In this study, Schumann et al. identify the penta-EF-hand protein PEF1 of the genetic model fungus Neurospora crassa as part of the cellular response to different types of membrane injury... Plasma membrane damage commonly occurs during cellular growth and development. To counteract these potentially lethal injuries, membrane repair mechanisms have evolved, which promote the integrity of the lipid bilayer. Although the membrane of fungi is the target of important clinical drugs and agricultural fungicides, the molecular mechanisms which mediate membrane repair in these organisms remain elusive. Here we identify the penta-EF-hand protein PEF1 of the genetic model fungus Neurospora crassa as part of a cellular response mechanism against different types of membrane injury. Deletion of the pef1 gene in the wild type and different lysis-prone gene knockout mutants revealed a function of the protein in maintaining cell integrity during cell-cell fusion and in the presence of pore-forming drugs, such as the plant defense compound tomatine. By fluorescence and live-cell imaging we show that green fluorescent protein (GFP)-tagged PEF1 accumulates at the sites of membrane injury in a Ca2+-dependent manner. Site-directed mutagenesis identified Ca2+-binding domains essential for the spatial dynamics and function of the protein. In addition, the subcellular localization of PEF1 revealed that the syncytial fungal colony undergoes compartmentation in response to antifungal treatment. We propose that plasma membrane repair in fungi constitutes an additional line of defense against membrane-disturbing drugs, thereby expanding the current model of fungal drug resistance mechanisms.

中文翻译：

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粗糙脉孢菌中的 Penta-EF-Hand 蛋白 PEF1 可促进细胞间融合和对成孔药物反应期间质膜的完整性。


在这项研究中，舒曼等人。鉴定遗传模型真菌粗糙脉孢菌的五-EF-手蛋白 PEF1 作为细胞对不同类型膜损伤反应的一部分......质膜损伤通常发生在细胞生长和发育过程中。为了抵消这些潜在的致命伤害，膜修复机制已经进化，从而促进脂质双层的完整性。尽管真菌膜是重要的临床药物和农业杀菌剂的靶标，但介导这些生物体膜修复的分子机制仍然难以捉摸。在这里，我们鉴定了遗传模型真菌粗糙脉孢菌的五手蛋白 PEF1，作为针对不同类型膜损伤的细胞反应机制的一部分。野生型和不同易裂解基因敲除突变体中 pef1 基因的删除揭示了该蛋白质在细胞-细胞融合过程中以及在存在成孔药物（例如植物防御化合物番茄碱）的情况下维持细胞完整性的功能。通过荧光和活细胞成像，我们发现绿色荧光蛋白 (GFP) 标记的 PEF1 以 Ca2+ 依赖性方式在膜损伤部位积聚。定点诱变鉴定出对于蛋白质的空间动力学和功能至关重要的 Ca2+ 结合域。此外，PEF1 的亚细胞定位表明，合胞体真菌菌落响应抗真菌治疗而发生区室化。我们认为真菌中的质膜修复构成了针对膜干扰药物的额外防线，从而扩展了当前真​​菌耐药机制的模型。