| Summary: | The RNA polymerase II degradation factor Degradation Factor 1 (Def1) is important for DNA damage repair and plays various roles in eukaryotes; however, the biological role in plant pathogenic fungi is still unknown. In this study, we investigated the role of Def1 during the development and infection of the rice blast fungus <i>Magnaporthe oryzae</i>. The deletion mutant of <i>Def1</i> displayed slower mycelial growth, less conidial production, and abnormal conidial morphology. The appressoria of Δ<i>def1</i> was impaired in the penetration into host cells, mainly due to blocking in the utilization of conidial storages, such as glycogen and lipid droplets. The invasive growth of the Δ<i>def1</i> mutant was also retarded and accompanied with the accumulation of reactive oxygen species (ROS) inside the host cells. Furthermore, compared with the wild type, Δ<i>def1</i> was more sensitive to multiple stresses, such as oxidative stress, high osmotic pressure, and alkaline/acidic pH. Interestingly, we found that Def1 was modified by <i>O</i>-GlcNAcylation at Ser232, which was required for the stability of Def1 and its function in pathogenicity. Taken together, the <i>O</i>-GlcNAc modified Def1 is required for hyphae growth, conidiation, pathogenicity, and stress response in <i>M. oryzae</i>. This study reveals a novel regulatory mechanism of <i>O</i>-GlcNAc-mediated Def1 in plant pathogenic fungi.
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