State Key Laboratory of North China Crop Improvement and Regulation, Hebei Agricultural University, Baoding, 071001, China.
BMC Plant Biol. 2021 May 15;21(1):220. doi: 10.1186/s12870-021-02992-w.
Verticillium wilt, caused by the soil borne fungus Verticillium dahliae, is a major threat to cotton production worldwide. An increasing number of findings indicate that WAK genes participate in plant-pathogen interactions, but their roles in cotton resistance to V. dahliae remain largely unclear.
Here, we carried out a genome-wide analysis of WAK gene family in Gossypium hirsutum that resulted in the identification of 81 putative GhWAKs, which were all predicated to be localized on plasma membrane. In which, GhWAK77 as a representative was further located in tobacco epidermal cells using transient expression of fluorescent fusion proteins. All GhWAKs could be classified into seven groups according to their diverse protein domains, indicating that they might sense different outside signals to trigger intracellular signaling pathways that were response to various environmental stresses. A lot of cis-regulatory elements were predicted in the upstream region of GhWAKs and classified into four main groups including hormones, biotic, abiotic and light. As many as 28 GhWAKs, playing a potential role in the interaction between cotton and V. dahliae, were screened out by RNA-seq and qRT-PCR. To further study the function of GhWAKs in cotton resistance to V. dahliae, VIGS technology was used to silence GhWAKs. At 20 dpi, VIGSed plants exhibited more chlorosis and wilting than the control plants. The disease indices of VIGSed plants were also significantly higher than those of the control. Furthermore, silencing of GhWAKs significantly affected the expression of JA- and SA-related marker genes, increased the spread of V. dahliae in the cotton stems, dramatically compromised V. dahliae-induced accumulation of lignin, HO and NO, but enhanced POD activity.
Our study presents a comprehensive analysis on cotton WAK gene family for the first time. Expression analysis and VIGS assay provided direct evidences on GhWAKs participation in the cotton resistance to V. dahliae.
由土壤传播真菌黄萎轮枝菌引起的黄萎病是全球棉花生产的主要威胁。越来越多的研究结果表明，WAK 基因参与了植物-病原体相互作用，但它们在棉花对黄萎轮枝菌的抗性中的作用在很大程度上仍不清楚。
在这里，我们对陆地棉基因组中的 WAK 基因家族进行了全基因组分析，共鉴定出 81 个推定的 GhWAKs，它们都被预测定位于质膜上。其中，GhWAK77 作为一个代表，通过瞬时表达荧光融合蛋白被进一步定位于烟草表皮细胞。根据不同的蛋白质结构域，所有的 GhWAKs 可分为 7 个组，表明它们可能感知不同的外部信号，以触发细胞内信号通路，从而对各种环境胁迫做出反应。在 GhWAKs 的上游区域预测到了大量的顺式调控元件，并分为激素、生物、非生物和光四个主要组。通过 RNA-seq 和 qRT-PCR 筛选出 28 个可能在棉花与黄萎轮枝菌相互作用中起作用的 GhWAKs。为了进一步研究 GhWAKs 在棉花对黄萎轮枝菌抗性中的功能，我们使用 VIGS 技术沉默 GhWAKs。在 20dpi 时，VIGSed 植物比对照植物表现出更多的黄化和萎蔫。VIGSed 植物的病情指数也明显高于对照。此外，GhWAKs 的沉默显著影响了 JA 和 SA 相关标记基因的表达，增加了黄萎轮枝菌在棉花茎中的扩散，大大削弱了黄萎轮枝菌诱导的木质素、HO 和 NO 的积累，但增强了 POD 活性。
我们的研究首次对棉花 WAK 基因家族进行了全面分析。表达分析和 VIGS 试验为 GhWAKs 参与棉花对黄萎轮枝菌的抗性提供了直接证据。
