Canker disease is caused by the fungus Cytospora chrysosperma and damages a wide range of woody plants, causing major losses to crops and native plants. Plant pathogens secrete virulence-related effectors into host cells during infection to regulate plant immunity and promote colonization. However, the functions of C. chrysosperma effectors remain largely unknown. In this study, we used Agrobacterium tumefaciens-mediated transient expression system in Nicotiana benthamiana and confocal microscopy to investigate the immunoregulation roles and subcellular localization of CcCAP1, a virulence-related effector identified in C. chrysosperma. CcCAP1 was significantly induced in the early stages of infection and contains cysteine-rich secretory proteins, antigen 5, and pathogenesis-related 1 proteins (CAP) superfamily domain with four cysteines. CcCAP1 suppressed the programmed cell death triggered by Bcl-2-associated X protein (BAX) and the elicitin infestin1 (INF1) in transient expression assays with Nicotiana benthamiana. The CAP superfamily domain was sufficient for its cell death-inhibiting activity and three of the four cysteines in the CAP superfamily domain were indispensable for its activity. Pathogen challenge assays in N. benthamiana demonstrated that transient expression of CcCAP1 promoted Botrytis cinerea infection and restricted reactive oxygen species accumulation, callose deposition, and defense-related gene expression. In addition, expression of green fluorescent protein-labeled CcCAP1 in N. benthamiana showed that it localized to both the plant nucleus and the cytoplasm, but the nuclear localization was essential for its full immune inhibiting activity. These results suggest that this virulence-related effector of C. chrysosperma modulates plant immunity and functions mainly via its nuclear localization and the CAP domain.

中文翻译：

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黄子细胞孢子毒力效应子 CcCAP1 主要定位于植物细胞核抑制植物免疫反应


溃疡病由真菌黄子细胞孢子引起，损害多种木本植物，对农作物和本土植物造成重大损失。植物病原体在感染过程中将毒力相关效应物分泌到宿主细胞中，以调节植物免疫并促进定植。然而， C. chrysosperma效应器的功能仍然很大程度上未知。在本研究中，我们使用本塞姆氏烟草中根癌农杆菌介导的瞬时表达系统和共聚焦显微镜来研究 CcCAP1 的免疫调节作用和亚细胞定位，CcCAP1 是在C. chrysosperma中鉴定的毒力相关效应子。 CcCAP1 在感染早期被显着诱导，包含富含半胱氨酸的分泌蛋白、抗原 5 和具有四个半胱氨酸的发病机制相关 1 蛋白 (CAP) 超家族结构域。在烟草瞬时表达测定中，CcCAP1 抑制了 Bcl-2 相关 X 蛋白 (BAX) 和 elicitin infestin1 (INF1) 触发的程序性细胞死亡。 CAP超家族结构域足以发挥其细胞死亡抑制活性，并且CAP超家族结构域中的四个半胱氨酸中的三个对于其活性是不可或缺的。本塞姆氏烟草中的病原体攻击测定表明，CcCAP1 的瞬时表达促进灰葡萄孢感染并限制活性氧的积累、胼胝质沉积和防御相关基因的表达。此外，绿色荧光蛋白标记的 CcCAP1 在N. 本塞米亚纳表明它定位于植物核和细胞质，但核定位对于其完全免疫抑制活性至关重要。这些结果表明， C. chrysosperma的毒力相关效应子主要通过其核定位和 CAP 结构域调节植物免疫和功能。