Tipping the Balance: <em>Sclerotinia sclerotiorum</em> Secreted Oxalic Acid Suppresses Host Defenses by Manipulating the Host Redox Environment

Sclerotinia sclerotiorum is a necrotrophic ascomycete fungus with an extremely broad host range. This pathogen produces the non-specific phytotoxin and key pathogenicity factor, oxalic acid (OA). Our recent work indicated that this fungus and more specifically OA, can induce apoptotic-like programmed cell death (PCD) in plant hosts, this induction of PCD and disease requires generation of reactive oxygen species (ROS) in the host, a process triggered by fungal secreted OA. Conversely, during the initial stages of infection, OA also dampens the plant oxidative burst, an early host response generally associated with plant defense. This scenario presents a challenge regarding the mechanistic details of OA function; as OA both suppresses and induces host ROS during the compatible interaction. In the present study we generated transgenic plants expressing a redox-regulated GFP reporter. Results show that initially, Sclerotinia (via OA) generates a reducing environment in host cells that suppress host defense responses including the oxidative burst and callose deposition, akin to compatible biotrophic pathogens. Once infection is established however, this necrotroph induces the generation of plant ROS leading to PCD of host tissue, the result of which is of direct benefit to the pathogen. In contrast, a non-pathogenic OA-deficient mutant failed to alter host redox status. The mutant produced hypersensitive response-like features following host inoculation, including ROS induction, callose formation, restricted growth and cell death. These results indicate active recognition of the mutant and further point to suppression of defenses by the wild type necrotrophic fungus. Chemical reduction of host cells with dithiothreitol (DTT) or potassium oxalate (KOA) restored the ability of this mutant to cause disease. Thus, Sclerotinia uses a novel strategy involving regulation of host redox status to establish infection. These results address a long-standing issue involving the ability of OA to both inhibit and promote ROS to achieve pathogenic success.核盘菌（Sclerotinia sclerotiorum）是一种寄主范围极广的死体营养型子囊真菌。该病原菌可产生非特异性植物毒素与关键致病因子——草酸（OA）。我们近期的研究表明，该真菌，更确切地说是其分泌的OA，可在植物宿主中诱导类凋亡程序性细胞死亡（PCD）；而这类PCD诱导与病害发生均依赖宿主活性氧（ROS）的产生，这一过程由真菌分泌的OA触发。与之相反，在侵染初期，OA还会抑制植物的氧化爆发——这是一种通常与植物防御相关的早期宿主应答。这种现象为解析OA功能的分子机制带来了挑战：因为在亲和互作过程中，OA既可抑制又可诱导宿主ROS的产生。本研究构建了表达氧化还原调控型绿色荧光蛋白（GFP）报告系统的转基因植物。实验结果显示，侵染初期，核盘菌通过OA在宿主细胞内营造还原环境，抑制包括氧化爆发与胼胝质沉积在内的宿主防御反应，这一特征与亲和性活体营养型病原菌类似。然而一旦侵染成功建立，该死体营养型真菌会诱导植物ROS的产生，进而引发宿主组织的PCD，而这一结果对病原菌自身极为有利。与之形成对比的是，一株非致病性的OA缺失突变体无法改变宿主的氧化还原状态。该突变体在接种宿主后会引发类过敏反应特征，包括ROS诱导、胼胝质形成、病原菌生长受限制以及细胞死亡。这些结果表明宿主可主动识别该突变体，进一步印证了野生型死体营养型真菌对宿主防御的抑制作用。用二硫苏糖醇（DTT）或草酸钾（KOA）对宿主细胞进行化学还原处理，可恢复该突变体的致病能力。综上，核盘菌采用了一种调控宿主氧化还原状态以成功建立侵染的全新策略。本研究结果解决了一个长期存在的科学问题：即OA如何通过同时抑制与促进ROS产生来实现致病成功。