Summary: Rice false smut caused by Ustilaginoidea virens is a devastating rice (Oryza sativa) disease worldwide. However, the molecular mechanisms underlying U. virens–rice interactions are largely unknown. In this study, we identified a secreted protein, Uv1809, as a key virulence factor. Heterologous expression of Uv1809 in rice enhanced susceptibility to rice false smut and bacterial blight. Host‐induced gene silencing of Uv1809 in rice enhanced resistance to U. virens, suggesting that Uv1809 inhibits rice immunity and promotes infection by U. virens. Uv1809 suppresses rice immunity by targeting and enhancing rice histone deacetylase OsSRT2‐mediated histone deacetylation, thereby reducing H4K5ac and H4K8ac levels and interfering with the transcriptional activation of defence genes. CRISPR‐Cas9 edited ossrt2 mutants showed no adverse effects in terms of growth and yield but displayed broad‐spectrum resistance to rice pathogens, revealing a potentially valuable genetic resource for breeding disease resistance. Our study provides insight into defence mechanisms against plant pathogens that inactivate plant immunity at the epigenetic level.

ARGONAUTE (AGO) proteins play crucial roles in plant defence against virus invasion. To date, the role of OsAGO2 in rice antiviral defence remains largely unknown. In this study, we determined that the expression of OsAGO2 in rice was induced upon rice black‐streaked dwarf virus (RBSDV) infection. Using transgenic rice plants overexpressing OsAGO2 and Osago2 mutants generated through transposon‐insertion or CRISPR/Cas9 technology, we found that overexpression of OsAGO2 enhanced rice susceptibility to RBSDV infection. Osago2 mutant lines exhibited strong resistance to RBSDV infection through the elicitation of an early defence response, including reprogramming defence gene expression and production of reactive oxygen species (ROS). Compared to Nipponbare control, the expression level of OsHXK1 (HEXOKINASE 1) increased significantly, and the methylation levels of its promoter decreased in the Osago2 mutant on RBSDV infection. The expression profile of OsHXK1 was the opposite to that of OsAGO2 during RBSDV infection. Overexpression of OsHXK1 in rice also induced ROS production and enhanced rice resistance to RBSDV infection. These results indicate that OsHXK1 controls ROS accumulation and is regulated by OsAGO2 through epigenetic regulation. It is noteworthy that the Osago2 mutant plants are also resistant to southern rice black‐streaked dwarf virus infection, another member of the genus Fijivirus. Based on the results presented in this paper, we conclude that OsAGO2 modulates rice susceptibility to fijivirus infection by suppressing OsHXK1 expression, leading to the onset of ROS‐mediated resistance. This discovery may benefit future rice breeding programmes for virus resistance.

Lysine β-hydroxybutyrylation (K_bhb) is an evolutionarily conserved and widespread post-translational modification that is associated with active gene transcription and cellular proliferation. However, its role in phytopathogenic fungi remains unknown. Here, we characterized K_bhb in the rice false smut fungus Ustilaginoidea virens. We identified 2204 K_bhb sites in 852 proteins, which are involved in diverse biological processes. The mitogen-activated protein kinase UvSlt2 is a K_bhb protein, and a strain harboring a point mutation at K72, the K_bhb site of this protein, had decreased UvSlt2 activity and reduced fungal virulence. Molecular dynamic simulations revealed that K72_bhb increases the hydrophobic solvent-accessible surface area of UvSlt2, thereby affecting its binding to its substrates. The mutation of K298_bhb in the septin UvCdc10 resulted in reduced virulence and altered the subcellular localization of this protein. Moreover, we confirmed that the NAD⁺-dependent histone deacetylases UvSirt2 and UvSirt5 are the major enzymes that remove K_bhb in U. virens. Collectively, our findings identify regulatory elements of the K_bhb pathway and reveal important roles for K_bhb in regulating protein localization and enzymatic activity. These findings provide insight into the regulation of virulence in phytopathogenic fungi via post-translational modifications.