Journal of Integrative Agriculture

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胁迫调控因子FgWhi2和磷酸酶FgPsr1在调控禾谷镰孢菌次生代谢物生物合成和对杀菌剂的响应中具有至关重要的作用

  

  • 修回日期:2023-12-19

The stress regulator FgWhi2 and phosphatase FgPsr1 play crucial roles in the regulation of secondary metabolite biosynthesis and the response to fungicides in Fusarium graminearum

Zhang Jie1,2, Gao Han1, Ren Fuhao1, Zhou Zehua3, Wu Huan1, Zhao Huahua1, Zhang Lu1,2, Zhou Mingguo1,2, Duan Yabing1,2#   

  1. 1 College of Plant Protection, Nanjing Agricultural University, Nanjing, 210095, China

    2 Sanya Institute, Nanjing Agricultural University, Sanya, 572025, China

    3 Hunan Provincial Key Laboratory for Biology and Control of Plant Pests, Hunan Agricultural University, Changsha, 410128, China

  • Revised:2023-12-19
  • About author: Zhang Jie, Tel.: +8618851173500, E-mail: 2018202061@njau.edu.cn; #Correspondence Duan Yabing, Tel: +86-25-84395641, E-mail: dyb@njau.edu.cn
  • Supported by:
    This work was supported by the National Key Research and Development Program of China (2022YFD1400100), Jiangsu Agriculture Science and Technology Innovation Fund (CX (21) 2037), the Guidance Foundation, the Hainan Institute of Nanjing Agricultural University (NAUSY-MS03), and Postgraduate Research & Practice Innovation Program of Jiangsu Province (KYCX20_0596).

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摘要：

在酵母中，胁迫应答蛋白Whi2与磷酸酶Psr1相互作用并以复合体形式调控生长、繁殖、侵染和胁迫应答，但它们在禾谷镰孢菌中的调控作用尚不明确。为了明确Whi2与Psr1在禾谷镰孢菌中的调控作用，本文鉴定了Whi2和Psr1在禾谷镰孢菌中的同源基因，并通过构建缺失突变体探究其功能。结果表明，FgWHI2与FgPSR1缺失突变体∆Fgwhi2、∆Fgpsr1及双缺失突变体∆Fgwhi2-Fgpsr1对1.2 M NaCl、1.2 M KCl、0.01% H2O2、0.05%刚果红及0.05% SDS敏感性显著降低，表明FgWHI2与FgPSR1对禾谷镰孢菌响应外界渗透压胁迫、氧化胁迫、细胞壁胁迫有重要作用。通过对突变体麦角甾醇产量及三唑类药剂敏感性的测定，我们发现FgWHI2与FgPSR1不调控禾谷镰孢菌对三唑类杀菌剂药敏性，但调控禾谷镰孢菌麦角甾醇生物合成。通过对禾谷镰孢菌FgCYP51s基因转录水平的测定，我们发现FgWHI2与FgPSR1正调控FgCYP51-C的转录表达；FgWHI2与FgPSR1过表达能够提高FgCYP51-A的转录水平，但FgWHI2与FgPSR1的缺失对FgCYP51-A的转录水平没有影响；FgWHI2与FgPSR1不调控FgCYP51-B的转录表达。同时，我们发现FgWHI2和FgPSR1参与禾谷镰孢菌的有性生殖和无性生殖。通过研究发现FgWHI2和FgPSR1缺失突变体的脱氧雪腐镰刀烯醇毒素（DON）产量降低超90%，且DON产量与FgWHI2和FgPSR1转录水平呈正相关。此外，我们还观察到FgWHI2和FgPSR1参与调控禾谷镰孢菌对百菌清、氟啶胺、嘧菌酯、氰烯菌酯和寡霉素的敏感性，并发现百菌清与氟啶胺之间存在交互抗性，且百菌清和氟啶胺通过抑制FgWHI2表达来阻碍DON毒素的生物合成。有趣的是，百菌清和氟啶胺处理后，FgWhi2和FgPsr1的亚细胞定位显著改变，并且共定位比例增加。综上所述，这些结果表明FgWHI2和FgPsr1在禾谷镰孢菌胁迫应激反应、生殖过程、次生代谢产物合成和对百菌清和氟啶胺的响应中起着至关重要的作用。通过本文的研究，首次明确了FgWhi2与FgPsr1对禾谷镰孢菌DON毒素生物合成的调控作用，拓展了禾谷镰孢菌DON毒素合成调控网络，为小麦赤霉病防控过程中兼顾“防病”与“降毒”及开发有效的杀菌剂组合提供了潜在靶标、理论依据及技术参考。

Abstract:

In yeast, the stress-responsive protein Whi2 interacts with phosphatase Psr1 to form a complex that regulates cell growth, reproduction, infection, and the stress response. However, the roles of Whi2 and Psr1 in Fusarium graminearum remain unclear. In this study, we identified homologous genes of WHI2 and PSR1 in F. graminearum and evaluated their functions by constructing deletion mutants. By comparing the responses of the mutants to different stressors, we found that FgWHI2 and FgPSR1 were involved in responding to osmotic, cell wall and cell membrane stresses, while also affecting the sexual and asexual reproduction in F. graminearum. Our studies demonstrated that FgWHI2 and FgPSR1 regulate the biosynthesis of ergosterol and the transcriptional level of FgCYP51C, which is a CYP51 paralogues unique to Fusarium species. This study also found that the deoxynivalenol (DON) production of FgWHI2 and FgPSR1 deletion mutants was reduced by ≥ 90% and DON production was positively correlated with the transcriptional levels of FgWHI2 and FgPSR1. In addition, we observed that FgWHI2 and FgPSR1 were involved in regulating the sensitivity of F. graminearum to chlorothalonil, fluazinam, azoxystrobin, phenamacril, and oligomycin. This study revealed the existence of cross-resistance between chlorothalonil and fluazinam. chlorothalonil and fluazinam inhibited DON biosynthesis by suppressing the expression of FgWHI2. Interestingly, the subcellular localization of FgWhi2 and FgPsr1 was significantly altered after treatment with chlorothalonil and fluazinam, with increased co-localization. Collectively, these findings indicate that FgWHI2 and FgPSR1 play crucial roles in stress response mechanisms, reproductive processes, secondary metabolite synthesis, and fungicide sensitivity in F. graminearum.

Key words: DON , Ergosterol , FgWHI2 ,  FgPSR1 ,  Fungicides ,  Fusarium graminearum ,  Stress responses