FreB is involved in the ferric metabolism and multiple pathogenicity-related traits of Verticillium dahliae
FreB is involved in the ferric metabolism and multiple pathogenicity-related traits of Verticillium dahliae
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FreB 参与大丽黄萎病菌的铁代谢和多种致病性相关性状
DOI：
10.1007/s00294-017-0780-x
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发表时间：
2018-06
期刊：
影响因子：
2.5
通讯作者：
Cheng Hongmei
中科院分区：
文献类型：
--
作者：
Rehman Latifur;Su Xiaofeng;Li Xiaokang;Qi Xiliang;Guo Huiming;Cheng Hongmei
铁还原酶是一种重要的膜蛋白，参与将环境中的三价铁还原为生物可利用的二价铁。在最主要的植物病原真菌大丽轮枝菌中，这些铁还原酶没有被详细研究。本研究通过原生质体转化法获得FreB基因敲除突变体（ΔFreB）和互补突变体（ΔFreB-C），探讨FreB基因（VDAG_06616）在大丽轮枝菌铁还原和毒力中的作用。当在添加FeSO 4、FeCl 3和不含铁的培养基上培养时，ΔFreB表现出显著降低的生长和孢子产生，尤其是在不含铁的培养基上。ΔFreB的跨膜铁还原酶活性比野生型菌株（Vd-wt）降低了50%。在ΔFreB-C中活性完全恢复。与此同时，其它相关基因（Frect-4、Frect-5、Frect-6和Met）在ΔFreB中的表达量也明显增加。与Vd-wt和ΔFreB-C相比，ΔFreB-1和ΔFreB-2在不同碳源（淀粉、蔗糖、半乳糖和木糖）上的菌落直径和孢子数均有所降低。当将100 µM H2 O2应用于真菌培养物时，平板扩散试验显示ΔFreB-1和ΔFreB-2对氧化应激也高度敏感。接种本氏烟后，ΔFreB表现出的病害症状比Vd-wt和ΔFreB-C少。因此，本研究结果不仅表明FreB介导了大丽轮枝菌铁代谢，是其致病力发挥的必要条件，而且为进一步揭示该菌致病机理奠定了基础。
Ferric reductases are integral membrane proteins involved in the reduction of environmental ferric iron into the biologically available ferrous iron. In the most overwhelming phytopathogenic fungus, Verticillium dahliae, these ferric reductase are not studied in details. In this study we explored the role of FreB gene (VDAG_06616) in the ferric reduction and virulence of V. dahliae by generating the knockout mutants (ΔFreB) and complementary strains (ΔFreB-C) using protoplast transformation. When cultured on media supplemented with FeSO4, FeCl3 and no iron, ΔFreB exhibited significantly reduced growth and spore production especially on media with no iron. Transmembrane ferric reductase activity of ΔFreB was decreased up to 50% than wild type strains (Vd-wt). The activity was fully restored in ΔFreB-C. Meanwhile, the expression levels of other related genes (Frect-4, Frect-5, Frect-6 and Met) were obviously increased in ΔFreB. Compared with the Vd-wt and ΔFreB-C, ΔFreB-1 and ΔFreB-2 were impaired in colony diameter and spore number on different carbon sources (starch, sucrose, galactose and xylose). ΔFreB-1 and ΔFreB-2 were also highly sensitive to oxidative stress as revealed by the plate diffusion assay when 100 µM H2O2 was applied to the fungal culture. When Nicotiana benthamiana plants were inoculated, ΔFreB exhibited less disease symptoms than Vd-wt and ΔFreB-C. In conclusion, the present findings not only indicate that FreB mediates the ferric metabolism and is required for the full virulence in V. dahliae, but would also accelerate future investigation to uncover the pathogenic mechanism of this fungus.
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