A Cell Wall Integrity-Related MAP Kinase Kinase Kinase AflBck1 Is Required for Growth and Virulence in Fungus Aspergillus flavus.

Aspergillus flavus represents an important fungal pathogen, causing severe economic losses in crops. The mitogen-activated protein (MAP) kinase signaling pathway contributes to many physiological processes, but its precise role in A. flavus is not yet fully understood. In this study, we focused on the AflBck1 gene, which encodes a MAP kinase kinase kinase of the Slt2-MAPK pathway. Targeted deletion of AflBck1 led to a significant defect in growth and development, and a AflBck1-deleted mutant (∆AflBck1) showed higher sensitivity to cell-wall stress than wild type (WT). Importantly, we observed that ∆AflBck1 displayed an enhanced ability to produce aflatoxin, a potential carcinogenic mycotoxin. However, the pathogenicity of the ∆AflBck1 mutant was markedly reduced in peanut seeds. We also presented evidence that AflBck1 was genetically epistatic to AflMkk2 in the Slt2-MAPK pathway. Finally, we found that loss of the proline-rich region at the N terminus of AflBck1 affected the reproduction of A. flavus. Collectively, this study not only extended the understanding that the MAPK pathway regulated A. flavus pathogenicity but also provided a possible strategy to control A. flavus contamination.

Contribution of peroxisomal protein importer AflPex5 to development and pathogenesis in the fungus Aspergillus flavus.

Peroxisomes are important organelles that have diverse metabolic functions and participate in the pathogenicity of fungal pathogens. Previous studies indicate that most functions of peroxisomes are dependent on peroxisomal matrix proteins, which are delivered from the cytoplasm into peroxisomes by peroxisomal protein importers. In this study, the roles of peroxisomal protein importer AflPex5 were investigated in Aspergillus flavus with the application of gene disruption. AflPex5 deletion mutants failed to localize the fluorescently fused peroxisomal targeting signal 1 (PTS1) proteins to peroxisomes. Deletion of AflPex5 caused defects in sporulation, sclerotial formation, aflatoxin biosynthesis, stress response, and plant infection. Moreover, AflPex5 null mutants exhibited a significant defect in carbon metabolism and oxidants' clearance. These results indicate that the PTS1 pathway mediated by AflPex5 serves as an important role in the development, metabolism, and pathogenesis of A. flavus.

The MAP kinase AflSlt2 modulates aflatoxin biosynthesis and peanut infection in the fungus Aspergillus flavus.

Aflatoxin contamination in food and feed products has been brought into sharp focus over the last few decades in the world. However, there is no effective strategy for solving the problem thus far. Therefore, basic research on the aflatoxin-producer Aspergillus flavus is an urgent need. The vital role of mitogen-activated protein kinases (MAPKs) in signal transduction has been documented in various pathogenic fungi, but their functions in A. flavus have rarely been investigated. Herein, we characterized the detailed function of one of these MAPKs, AflSlt2. Targeted deletion of AflSlt2 gene indicates that this kinase is required for vegetative growth, conidia generation, and sclerotium formation. The analysis of AflSlt2 deletion mutant revealed hypersensitivity to cell wall-damaging chemicals and resistance against hydrogen peroxide. Interestingly, the ability of the ΔAflSlt2 mutant to generate aflatoxins in medium was significantly increased compared to wild type. However, a pathogenicity assay indicated that the ΔAflSlt2 mutant was deficient in peanut infection. Site-directed mutation study uncovered that the function of AflSlt2 was dependent on the phosphorylated residues (Thr-186 and Tyr-188) within the activation loop and the phosphotransfer residue (Lys-52) within the subdomain II. Interestingly, an autophosphorylation mutant of AflSlt2 (AflSlt2R66S) displayed wild type-like phenotypes. Bringing these observations together, we propose that Slt2-MAPK pathway is involved in development, stress response, aflatoxin biosynthesis, and pathogenicity in A. flavus. This study may be useful to unveil the regulation mechanism of aflatoxin biosynthesis and provide strategy to control A. flavus contamination.