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Molecular Methods for Studying the Cryphonectria parasitica – Hypovirus Experimental System

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Fungal Genomics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 722))

Abstract

The interaction of the filamentous fungal plant pathogen Cryphonectria parasitica with its virulence-attenuating viruses provides a unique platform to explore the molecular biology and genetics of virus–host interactions. Following the development of transformation procedures for this fungus, subsequent advances include infectious cDNA clones of several members of the Hypoviridae and an imminently complete fungal genome project. Presented here are basic protocols for growth of the organism and the extraction of DNA, RNA, and protein. Additionally, two further protocols are provided for investigations of host protein phosphorylation and for viral genome secondary structure.

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References

  1. Merkel, H. W. (1906) A deadly fungus on the American chestnut., NY Zool. Soc. Annu. Rep. 10, 97–103.

    Google Scholar 

  2. Biraghi, A. (1946) Il cancro del castagno causato da Endothia parasitica., Ital. Agric. 7, 1–9.

    Google Scholar 

  3. Biraghi, A. (1953) Possible active resistance to Endothia parasitica in Castenea sativa., in 11th Rep. Congr. Int. Union For. Res. Org., pp 149–157, Rome.

    Google Scholar 

  4. Grente, J. (1965) Les formes hypovirulentes d’Endothia parasitica et les espoires de lutte contre le chancre du chataigner., C.R. Seances Acad. Agric. Fr. 51, 1033–1037.

    Google Scholar 

  5. Day, P. R., Dodds, J. A., Elliston, J. E., Jaynes, R. A., and Anagnostakis, S. L. (1977) Double-Stranded RNA in Endothia parasitica [from Europe and North America, Biological Control, Chestnut Blight], Phytopathology 67, 1393–1396.

    Article  Google Scholar 

  6. Anagnostakis, S. L., and Day, P. R. (1979) Hypovirulence conversion in Endothia parasitica., Phytopathology 69, 1226–1229.

    Article  CAS  Google Scholar 

  7. Hillman, B. I., Fulbright, D. W., Nuss, D. L., and Van Alfen, N. K. (1995) Hypoviridae, in Virus taxonomy (F.A. Murphy, C. M. F., D.H.L. Bishop, S.A. Ghabrial, A.W. Jarvis, G.P. Martelli, M.A. Mayo, and M.D. Summers (ed.), Ed.), pp 261–264, Springer-Verlag, New York, N.Y.

    Google Scholar 

  8. Dawe, A. L., and Nuss, D. L. (2001) Hypoviruses and chestnut blight: exploiting viruses to understand and modulate fungal pathogenesis, Annu Rev Genet 35, 1–29.

    Article  PubMed  CAS  Google Scholar 

  9. Anagnostakis, S. L. (1982) Biological control of chestnut blight, Science 215, 466–471.

    Article  PubMed  CAS  Google Scholar 

  10. Shapira, R., Choi, G. H., and Nuss, D. L. (1991) Virus-like genetic organization and expression strategy for a double- stranded RNA genetic element associated with biological control of chestnut blight, EMBO J. 10, 731–739.

    PubMed  CAS  Google Scholar 

  11. Choi, G. H., Pawlyk, D. M., and Nuss, D. L. (1991) The autocatalytic protease p29 encoded by a hypovirulence-associated virus of the chestnut blight fungus resembles the potyvirus-encoded protease HC-Pro, Virology 183, 747–752.

    Article  PubMed  CAS  Google Scholar 

  12. Koonin, E. V., Choi, G. H., Nuss, D. L., Shapira, R., and Carrington, J. C. (1991) Evidence for common ancestry of a chestnut blight hypovirulence- associated double-stranded RNA and a group of positive-strand RNA plant viruses, Proc. Natl. Acad. Sci. U.S.A 88, 10647–10651.

    Article  PubMed  CAS  Google Scholar 

  13. Churchill, A. C. L., Ciufetti, L. M., Hansen, D. R., Van Etten, H. D., and Van Alfen, N. K. (1990) Transformation of the fungal pathogen Cryphonectria parasitica with a variety of heterologous plasmids., Curr Genet 17, 25–31.

    Article  CAS  Google Scholar 

  14. Choi, G. H., and Nuss, D. L. (1992) Hypovirulence of chestnut blight fungus conferred by an infectious viral cDNA, Science 257, 800–803.

    Article  PubMed  CAS  Google Scholar 

  15. Chen, B., and Nuss, D. L. (1999) Infectious cDNA clone of hypovirus CHV1-Euro7: a comparative virology approach to investigate virus-mediated hypovirulence of the chestnut blight fungus Cryphonectria parasitica, J. Virol. 73, 985–992.

    PubMed  CAS  Google Scholar 

  16. Pagano, M. A., Meggio, F., Ruzzene, M., Andrzejewska, M., Kazimierczuk, Z., and Pinna, L. A. (2004) 2-Dimethylamino-4,5,6,7-tetrabromo-1H-benzimidazole: a novel powerful and selective inhibitor of protein kinase CK2, Biochem Biophys Res Commun 321, 1040–1044.

    Article  PubMed  CAS  Google Scholar 

  17. Romero, T. A., Tumban, E., Jun, J., Lott, W. B., and Hanley, K. A. (2006) Secondary structure of dengue virus type 4 3′ untranslated region: impact of deletion and substitution mutations, J Gen Virol 87, 3291–3296.

    Article  PubMed  CAS  Google Scholar 

  18. Van Den Born, E., Gultyaev, A. P., and Snijder, E. J. (2004) Secondary structure and function of the 5′-proximal region of the equine arteritis virus RNA genome, RNA 10, 424–437.

    Article  Google Scholar 

  19. Zuker, M. (2003) Mfold web server for nucleic acid folding and hybridization prediction, Nucleic Acids Res 31, 3406–3415.

    Article  PubMed  CAS  Google Scholar 

  20. Hillman, B. I., Shapira, R., and Nuss, D. L. (1990) Hypovirulence-associated suppression of host functions in Cryphonectria parasitica can be partially relieved by high light intensity., Phytopathology 80, 950–956.

    Article  Google Scholar 

  21. Sambrook, J., Fritsch, E. F., and Maniatis, T. (1989) Molecular cloning: a laboratory manual, 2nd ed., Cold Spring Harbor Press, Cold Spring Harbor, NY.

    Google Scholar 

  22. Parsley, T. B., Segers, G. C., Nuss, D. L., and Dawe, A. L. (2003) Analysis of altered G-protein subunit accumulation in Cryphonectria parasitica reveals a third Gα homologue, Curr Genet 43, 24–33.

    PubMed  CAS  Google Scholar 

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Acknowledgments

This work was supported by NSF award MCB-0718735 (to ALD). GR was supported by a fellowship from the NMSU-NIH MBRS-RISE program (R25GM061222).

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Correspondence to Angus L. Dawe .

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Dawe, A.L., Mu, R., Rivera, G., Salamon, J.A. (2011). Molecular Methods for Studying the Cryphonectria parasitica – Hypovirus Experimental System. In: Xu, JR., Bluhm, B. (eds) Fungal Genomics. Methods in Molecular Biology, vol 722. Humana Press. https://doi.org/10.1007/978-1-61779-040-9_17

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  • DOI: https://doi.org/10.1007/978-1-61779-040-9_17

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  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-039-3

  • Online ISBN: 978-1-61779-040-9

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