P-203 - Engineering Pathogen Resistance: Enhancing metabolic Flux through the Indole-Glucosinolate Pathway by overexpression of a single Pathway Enzyme

Indole-Glucosinolates (IGs) are specialized metabolites produced in Brassicaceae, which possess valuable antimicrobial properties. In the Arabidopsis root, the complex biosynthetic pathway is induced upon infection generally controlled on transcriptional level. IGs have been identified as crucial defense compound against the vascular pathogenic fungus Verticillium longisporum, which is an agronomical thread in rape seed cultures. Aiming at metabolic pathway engineering, we obtained increased resistance by overexpression of a single bottleneck enzyme CYP81F2 (CYTOCHROME P450 MONOOXYGENASE 81F2). Mutant analyses of cyp81f2, as well as of the respective ABC transporters confirmed that resistance to V. longisporum depends on IGs and their cellular export. Indeed, metabolic analyses revealed an increased flux through the pathway, which is even more pronounced upon infection. Important for its biotechnological application, detailed phenotypical inspection revealed no growth alteration between wt and overexpressing plants. Screening of an Arabidopsis ecotype collection harboring mutations in the CYP81F2 promoter identified lines showing increased resistance, which was correlated with overexpression of the CYP81F2 gene. Hence, this proof-of-concept approach provides a strategy how pathogen resistance can be gained by metabolic pathway engineering. Moreover, targeted promoter editing offers an opportunity to avoid transgenic overexpression approaches for agricultural application.