Identification and study of promoters induced by Asian soybean rust : application in an artificial cell death system

Lisa Cabre 1
Abstract : Phakopsora pachyrhizi Syd.& P.Syd, the fungus responsible for Asian soybean rust, is the most devastating soybean (Glycine max (L.) Merrill) pathogen. First observed in the 2000s in Latin America, the pathogen has spread throughout the Americas. The control of this pathogen depends on the use of fungicides and strict agricultural practices. The repetitive use of the 3 classes of fungicides and the genome plasticity of the pathogen have led to a decreased efficacy of certain molecules. Although vertical resistance genes have been mapped in the soybean germplasm, most of them are not effective against all Asian soybean rust isolates. A deeper understanding of plant immunity facilitates the development of biotechnological approaches for plant disease control. Artificial cell death was previously developed to control Phytophthora infestans development in potato. The technology was based on a barnase ribonuclease that is highly toxic to the plant cell and that consequently needed to be expressed only in the presence of the pathogen. The lethal expression of barnase was counterbalanced by barstar, a highly specific inhibitor of barnase. We propose to evaluate this technology in soybean to control P. pachyrhizi. The key objective is the modulation of the ratio of barnase/barstar based on the identification of an adequate inducible promoter to control the expression of barnase. The previous literature and transcriptomic data were used to identify candidate promoters for barnase expression. Stable transgenic soybean expressing the different promoter:GFP fusions were generated to test the spatiotemporal activity of the promoters in the presence of the pathogen. pGmCHIT1 (from G. max) and pgst1 (from Solanum tuberosum) promoters controlling a chitinase and a glutathione-Stransferase, respectively, were identified as induced by soybean rust. The impacts of different stresses on these promoters were evaluated. Molecular constructs with different promoters driving the barnase and barstar gene combination were generated. Nicotiana benthamiana was used to evaluate construct toxicity in the absence of the pathogen. One single construct containing the promoter pgst1 was shown to be non-phytotoxic. This construct was successfully introduced in soybean plants. The generated soybeans were challenged with rust, but no protection was observed. Based on these results, we discuss how to improve the barnase/barstar system to control soybean rust
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Submitted on : Monday, September 9, 2019 - 4:10:09 PM
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Lisa Cabre. Identification and study of promoters induced by Asian soybean rust : application in an artificial cell death system. Microbiology and Parasitology. Université de Lyon, 2019. English. ⟨NNT : 2019LYSE1054⟩. ⟨tel-02281993⟩

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