MoEnd3 regulates appressorium formation and virulence through mediating endocytosis in rice blast fungus Magnaporthe oryzae

Eukaryotic cells respond to environmental stimuli when cell surface receptors are bound by environmental ligands. The binding initiates a signal transduction cascade that results in the appropriate intracellular responses. Studies have shown that endocytosis is critical for receptor internalization and signaling activation. In the rice blast fungus Magnaporthe oryzae , a non-canonical G-protein coupled receptor, Pth11, and membrane sensors MoMsb2 and MoSho1 are thought to function upstream of G-protein/cAMP signaling and the Pmk1 MAPK pathway to regulate appressorium formation and pathogenesis. However, little is known about how these receptors or sensors are internalized and transported into intracellular compartments. We found that the MoEnd3 protein is important for endocytic transport and that the Δ Moend3 mutant exhibited defects in efficient internalization of Pth11 and MoSho1. The Δ Moend3 mutant was also defective in Pmk1 phosphorylation, autophagy, appressorium formation and function. Intriguingly, restoring Pmk1 phosphorylation levels in Δ Moend3 suppressed most of these defects. Moreover, we demonstrated that MoEnd3 is subject to regulation by MoArk1 through protein phosphorylation. We also found that MoEnd3 has additional functions in facilitating the secretion of effectors, including Avr-Pia and AvrPiz-t that suppress rice immunity. Taken together, our findings suggest that MoEnd3 plays a critical role in mediating receptor endocytosis that is critical for the signal transduction-regulated development and virulence of M . oryzae . During the interaction between the rice blast fungus Magnaporthe oryzae and the host, the pathogen employs a series of receptors and sensors at the plasma membrane to recognize host surface cues and to activate signal transduction pathways required for appressorium formation and pathogenicity. We found that MoEnd3-mediated endocytosis is responsible for internalization of non-canonical GPCR Pth11 and the sensor MoSho1 to endosomal compartments. This is important for activating the downstream Pmk1 MAPK pathway to control appressorium formation and penetration. MoEnd3 is regulated through phosphorylation by the actin-regulating kinase MoArk1. In addition, MoEnd3 has a role in establishing effector secretion required for suppressing rice immunity. Our studies provide evidence that endocytosis is required for normal signaling and virulence in M . oryzae .