Identification of the putative RNA polymerase of Cryphonectria hypovirus in a solubilized replication complex
1994
Fahima, T. | Wu, Y. | Zhang, L. | Van Alfen, N.K.
Hypovirulence of the pathogenic fungus Cryphonectria parasitica, caused by the unencapsulated viral double-stranded RNA of Cryphonectria hypovirus (CHV1), provides a means for biological control of chestnut blight. We report here the isolation of a replication complex of the virus solubilized from host membranes. The conserved regions of the putative RNA polymerase encoded by strain CHV1-713 were cloned and expressed, and the recombinant protein was purified and used to produce polyclonal antibodies. The CHV1 replication complex was solubilized from a membrane fraction of CHV1-infected C. parasitica hyphae. Antibodies raised against the putative viral polymerase reacted on a Western immunoblot with an 87-kDa polypeptide of the replication complex but not with comparable preparations from an isogeneic uninfected strain. Analysis of the polypeptide composition of the complex by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining revealed a number of other polypeptides along with the double=stranded RNA of the virus. We conclude that this 87-kDa polypeptide is the putative RNA polymerase encoded on open reading frame B of CHV1.
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