Increase Tomato Disease Resistance by Viral Vector- Mediated Expression of Tomato Defense Genes

Using transgenic approaches we have shown that overexpression of the Arabidopsis ELP4/SNT2 gene in tomato increases resistance to the bacterial pathogen Pseudomonas syringae pv. tomato, which causes bacterial speck disease in tomato. We have also shown that the tomato ELP4 gene is biologically functional. While we are planning field trials for the resistant transgenic lines, we analyzed P. syringae pv. tomato-induced transcriptome changes in the most resistant line 61–5. Results show that a group of genes that have previously been shown to be associated with disease resistance were induced to higher levels in the transgenic plants than in the control. The induction of several genes was further confirmed by qPCR analysis. Moreover, in order to avoid genetically modified organism (GMO) issues, we attempted to use the viral overexpression vector pTRV2e to deliver ELP4 into tomato. To this end, we cloned the tomato ELP4 gene into pTRV2e. The resulting plasmid pTRV2e-ELP4 and the control plasmid pTRVe-GFP were transformed into Agrobacterium, which was then used to infiltrate tomato plants. Although multiple experiments under different conditions were conducted, we did not detect any expression of either the ELP4 gene or the control GFP protein, indicating that the viral vector did not work in the experiments. Currently, instead of using the viral vector, we are establishing the state-of-theart CRISPR/Cas9 gene editing technology in tomato, which will likely better avoid GMO issues than any other approaches. INTRODUCTION Disease is one of the major management problems in tomato production areas including Florida. Severe diseases not only lead to direct yield losses but also in many cases result in unmarketable fresh fruit. In Florida, bacterial, fungal, and viral pathogens can all cause serious diseases in tomato. Unfortunately, no major commercial varieties have high levels of resistance to multiple pathogens. As a result, production of tomato heavily relies on application of fungicides and bactericides. However, it has been well documented that application of fungicides and bactericides in agriculture causes significant environmental pollution and raises public concerns on chemical residues. Thus, production and/or activation of broad-spectrum disease resistance using approaches other than fungicides and bactericides are desired in managing tomato diseases. ELP4/SNT2 encodes a subunit of the six-subunit protein complex called Elongator, which functions in multiple biological processes including plant defense responses (Ding and Mou, 2015; Woloszynska et al., 2016). Loss-of-function mutations in the Elongator complex make the plants highly susceptible to both bacterial and fungal pathogens (DeFraia et al., 2010; 2013; Wang et al., 2013; Wang et al., 2015; An et al., 2016), demonstrating an important function for Elongator in plant disease resistance. We have recently shown that overexpression of ELP4 increases basal expression of a number of defense genes and enhances resistance to the bacterial pathogen P. syringae pv. maculicola (Psm) ES4326 in Arabidopsis (Ding et al., unpublished data). We also found that overexpression of the Arabidopsis ELP3 and ELP4 genes in strawberry increases resistance to multiple pathogens including Colletotrichum gloeosporioides (causing anthracnose), Podosphaera aphanis (causing powdery mildew), and Xanthomonas fragariae (causing angular leaf spot) (Silva et al., unpublished data). Moreover, overexpression of ELP3 or ELP4 does not drastically affect plant growth and development. We further generated transgenic tomato plants expressing the Arabidopsis ELP3 and ELP4 genes and tested their disease resistance. Results show that several transgenic tomato lines expressing the ELP4 gene are more resistant than the control plants to P. syringae pv. tomato. We also demonstrated that the tomato ELP3 and ELP4 genes both encode a functional protein. Here, we compared P. syringae pv. tomato-induced transcriptome changes in the most resistant transgenic line 61–5 and the control plants