Nucleoside Diphosphate Kinase FgNdpk Is Required for DON Production and Pathogenicity by Regulating the Growth and Toxisome Formation of Fusarium graminearum.

Nucleoside diphosphate kinases (NDPKs) are nucleotide metabolism enzymes that play different physiological functions in different species. However, the roles of NDPK in phytopathogen and mycotoxin production are not well understood. In this study, we showed that Fusarium graminearum FgNdpk is important for vegetative growth, conidiation, sexual development, and pathogenicity. Furthermore, FgNdpk is required for deoxynivalenol (DON) production; deletion of FgNDPK downregulates the expression of DON biosynthesis genes and disrupts the formation of FgTri4-GFP-labeled toxisomes, while overexpression of FgNDPK significantly increases DON production. Interestingly, FgNdpk colocalizes with the DON biosynthesis proteins FgTri1 and FgTri4 in the toxisome, and coimmunoprecipitation (Co-IP) assays show that FgNdpk associates with FgTri1 and FgTri4 in vivo and regulates their localizations and expressions, respectively. Taken together, these data demonstrate that FgNdpk is important for vegetative growth, conidiation, and pathogenicity and acts as a key protein that regulates toxisome formation and DON biosynthesis in F. graminearum.

ESCRT-III accessory proteins regulate fungal development and plant infection in Fusarium graminearum.

Ubiquitinated biosynthetic and surface proteins destined for degradation are sorted into the lysosome/vacuole via the multivesicular body sorting pathway, which depends on the function of ESCRT machinery. Fusarium head blight (FHB) caused by Fusarium graminearum is one of the most devastating diseases for wheat and barley worldwide. To better understand the role of ESCRT machinery in F. graminearum, we investigated the function of ESCRT-III accessory proteins FgVps60, FgDid2 and FgIst1 in this study. FgVps60-GFP, FgDid2-GFP and FgIst1-GFP are localized to punctate structures adjacent to the vacuolar membrane except for FgIst1-GFP that is also found in the nucleus. Then, the gene deletion mutants ΔFgvps60, ΔFgdid2 and ΔFgist1 displayed defective growth to a different extent. ΔFgvps60 and ΔFgdid2 but not ΔFgist1 also showed significant reduction in hydrophobicity on cell surface, conidiation, perithecia production and virulence. Interestingly, ΔFgist1 mutant produced a significantly higher level of DON while showing a minor reduction in pathogenicity. Microscopic analyses revealed that FgVps60 but not FgIst1 and FgDid2 is necessary for endocytosis. Moreover, spontaneous mutations were identified in the ΔFgvps60 mutant that partially rescued its defects in growth and conidiation. Taken together, we conclude that ESCRT-III accessory proteins play critical roles in growth, reproduction and plant infection in F. graminearum.

FgBud3, a Rho4-Interacting Guanine Nucleotide Exchange Factor, Is Involved in Polarity Growth, Cell Division and Pathogenicity of Fusarium graminearum.

Rho GTPases are signaling macromolecules that are associated with developmental progression and pathogenesis of Fusarium graminearum. Generally, enzymatic activities of Rho GTPases are regulated by Rho GTPase guanine nucleotide exchange factors (RhoGEFs). In this study, we identified a putative RhoGEF encoding gene (FgBUD3) in F. graminearum database and proceeded further by using a functional genetic approach to generate FgBUD3 targeted gene deletion mutant. Phenotypic analysis results showed that the deletion of FgBUD3 caused severe reduction in growth of FgBUD3 mutant generated during this study. We also observed that the deletion of FgBUD3 completely abolished sexual reproduction and triggered the production of abnormal asexual spores with nearly no septum in ΔFgbud3 strain. Further results obtained from infection assays conducted during this research revealed that the FgBUD3 defective mutant lost its pathogenicity on wheat and hence, suggests FgBud3 plays an essential role in the pathogenicity of F. graminearum. Additional, results derived from yeast two-hybrid assays revealed that FgBud3 strongly interacted with FgRho4 compared to the interaction with FgRho2, FgRho3, and FgCdc42. Moreover, we found that FgBud3 interacted with both GTP-bound and GDP-bound form of FgRho4. From these results, we subsequently concluded that, the Rho4-interacting GEF protein FgBud3 crucially promotes vegetative growth, asexual and sexual development, cell division and pathogenicity in F. graminearum.