Submitted to: Journal of General Virology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: April 25, 2006
Publication Date: September 1, 2006
Citation: Stenger, D.C., Young, B.A., French, R.C. 2006. Random mutagenesis of wheat streak mosaic virus HC-Pro: noninfectious interfering mutations in a gene dispensable for systemic infection of plants. Journal of General Virology 87: 2741-2747. Interpretive Summary: The HC-Pro gene of wheat streak mosaic virus (WSMV) may be completely deleted without affecting the ability of the virus to systemically infect plants. However, 11 of 35 point mutations in HC-Pro rendered WSMV unable to systemically infect plants. Some systemic infection deficient mutants remained able to initiate primary infection foci on inoculated wheat leaves but were unable to spread to other parts of the plant. Other noninfectious mutants were unable to establish primary infection foci on inoculated leaves. These results indicated that some HC-Pro mutants interfered with long distance movement of virus within plants and/or blocked replication of the virus within wheat cells. Because absence of HC-Pro did not affect WSMV pathogenicity on wheat plants, these systemic infection deficient mutants most likely acted via mutant interference.
Technical Abstract: Mutations within the HC-Pro coding region of wheat streak mosaic virus (WSMV) were introduced by misincorporation during PCR and evaluated for phenotype within the context of an infectious clone. Nine synonymous substitutions and 15 of 24 nonsynonymous substitutions had no phenotypic effect. Four nonsynonymous substitutions, including one that consistently reverted to wild type, resulted in attenuated systemic infection. Six nonsynonymous substitutions and one nonsense substitution abolished systemic infectivity. Mutants bearing the GUS reporter gene were evaluated for the ability to establish primary infection foci. All attenuated mutants and two systemic infection deficient mutants produced localized regions of GUS expression. In vitro assays revealed that mutants able to establish infection foci retained HC-Pro proteinase activity. Among mutants unable to establish infection foci, HC-Pro proteinase activity was retained, reduced, or absent. As a complete HC-Pro deletion mutant can systemically infect plants, certain substitutions in this dispensable gene likely prevented infection via interference.