Abstract
Viral double-stranded RNAs (dsRNAs) responsible for virulence attenuation (hypovirulence) of the chestnut blight fungus, Cryphonectria parasitica, profoundly influence a range of host functions in addition to virulence. The 5'-proximal open reading frame, A, of the prototypical hypovirulence-associated viral dsRNA, L-dsRNA, present in hypovirulent strain EP713, was recently shown by DNA-mediated transformation analysis to suppress fungal sporulation, pigmentation, and accumulation of the enzyme laccase (G. H. Choi and D. L. Nuss, EMBO J. 11:473-477, 1992). We mapped this suppressive activity to the autocatalytic papain-like protease, p29, present within the amino-terminal portion of open reading frame A-encoded polyprotein p69. Mutational analysis revealed that the ability of p29 to alter fungal phenotype is dependent upon release from the polyprotein precursor but is independent of intrinsic proteolytic activity. Deletion of the p29-coding domain within the context of an infectious L-dsRNA cDNA clone resulted in a replication-competent viral dsRNA that exhibited intermediate suppressive activity while retaining the ability to confer hypovirulence. Thus, p29 is necessary but not sufficient for the level of virus-mediated suppression of fungal pigmentation, sporulation, and laccase accumulation observed for wild-type hypovirulent strain EP713 and is nonessential for viral RNA replication and virulence attenuation. These results also illustrate the feasibility of engineering infectious viral cDNA for construction of hypovirulent fungal strains with specific phenotypic traits.
Full text
PDF








Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Anagnostakis S. L. Biological control of chestnut blight. Science. 1982 Jan 29;215(4532):466–471. doi: 10.1126/science.215.4532.466. [DOI] [PubMed] [Google Scholar]
- Bernstein H. D., Sonenberg N., Baltimore D. Poliovirus mutant that does not selectively inhibit host cell protein synthesis. Mol Cell Biol. 1985 Nov;5(11):2913–2923. doi: 10.1128/mcb.5.11.2913. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chen B., Choi G. H., Nuss D. L. Mitotic stability and nuclear inheritance of integrated viral cDNA in engineered hypovirulent strains of the chestnut blight fungus. EMBO J. 1993 Aug;12(8):2991–2998. doi: 10.1002/j.1460-2075.1993.tb05967.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Choi G. H., Larson T. G., Nuss D. L. Molecular analysis of the laccase gene from the chestnut blight fungus and selective suppression of its expression in an isogenic hypovirulent strain. Mol Plant Microbe Interact. 1992 Mar-Apr;5(2):119–128. doi: 10.1094/mpmi-5-119. [DOI] [PubMed] [Google Scholar]
- Choi G. H., Nuss D. L. A viral gene confers hypovirulence-associated traits to the chestnut blight fungus. EMBO J. 1992 Feb;11(2):473–477. doi: 10.1002/j.1460-2075.1992.tb05077.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Choi G. H., Nuss D. L. Hypovirulence of chestnut blight fungus conferred by an infectious viral cDNA. Science. 1992 Aug 7;257(5071):800–803. doi: 10.1126/science.1496400. [DOI] [PubMed] [Google Scholar]
- Choi G. H., Pawlyk D. M., Nuss D. L. The autocatalytic protease p29 encoded by a hypovirulence-associated virus of the chestnut blight fungus resembles the potyvirus-encoded protease HC-Pro. Virology. 1991 Aug;183(2):747–752. doi: 10.1016/0042-6822(91)91004-z. [DOI] [PubMed] [Google Scholar]
- Choi G. H., Shapira R., Nuss D. L. Cotranslational autoproteolysis involved in gene expression from a double-stranded RNA genetic element associated with hypovirulence of the chestnut blight fungus. Proc Natl Acad Sci U S A. 1991 Feb 15;88(4):1167–1171. doi: 10.1073/pnas.88.4.1167. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Clark M. E., Dasgupta A. A transcriptionally active form of TFIIIC is modified in poliovirus-infected HeLa cells. Mol Cell Biol. 1990 Oct;10(10):5106–5113. doi: 10.1128/mcb.10.10.5106. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Clark M. E., Hämmerle T., Wimmer E., Dasgupta A. Poliovirus proteinase 3C converts an active form of transcription factor IIIC to an inactive form: a mechanism for inhibition of host cell polymerase III transcription by poliovirus. EMBO J. 1991 Oct;10(10):2941–2947. doi: 10.1002/j.1460-2075.1991.tb07844.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cullen D., Leong S. A., Wilson L. J., Henner D. J. Transformation of Aspergillus nidulans with the hygromycin-resistance gene, hph. Gene. 1987;57(1):21–26. doi: 10.1016/0378-1119(87)90172-7. [DOI] [PubMed] [Google Scholar]
- Daubert S. D., Schoelz J., Debao L., Shepherd R. J. Expression of disease symptoms in cauliflower mosaic virus genomic hybrids. J Mol Appl Genet. 1984;2(6):537–547. [PubMed] [Google Scholar]
