Functional Analysis of Oomycete Plasmopara viticola SecretedRxLR Effector Repertoire in Korean Vitis Germplasm
Marc Semunyana, Inyong Jeong, Sun Ha Kim, Jiyoung Min and Sang-Keun Oh*
Department of Applied Biology, Chungnam National University, Daejeon 34134, Korea
Plasmopara viticola causes grapevine downy mildew, one of the most notorious diseases threatening cultivated grapes, and causing enormous damage to vineyards worldwide. The pathogen secretes an array of effector molecules necessary to modulate host biological processes, thus establishing successful infection. In this study, 11 isolates were collected from four grape production areas in South Korea. One of the isolates, JN-9, showed the highest virulence potential, and grape cultivars displayed varying phenotypic reactions to P. viticola JN-9 infection; approximately 66.16% were susceptible, 9.7% were resistant, and 24.14% exhibited extreme resistance. Phylogenetic analysis resulted in a highly common evolutionary relationship, with all Korean isolates clustered in a single monophyletic lineage. JN-9 encodes 1,972 genes for secreted proteins, among which 87 genes were classified as putative RxLR effectors. We tested the function of 45 randomly selected PvRxLR effectors against BAX– and INF1-elicited apoptosis in the non-host Nicotiana benthamiana and resistant grape host cultivars through Agrobacterium-mediated transient transformation. As a result, 37 effectors interfered with both BAX– and INF1-induced apoptosis, while 8 effectors induced hypersensitive cell death (HCD) and activated the accumulation of reactive oxygen species (ROS). Based on electrolytic leakage and the ability of the candidate effectors to accumulate ROS, PvRxL22, PvRxL27, and PvRxL35 were selected as core effectors inducing strong HCD in N. benthamiana. All three effectors rapidly triggered HCD in two grape cultivars, confirming their resistance reaction against downy mildew infection. Our findings suggest that these three PvRxLR effectors serve as potential avurulence factors recognized by resistant vitis cultivars thereby resulting in accumulation of resistance mechanisms to limit pathogen development.