Label-free quantitative proteomic analysis for identification of in-planta Magnaporthe oryzae proteins induced cell death

The rice blast disease caused by Magnaporthe oryzae (M.oryzae) is one of the most important disease in cultivated rice (Oryza sativa L.) results in extensive loss of productivity worldwide. Therefore, here we utilized the label-free quantitative proteomic approach to identify the novel M. oryzae effectors. A total of 114 were significantly modulated M. oryzae proteins derived from infected rice leaves. Among them, twenty-nine proteins were predicted as secreted proteins by SignalP 4.0. In addition, functional annotation proteins revealed that five proteins were mainly associated with hydrolase family. Consequently, five putative effector genes were cloned into a plant expression vector and fused with MYC tag in the C-terminal. Finally, we found that three in-planta M.oryzae hydrolase proteins were induced cell death after Agrobacterium tumefaciens-mediated transient expression in Nicotiana benthamiana (N.benthamiana). MGP1 (endo-1,4-beta-xylanase) and MGP4 (cellulose-binding protein) were localized in extracellular. Otherwise, MGP5 (cutinase) localized in intracellular space induced cell death. Thus, our results suggest that in-planta M. oryzae hydrolase proteins may play a key role in innate immunity via PAMP (Pathogen-Associated Molecular Pattern) in rice.