- Devaney M. A., Vakharia V. N., Lloyd R. E., Ehrenfeld E., Grubman M. J. Leader protein of foot-and-mouth disease virus is required for cleavage of the p220 component of the cap-binding protein complex. J Virol. 1988 Nov;62(11):4407–4409. doi: 10.1128/jvi.62.11.4407-4409.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Falk M. M., Grigera P. R., Bergmann I. E., Zibert A., Multhaup G., Beck E. Foot-and-mouth disease virus protease 3C induces specific proteolytic cleavage of host cell histone H3. J Virol. 1990 Feb;64(2):748–756. doi: 10.1128/jvi.64.2.748-756.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hellen C. U., Fäcke M., Kräusslich H. G., Lee C. K., Wimmer E. Characterization of poliovirus 2A proteinase by mutational analysis: residues required for autocatalytic activity are essential for induction of cleavage of eukaryotic initiation factor 4F polypeptide p220. J Virol. 1991 Aug;65(8):4226–4231. doi: 10.1128/jvi.65.8.4226-4231.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hillman B. I., Tian Y., Bedker P. J., Brown M. P. A North American hypovirulent isolate of the chestnut blight fungus with European isolate-related dsRNA. J Gen Virol. 1992 Mar;73(Pt 3):681–686. doi: 10.1099/0022-1317-73-3-681. [DOI] [PubMed] [Google Scholar]
- Kräusslich H. G., Nicklin M. J., Toyoda H., Etchison D., Wimmer E. Poliovirus proteinase 2A induces cleavage of eucaryotic initiation factor 4F polypeptide p220. J Virol. 1987 Sep;61(9):2711–2718. doi: 10.1128/jvi.61.9.2711-2718.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Larson T. G., Choi G. H., Nuss D. L. Regulatory pathways governing modulation of fungal gene expression by a virulence-attenuating mycovirus. EMBO J. 1992 Dec;11(12):4539–4548. doi: 10.1002/j.1460-2075.1992.tb05555.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lloyd R. E., Grubman M. J., Ehrenfeld E. Relationship of p220 cleavage during picornavirus infection to 2A proteinase sequencing. J Virol. 1988 Nov;62(11):4216–4223. doi: 10.1128/jvi.62.11.4216-4223.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nuss D. L. Biological control of chestnut blight: an example of virus-mediated attenuation of fungal pathogenesis. Microbiol Rev. 1992 Dec;56(4):561–576. doi: 10.1128/mr.56.4.561-576.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rigling D., Van Alfen N. K. Regulation of laccase biosynthesis in the plant-pathogenic fungus Cryphonectria parasitica by double-stranded RNA. J Bacteriol. 1991 Dec;173(24):8000–8003. doi: 10.1128/jb.173.24.8000-8003.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Saiki R. K., Gelfand D. H., Stoffel S., Scharf S. J., Higuchi R., Horn G. T., Mullis K. B., Erlich H. A. Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science. 1988 Jan 29;239(4839):487–491. doi: 10.1126/science.2448875. [DOI] [PubMed] [Google Scholar]
- Shapira R., Choi G. H., Hillman B. I., Nuss D. L. The contribution of defective RNAs to the complexity of viral-encoded double-stranded RNA populations present in hypovirulent strains of the chestnut blight fungus Cryphonectria parasitica. EMBO J. 1991 Apr;10(4):741–746. doi: 10.1002/j.1460-2075.1991.tb08005.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Shapira R., Choi G. H., Nuss D. L. Virus-like genetic organization and expression strategy for a double-stranded RNA genetic element associated with biological control of chestnut blight. EMBO J. 1991 Apr;10(4):731–739. doi: 10.1002/j.1460-2075.1991.tb08004.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sun X. H., Baltimore D. Human immunodeficiency virus tat-activated expression of poliovirus protein 2A inhibits mRNA translation. Proc Natl Acad Sci U S A. 1989 Apr;86(7):2143–2146. doi: 10.1073/pnas.86.7.2143. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Van Alfen N. K., Jaynes R. A., Anagnostakis S. L., Day P. R. Chestnut Blight: Biological Control by Transmissible Hypovirulence in Endothia parasitica. Science. 1975 Sep 12;189(4206):890–891. doi: 10.1126/science.189.4206.890. [DOI] [PubMed] [Google Scholar]
- Wickner R. B. Double-stranded and single-stranded RNA viruses of Saccharomyces cerevisiae. Annu Rev Microbiol. 1992;46:347–375. doi: 10.1146/annurev.mi.46.100192.002023. [DOI] [PubMed] [Google Scholar]
- Wickner R. B. Yeast virology. FASEB J. 1989 Sep;3(11):2257–2265. doi: 10.1096/fasebj.3.11.2550303. [DOI] [PubMed] [Google Scholar]
- Yu S. F., Lloyd R. E. Identification of essential amino acid residues in the functional activity of poliovirus 2A protease. Virology. 1991 Jun;182(2):615–625. doi: 10.1016/0042-6822(91)90602-8. [DOI] [PubMed] [Google Scholar]