The study of plum eukaryotic translation initiation factor 4e genes for plum pox virus resistance

Plum pox virus (PPV) causes the most economically-devastating viral disease in Prunus species, including fruit crops and ornamental species. Resistance through screening natural resistant germplasm has long been sought for the control of PPV but few natural resistance genes are available. Viruses encode only a limited number of proteins thus have to recruit host factors for their life cycles. This indicates that manipulating host factors is a potential approach for the development of virus resistance in plants. The present research focused on studying of plum (Prunus domestic L.) eukaryotic translation initiation factor 4E (eIF4E) and its isoform eIF(iso)4E genes for the development of strategies for PPV resistance in its natural host Prunus species. The eIF4E and eIF(iso)4E genes were cloned from plum. The expression profiles of the two genes were shown to be similar in different tissues analyzed by real-time PCR. The plasmids containing hair-pin RNAs of plum eIF4E and eIF(iso)4E were constructed and introduced into plum via Agrobacterium-mediated transformation to silence the endogenous eIF4E and eIF(iso)4E genes. Gene expression analysis in transgenic plum plants expressing the corresponding hairpin RNAs demonstrated that mRNAs of eIF4E and eIF(iso)4E were significantly decreased in comparison with the wild type plant. Two species of siRNAs, the hallmarks of gene silencing, were detected in transgenic plum plants. Transgenic plants were challenged with PPV-D strain and the resistance was evaluated by the presence of PPV symptoms and lack of viral RNA. It was found that 82% of the eIF(iso)4E transgenic plants were resistant to PPV. In contrast, all the wild type and eIF4E transgenic plants were susceptible to PPV. These results indicate that eIF(iso)4E is involved in PPV infection and silencing of eIF(iso)4E gene leads to plants resistant to the virus. These transgenic plants may also be resistant to other viruses which require eIF(iso)4E for their infections. Results from this research provide a superior strategy for engineering PPV resistance, which will contribute to the eradication of PPV in Canada and also to the global research effort for the control of the disease.;Keywords: Plum pox virus (PPV), eukaryotic translation initiation factor 4E (eIF4E), eIF(iso)4E, gene silencing, small RNA, intron-spliced hairpin RNA (ihp-RNA), host factor, recessive resistance, transgenic plant.