Pathological hypertrophy (cell enlargement) plays an important role in the development of citrus canker, but its regulators are largely unknown. Although WRKY22 is known to be involved in pathogen-triggered immunity and positively regulates resistance to bacterial pathogens in Arabidopsis, rice and pepper, the CRISPR/Cas9-mediated partial knockout of CsWRKY22 improves resistance to Xanthomonas citri subsp. citri (Xcc) in Wanjincheng orange (Citrus sinensis Osbeck). Here, we demonstrate that CsWRKY22 is a nucleus-localized transcriptional activator. CsWRKY22-overexpressing plants exhibited dwarf phenotypes that had wrinkled and thickened leaves and were more sensitive to Xcc, whereas CsWRKY22-silenced plants showed no visible phenotype changes and were more resistant to Xcc. Microscopic observations revealed that the overexpression of CsWRKY22 increased cell size in the spongy mesophyll. Transcriptome analysis showed that cell growth-related pathways, such as the auxin and brassinosteroid hormonal signaling and cell wall organization and biogenesis pathways, were significantly upregulated upon CsWRKY22 overexpression. Interestingly, CsWRKY22 activated the expression of CsLOB1, which is a key gene regulating susceptibility to citrus canker. We further confirmed that CsWRKY22 bound directly to the W-boxes just upstream of the transcription start site of CsLOB1 in vivo and in vitro. We conclude that CsWRKY22 enhances susceptibility to citrus canker by promoting host hypertrophy and CsLOB1 expression. Thus, our study provides new insights into the mechanism regulating pathological hypertrophy and the function of WRKY22 in citrus.

中文翻译：

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转录因子 WRKY22 通过增强细胞增大和 CsLOB1 表达来调节甜橙 (Citrus sinensis Osbeck) 的溃疡病易感性

病理性肥大（细胞增大）在柑橘溃疡病的发展中起重要作用，但其调节因子在很大程度上是未知的。尽管已知 WRKY22 参与病原体触发的免疫并正调节拟南芥、水稻和辣椒对细菌病原体的抗性，但 CRISPR/Cas9 介导的部分敲除CsWRKY22提高抵抗力柑橘黄单胞菌亚种柑橘(Xcc) 万金城橙 (柑橘奥斯贝克）。在这里，我们证明 CsWRKY22 是一种定位于细胞核的转录激活因子。CsWRKY22- 过度表达的植物表现出矮化表型，叶片起皱和增厚，对 Xcc 更敏感，而CsWRKY22- 沉默的植物没有表现出明显的表型变化，并且对 Xcc 更具抗性。显微镜观察显示，过表达CsWRKY22海绵状叶肉中的细胞大小增加。转录组分析表明，细胞生长相关通路，如生长素和油菜素内酯激素信号传导以及细胞壁组织和生物发生通路，在CsWRKY22过度表达。有趣的是，CsWRKY22 激活了CsLOB1，是调控柑橘溃疡病易感性的关键基因。我们进一步证实 CsWRKY22 直接与转录起始位点上游的 W 盒结合。CsLOB1在体内和体外。我们得出结论，CsWRKY22 通过促进宿主肥大和增强对柑橘溃疡病的易感性CsLOB1表达。因此，我们的研究为柑橘中调节病理性肥大的机制和 WRKY22 的功能提供了新的见解。