Aflatoxin, a carcinogenic secondary metabolite produced by Aspergillus flavus, is a significant threat to human health and agricultural production. Histone 2-hydroxyisobutyrylation is a novel post-translational modification that regulates various biological processes, including secondary metabolism. In this study, we identified the novel histone 2-hydroxyisobutyryltransferase Afngg1 in A. flavus, and explored its role in cell growth, development and aflatoxin biosynthesis. Afngg1 gene deletion markedly decreased lysine 2-hydroxyisobutyrylation modification of histones H4K5 and H4K8 compared with the control strain. Additionally, Afngg1 deletion inhibited mycelial growth of A. flavus, and the number of conidia and hydrophobicity were significantly decreased. Notably, aflatoxin B1 biosynthesis and sclerotia production were completely inhibited in the ΔAfngg1 strain. Furthermore, the pathogenicity of the ΔAfngg1 strain infecting peanut and corn grains was also diminished, including reduced spore production and aflatoxin biosynthesis compared with A. flavus control and Afngg1 complementation strains. Transcriptome analysis showed that, compared with control strains, differentially expressed genes in ΔAfngg1 were mainly involved in chromatin remodelling, cell development, secondary metabolism and oxidative stress. These results suggest that Afngg1 is involved in histone 2-hydroxyisobutyrylation and chromatin modification, and thus affects cell development and aflatoxin biosynthesis in A. flavus. Our results lay a foundation for in-depth research on the 2-hydroxyisobutyrylation modification in A. flavus, and may provide a novel target for aflatoxin contamination prevention.
Lysine acetylation (Kac) is a protein post-translational modification (PTM) widely found in plants that plays vital roles in metabolic pathways. Although seed germination and development are regulated by Kac, its potential function in seed ageing remains to be investigated. Our preliminary study demonstrated that Kac levels were altered during wheat seed artificial ageing. However, its specific role in this process still needs to be elucidated. Here, we performed quantitative acetylation proteomics analysis of soft wheat seeds with different germination rates during artificial ageing. A total of 175 acetylation proteins and 255 acetylation modification sites were remarkably changed. The differentially acetylated proteins were enriched in metabolism; response to harsh intracellular environment, such as ROS; protein storage and processing. Notably, expression, point mutation to mimic Kac by K to Q mutation at K80 and K138, protein purification and enzyme activity detection revealed that the Kac of ROS-scavenging glutathione transferase attenuated its activity, indicating that the defense ability of wheat seeds to stress gradually diminished, and the ageing process was inevitable. Collectively, our data provide a basis for further understanding the roles of Kac in seed ageing and might aid in the development of new techniques to prolong seed viability and food quality.
Aflatoxin is a carcinogenic secondary metabolite that poses a serious threat to human and animal health. Some C2H2 transcription factors are associated with fungal growth and secondary metabolic regulation. In this study, we characterized the role of AflZKS3, a putative C2H2 transcription factor based on genome annotation, in the growth and aflatoxin biosynthesis of A. flavus and explored its possible mechanisms of action. Surprisingly, the protein was found to be located in the cytoplasm, and gene deletion in A. flavus resulted in defective growth and conidia formation, as well as increased sensitivity to the fluorescent brightener Calcofluor white, Congo red, NaCl, and sorbitol stress. Notably, the biosynthesis of aflatoxin B1 was completely inhibited in the ΔAflZKS3 deletion strain, and its ability to infect peanut and corn seeds was also reduced. RNA sequencing showed that differentially expressed genes in the ΔAflZKS3 strain compared with the control and complementation strains were mainly associated with growth, aflatoxin biosynthesis, and oxidative stress. Thus, AflZKS3 likely contributes to growth, cell development, and aflatoxin synthesis in A. flavus. These findings lay the foundation for a deeper understanding of the roles of C2H2 transcription factors in A. flavus and provide a potential biocontrol target for preventing aflatoxin contamination.
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