Submitted to: Molecular Plant-Microbe Interactions
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/26/1995
Publication Date: N/A
Citation: Interpretive Summary: The complexity of molecular interactions between a virus and its plant hosts has been, and continues to be, a significant barrier to the development of better control measures. Yet we really know very little about how viruses cause disease and the mechanisms which determine a virus' ability to infect a given host. This research was designed to determine which parts of a particular viral gene (the alpha-a gene of barley stripe mosaic virus) are responsible for the ability of one viral strain to infect oat, while another viral strain cannot. We identified the specific amino acids (the basic building blocks of proteins) encoded by this gene which determine BSMV pathogenicity to oat. Furthermore, we found that changing a single amino acid in the alpha-a protein was all that was required to confer minimal pathogenicity to a BSMV strain which otherwise would not be pathogenic to oat. Other amino acid changes influenced the rate of virus accumulation within plants and the efficiency of virus transmission. These results will help us to better understand host-virus interactions and will provide valuable clues for the future design of molecular approaches to disease control.
Technical Abstract: Cloned DNA of the BSMV RNA genomes of strains ND18 and CV42 was used to examine the molecular aspects of virus host range. Previously, we showed that a 236 nucleotide region from the alpha-a gene of strain CV42, when substituted for the homologous region in strain ND18, conferred a near wild type oat pathogenicity to this strain. Six amino acid substitutions were found in this region of the alpha-a gene. In the present work, codons specifying T-724, I-764, and N-785 (encoded by CV42 RNA alpha) were substituted individually and in combination for those coding for P-724, T-764, and K-785 (encoded by ND18 RNA alpha) by site-directed mutagenesis. The core K-733, T-734, and K-736 positions (CV42) were substituted for Q-733, S-734, and Q-736 (ND18) as a single block. Transcripts generated from these clones were mixed with ND18 beta and gamma transcripts and were inoculated to oat protoplasts and plants. Inoculated plants were analyzed for symptom type and the accumulation of virus antigen, and extracts from inoculated protoplasts were subjected to Northern and western blot analysis. The results indicate that the C-2261 to A-2261 nucleotide substitution (P-724 to T-724) by itself is sufficient to enable strain ND18 to infect oat plants, although poorly. Additional substitution of CV42 codons into ND18 RNA alpha at the remaining five positions altered symptom type, decreased the timing of the appearance of symptoms, and increased the percentage of plants infected per inoculation. Nevertheless, all mutants accumulated to similar levels in inoculated oat protoplasts after a 24 hr period. Additional evidence suggested that strain ND18 and the non-pathogenic variants generated in the study are either unable to move cell-to-cell or unable to replicate efficiently in intact oat plant leaves.