Post-Translational Modification β-Hydroxybutyrylation Regulates Ustilaginoidea virens Virulence

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.