Targeting of anti-microbial proteins to the hyphal surface amplifies protection of crop plants against Phytophthora pathogens

Phytophthora pathogens are a persistent threat to the world's commercially important agricultural crops, including potato and soybean. Current strategies aim at reducing crop losses rely mostly on disease-resistance breeding and chemical pesticides, which can be frequently overcome by the rapid adaptive evolution of pathogens. Transgenic crops with intrinsic disease resistance offer a promising alternative and continue to be developed. Here, we explored Phytophthora-derived PI3P (phosphatidylinositol 3-phosphate) as a novel control target, using proteins that bind this lipid to direct secreted anti-microbial peptides and proteins (AMPs) to the surface of Phytophthora pathogens. In transgenic Nicotiana benthamiana, soybean, and potato plants, significantly enhanced resistance to different pathogen isolates was achieved by expression of two AMPs (GAFP1 or GAFP3 from the Chinese medicinal herb Gastrodia elata) fused with a PI3P-specific binding domain (FYVE). Using the soybean pathogen P. sojae as an example, we demonstrated that the FYVE domain could boost the activities of GAFPs in multiple independent assays, including those performed in vitro, in vivo, and in planta. Mutational analysis of P. sojae PI3K1 and PI3K2 genes of this pathogen confirmed that the enhanced activities of the targeted GAFPs were correlated with PI3P levels in the pathogen. Collectively, our study provides a new strategy that could be used to confer resistance not only to Phytophthora pathogens in many plants but also potentially to many other kinds of plant pathogens with unique targets.