Filamentous fungi produce small cysteine-rich proteins with potent, specific antifungal activity, offering the potential to fight fungal infections that severely threaten human health and food safety and security. The genome of the citrus postharvest fungal pathogen Penicillium digitatum encodes one of these antifungal proteins, namely AfpB. Biotechnologically produced AfpB inhibited the growth of major pathogenic fungi at minimal concentrations, surprisingly including its parental fungus, and conferred protection to crop plants against fungal infections. This study reports an in-depth characterization of the AfpB mechanism of action, showing that it is a cell-penetrating protein that triggers a regulated cell death program in the target fungus. We prove the importance of AfpB interaction with the fungal cell wall to exert its killing activity, for which protein mannosylation is required. We also show that the potent activity of AfpB correlates with its rapid and efficient uptake by fungal cells through an energy-dependent process. Once internalized, AfpB induces a transcriptional reprogramming signaled by reactive oxygen species that ends in cell death. Our data show that AfpB activates a self-injury program, suggesting that this protein has a biological function in the parental fungus beyond defense against competitors, presumably more related to regulation of the fungal population. Our results demonstrate that this protein is a potent antifungal that acts through various targets to kill fungal cells through a regulated process, making AfpB a promising compound for the development of novel biofungicides with multiple fields of application in crop and postharvest protection, food preservation, and medical therapies.

中文翻译：

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抗真菌蛋白 AfpB 在其亲本真菌 Penicillium digitatum 中诱导调节性细胞死亡。

丝状真菌产生富含半胱氨酸的小蛋白质，具有强效、特定的抗真菌活性，有可能对抗严重威胁人类健康和食品安全的真菌感染。柑橘采后真菌病原体指状青霉的基因组编码这些抗真菌蛋白之一，即 AfpB。生物技术生产的 AfpB 在最低浓度下抑制主要病原真菌的生长，令人惊讶的是包括其亲本真菌，并赋予作物植物免受真菌感染的保护。该研究报告了对 AfpB 作用机制的深入表征，表明它是一种细胞穿透蛋白，可在目标真菌中触发受调节的细胞死亡程序。我们证明了 AfpB 与真菌细胞壁相互作用以发挥其杀伤活性的重要性，为此需要蛋白质甘露糖基化。我们还表明，AfpB 的有效活性与其通过能量依赖过程被真菌细胞快速有效地吸收有关。一旦内化，AfpB 诱导由活性氧发出信号的转录重编程，最终导致细胞死亡。我们的数据显示 AfpB 激活了一个自伤程序，这表明这种蛋白质在亲本真菌中具有超越防御竞争者的生物学功能，可能与真菌种群的调节更相关。我们的研究结果表明，这种蛋白质是一种有效的抗真菌剂，可通过各种靶点通过受调控的过程杀死真菌细胞，从而使 AfpB 成为开发新型生物杀真菌剂的有前途的化合物，在作物和收获后保护、食品保鲜和医学疗法。表明这种蛋白质在亲本真菌中具有超越对抗竞争者防御的生物学功能，可能与真菌种群的调节更相关。我们的研究结果表明，这种蛋白质是一种有效的抗真菌剂，可通过各种靶点通过受调控的过程杀死真菌细胞，从而使 AfpB 成为开发新型生物杀真菌剂的有前途的化合物，在作物和收获后保护、食品保鲜和医学疗法。表明这种蛋白质在亲本真菌中具有超越对抗竞争者防御的生物学功能，可能与真菌种群的调节更相关。我们的研究结果表明，这种蛋白质是一种有效的抗真菌剂，可通过各种靶点通过受调控的过程杀死真菌细胞，从而使 AfpB 成为开发新型生物杀真菌剂的有前途的化合物，在作物和收获后保护、食品保鲜和医学疗法。