An antifungal protein from Ginkgo biloba binds actin and can trigger cell death
An antifungal protein from Ginkgo biloba binds actin and can trigger cell death
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DOI：
10.1007/s00709-015-0876-4
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发表时间：
2016-07-01
期刊：
影响因子：
2.9
通讯作者：
Nick, Peter
中科院分区：
文献类型：
--
作者：
Gao, Ningning;Wadhwani, Parvesh;Nick, Peter
银杏素是从银杏种子中分离纯化的一种短链抗真菌蛋白。该蛋白的同源物可以在所有种子植物和异孢蕨类卷柏中检测到，并且在结构域结构、肽基序和特定的半胱氨酸特征方面是保守的。为了深入了解这些保守基序的细胞功能，我们在烟草BY-2细胞中表达了全长和截短的银杏黄酮的绿色荧光蛋白融合体。我们表明，信号肽赋予银杏黄酮的有效分泌。当这个信号肽被切割或掩蔽时，银杏黄酮结合并显现肌动蛋白细胞骨架。银杏黄酮的肌动蛋白结合活性是由信号肽下游的一个特异性亚结构域介导的，该亚结构域也可以在体外与肌动蛋白共组装。在此域的稳定过度表达，我们观察到一个特定的延迟有丝分裂前核定位表明肌动蛋白的动态性降低。为了阐明细胞对该亚结构域与肌动蛋白结合的反应，我们使用基于合成肽的化学工程，所述合成肽包括与细胞穿透肽BP 100缀合的肌动蛋白结合亚结构域的不同部分，并与罗丹明B作为荧光报告物。这种合成结构与肌动蛋白的结合有效地诱导了程序性细胞死亡。我们讨论了这些发现的工作模型，银杏黄酮可以激活肌动蛋白依赖性细胞死亡。
Ginkbilobin is a short antifungal protein that had been purified and cloned from the seeds of the living fossil Ginkgo biloba. Homologues of this protein can be detected in all seed plants and the heterosporic fern Selaginella and are conserved with respect to domain structures, peptide motifs, and specific cysteine signatures. To get insight into the cellular functions of these conserved motifs, we expressed green fluorescent protein fusions of full-length and truncated ginkbilobin in tobacco BY-2 cells. We show that the signal peptide confers efficient secretion of ginkbilobin. When this signal peptide is either cleaved or masked, ginkbilobin binds and visualizes the actin cytoskeleton. This actin-binding activity of ginkbilobin is mediated by a specific subdomain just downstream of the signal peptide, and this subdomain can also coassemble with actin in vitro. Upon stable overexpression of this domain, we observe a specific delay in premitotic nuclear positioning indicative of a reduced dynamicity of actin. To elucidate the cellular response to the binding of this subdomain to actin, we use chemical engineering based on synthetic peptides comprising different parts of the actin-binding subdomain conjugated with the cell-penetrating peptide BP100 and with rhodamine B as a fluorescent reporter. Binding of this synthetic construct to actin efficiently induces programmed cell death. We discuss these findings in terms of a working model, where ginkbilobin can activate actin-dependent cell death.