Abstract
Helper component (HC) is a virus-encoded nonstructural protein that is required for transmission of potyviruses by their aphid vectors. As a prelude to studies on the molecular basis of HC activity, a cDNA clone (pPB-3) was constructed that contained the first three cistrons (34 kDa-HC-42 kDa) of the RNA genome of the potyvirus tobacco vein mottling virus, the first six nucleotides of the adjacent cylindrical inclusion body protein cistron, and a synthetic translation termination codon. This construction was introduced into tobacco cells via a Ti plasmid-based vector. Northern blot analysis of transgenic plants demonstrated the presence of an RNA of the size expected from the construction of pPB-3, and Western blot analysis revealed the presence of a protein that comigrated with authentic HC, indicating that the proteolytic activity necessary to produce mature-sized HC was encoded by pPB-3. The HC produced in the transgenic plants was demonstrated to be active in a virus transmission bioassay with aphids.
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- Carrington J. C., Cary S. M., Parks T. D., Dougherty W. G. A second proteinase encoded by a plant potyvirus genome. EMBO J. 1989 Feb;8(2):365–370. doi: 10.1002/j.1460-2075.1989.tb03386.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
- Domier L. L., Franklin K. M., Hunt A. G., Rhoads R. E., Shaw J. G. Infectious in vitro transcripts from cloned cDNA of a potyvirus, tobacco vein mottling virus. Proc Natl Acad Sci U S A. 1989 May;86(10):3509–3513. doi: 10.1073/pnas.86.10.3509. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Domier L. L., Franklin K. M., Shahabuddin M., Hellmann G. M., Overmeyer J. H., Hiremath S. T., Siaw M. F., Lomonossoff G. P., Shaw J. G., Rhoads R. E. The nucleotide sequence of tobacco vein mottling virus RNA. Nucleic Acids Res. 1986 Jul 11;14(13):5417–5430. doi: 10.1093/nar/14.13.5417. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
- Govier D. A., Kassanis B. A virus-induced component of plant sap needed when aphids acquire potato virus Y from purified preparations. Virology. 1974 Oct;61(2):420–426. doi: 10.1016/0042-6822(74)90278-5. [DOI] [PubMed] [Google Scholar]
- Hanahan D. Studies on transformation of Escherichia coli with plasmids. J Mol Biol. 1983 Jun 5;166(4):557–580. doi: 10.1016/s0022-2836(83)80284-8. [DOI] [PubMed] [Google Scholar]
- Hellmann G. M., Shaw J. G., Rhoads R. E. In vitro analysis of tobacco vein mottling virus NIa cistron: evidence for a virus-encoded protease. Virology. 1988 Apr;163(2):554–562. doi: 10.1016/0042-6822(88)90296-6. [DOI] [PubMed] [Google Scholar]
- Kozak M. Compilation and analysis of sequences upstream from the translational start site in eukaryotic mRNAs. Nucleic Acids Res. 1984 Jan 25;12(2):857–872. doi: 10.1093/nar/12.2.857. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Leary J. J., Brigati D. J., Ward D. C. Rapid and sensitive colorimetric method for visualizing biotin-labeled DNA probes hybridized to DNA or RNA immobilized on nitrocellulose: Bio-blots. Proc Natl Acad Sci U S A. 1983 Jul;80(13):4045–4049. doi: 10.1073/pnas.80.13.4045. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pirone T. P., Thornbury D. W. The involvement of a helper component in nonpersistent transmission of plant viruses by aphids. Microbiol Sci. 1984 Nov;1(8):191–193. [PubMed] [Google Scholar]
- Schardl C. L., Byrd A. D., Benzion G., Altschuler M. A., Hildebrand D. F., Hunt A. G. Design and construction of a versatile system for the expression of foreign genes in plants. Gene. 1987;61(1):1–11. doi: 10.1016/0378-1119(87)90359-3. [DOI] [PubMed] [Google Scholar]
- Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Zambryski P., Joos H., Genetello C., Leemans J., Montagu M. V., Schell J. Ti plasmid vector for the introduction of DNA into plant cells without alteration of their normal regeneration capacity. EMBO J. 1983;2(12):2143–2150. doi: 10.1002/j.1460-2075.1983.tb01715.x. [DOI] [PMC free article] [PubMed] [Google Scholar]




