology 75:259–267
    [Google Scholar]
  43. Sonenberg N., Meerovitch K. 1990; Translation of poliovirus mRNA. Enzyme 44:278–291
    [Google Scholar]
  44. Thomas A. A., Haar E., Wellink J., Voorma H. O. 1991; Cowpea mosaic virus middle component RNA contains a sequence that allows internal binding of ribosomes and that requires eukaryotic initiation factor 4F for optimal translation. Journal of Virology 65:2953–2959
    [Google Scholar]
  45. White K. A., Mackie G. A. 1990; Control and expression of 3′ open reading frames in clover yellow mosaic virus. Virology 179:576–584
    [Google Scholar]
  46. Witherell G. W., Wimmer E. 1994; Encephalomyocarditis virus internal ribosomal entry site RNA-proteininteractions. JournalofVirology 68:3183–3192
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/0022-1317-78-11-3051
Loading
/content/journal/jgv/10.1099/0022-1317-78-11-3051
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error