Plant pathogens cause disease through secreted effector proteins, which act to modulate host physiology and promote infection. Typically, the sequences of effectors provide little functional information and further targeted experimentation is required. Here, we utilised a structure/function approach to study SnTox3, an effector from the necrotrophic fungal pathogen Parastagonospora nodorum, which causes cell death in wheat-lines carrying the sensitivity gene Snn3. We developed a workflow for the production of SnTox3 in a heterologous host that enabled crystal structure determination. We show this approach can be successfully applied to effectors from other pathogenic fungi. Complementing this, an in-silico study uncovered the prevalence of an expanded subclass of effectors from fungi. The β-barrel fold of SnTox3 is a novel fold among fungal effectors. We demonstrate that SnTox3 is a pre-pro-protein and that the protease Kex2 removes the pro-domain. Our in-silico studies suggest that Kex2-processed pro-domain (designated here as K2PP) effectors are common in fungi, and we demonstrate this experimentally for effectors from Fusarium oxysporum f sp. lycopersici. We propose that K2PP effectors are highly prevalent among fungal effectors. The identification and classification of K2PP effectors has broad implications for the approaches used to study their function in fungal virulence.

中文翻译：

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坏死性真菌Nostumparaporagonospora nodorum的SnTox3的晶体结构揭示了独特的效应子折叠和对真菌效应子的Kex2蛋白酶加工的见解。

植物病原体通过分泌的效应蛋白引起疾病，该蛋白调节宿主生理并促进感染。通常，效应子序列提供的功能信息很少，并且需要进一步的靶向实验。在这里，我们利用结构/功能方法来研究SnTox3，这是一种来自坏死性真菌病原体Parastagonospora nodorum的效应物，它导致携带敏感基因Snn3的小麦品系中的细胞死亡。我们开发了在异源宿主中生产SnTox3的工作流程，该流程可确定晶体结构。我们显示这种方法可以成功地应用于其他致病真菌的效应子。与此相辅相成的是，一项计算机模拟研究发现了来自真菌的效应子扩展类的普遍性。SnTox3的β-桶状折叠是真菌效应子中的新型折叠。我们证明，SnTox3是前原蛋白，蛋白酶Kex2删除了前域。我们的计算机模拟研究表明，经过Kex2处理的结构域（在这里称为K2PP）效应子在真菌中很常见，我们通过实验证明了尖镰孢（Fusarium oxysporum fsp）的效应子。lycopersici。我们提出，K2PP效应子在真菌效应子中非常普遍。K2PP效应子的鉴定和分类对研究其在真菌毒力中的功能的方法具有广泛的意义。并且我们通过实验证明了尖孢镰刀菌f sp的效应子。lycopersici。我们提出，K2PP效应子在真菌效应子中非常普遍。K2PP效应子的鉴定和分类对研究其在真菌毒力中的功能的方法具有广泛的意义。并且我们通过实验证明了尖孢镰刀菌f sp的效应子。lycopersici。我们提出，K2PP效应子在真菌效应子中非常普遍。K2PP效应子的鉴定和分类对研究其在真菌毒力中的功能的方法具有广泛的意义。