Gai Yunpeng, Li Lei, Ma Haijie, Riely Brendan K, Liu Bing, Li Hongye
Key Lab of Molecular Biology of Crop Pathogens and Insects, Ministry of Agriculture, Institute of Biotechnology, Zhejiang University, Hangzhou 310058, China.
School of Agriculture and Food Sciences, Zhejiang Agriculture & Forestry University, Hangzhou 311300, China.
Appl Environ Microbiol. 2021 Mar 1;87(4). doi: 10.1128/AEM.01911-20. Epub 2020 Dec 4.
Methionine is a unique sulfur-containing amino acid, which plays an important role in biological protein synthesis and various cellular processes. Here, we characterized the biological functions of , , and in the tangerine pathotype of Morphological analysis showed that the mutants lacking , , or resulted in less aerial hypha and fewer conidia in artificial media. Pathogenicity analysis showed that , , and are required for full virulence. The defects in vegetative growth, conidiation and virulence of Δ, Δ, and Δ can be restored by exogenous methionine and homocysteine, indicating that , , and are required for methionine biosynthesis. However, exogenous cysteine only restored the growth and virulence defects of Δ but not Δ, suggesting that is essential for cysteine biosynthesis. Oxidant sensitivity assay showed that only Δ is sensitive to HO and many ROS-generating compounds, indicating that is essential for oxidative tolerance. Interestingly, fungicides indoor bioassays showed that only the Δ mutants are susceptive to chlorothalonil, a fungicide that could bind to the cysteine of glyceraldehyde-3-phosphate dehydrogenase. Comparative transcriptome analysis showed that the inactivation of , , , or significantly affected the expression of methionine metabolism-related genes. Moreover, the inactivation of Aa significantly affected the expression of many genes related to glutathione metabolism, which is essential for ROS tolerance. Taken together, our study provides genetic evidence to define the critical roles of , , , and in cysteine and methionine metabolism, fungal development and virulence of The transcription factor METR regulating methionine metabolism is essential for reactive oxygen species (ROS) tolerance and virulence in many phytopathogenic fungi. However, the underlying regulatory mechanism of METR involved in this process is still unclear. In the present study, we generated , and deletion mutants and compared these mutants with disrupted mutants. Interestingly, we found that , and are required for vegetative growth, conidiation, and pathogenicity in , but not for ROS tolerance and cysteine metabolism. Furthermore, we found that METR is involved in the biosynthesis of cysteine, which is an essential substrate for the biosynthesis of methionine and glutathione. This study emphasizes the critical roles of , , , in the regulation of cysteine and methionine metabolism, as well as the cross-link with glutathione-mediated ROS tolerance in phytopathogenic fungi, which provides a foundation for future investigations.
甲硫氨酸是一种独特的含硫氨基酸，在生物蛋白质合成和各种细胞过程中发挥着重要作用。在此，我们对[具体基因名称]在[病原菌名称]的柑橘致病型中的生物学功能进行了表征。形态学分析表明，缺失[具体基因名称]的突变体在人工培养基中产生的气生菌丝较少，分生孢子也较少。致病性分析表明，[具体基因名称]对于完全致病力是必需的。Δ[具体基因名称]、Δ[具体基因名称]和Δ[具体基因名称]在营养生长、产孢和致病力方面的缺陷可通过外源甲硫氨酸和同型半胱氨酸恢复，这表明[具体基因名称]对于甲硫氨酸生物合成是必需的。然而，外源半胱氨酸仅恢复了Δ[具体基因名称]的生长和致病力缺陷，而未恢复Δ[具体基因名称]的，这表明[具体基因名称]对于半胱氨酸生物合成至关重要。氧化剂敏感性测定表明，只有Δ[具体基因名称]对H₂O₂和许多产生活性氧的化合物敏感，这表明[具体基因名称]对于氧化耐受至关重要。有趣的是，杀菌剂室内生物测定表明，只有Δ[具体基因名称]突变体对百菌清敏感，百菌清是一种可与3-磷酸甘油醛脱氢酶的半胱氨酸结合的杀菌剂。比较转录组分析表明，[具体基因名称]、[具体基因名称]、[具体基因名称]或[具体基因名称]的失活显著影响了甲硫氨酸代谢相关基因的表达。此外，Aa的失活显著影响了许多与谷胱甘肽代谢相关基因的表达，而谷胱甘肽代谢对于活性氧耐受至关重要。综上所述，我们的研究提供了遗传学证据，以确定[具体基因名称]、[具体基因名称]、[具体基因名称]和[具体基因名称]在半胱氨酸和甲硫氨酸代谢、真菌发育以及[病原菌名称]致病力中的关键作用。调节甲硫氨酸代谢的转录因子METR对于许多植物病原真菌中的活性氧（ROS）耐受和致病力至关重要。然而，METR参与此过程的潜在调控机制仍不清楚。在本研究中，我们构建了[具体基因名称]、[具体基因名称]和[具体基因名称]缺失突变体，并将这些突变体与[具体基因名称]破坏突变体进行了比较。有趣的是，我们发现[具体基因名称]、[具体基因名称]和[具体基因名称]对于[病原菌名称]的营养生长、产孢和致病性是必需的，但对于活性氧耐受和半胱氨酸代谢并非必需。此外，我们发现METR参与半胱氨酸的生物合成，而半胱氨酸是甲硫氨酸和谷胱甘肽生物合成的必需底物。本研究强调了[具体基因名称]、[具体基因名称]、[具体基因名称]、[具体基因名称]在调节半胱氨酸和甲硫氨酸代谢以及与植物病原真菌中谷胱甘肽介导的活性氧耐受的交联中的关键作用，为未来的研究奠定了基础。
