Plasticity of the β-trefoil protein fold in the recognition and control of invertebrate predators and parasites by a fungal defence system.
Plasticity of the β-trefoil protein fold in the recognition and control of invertebrate predators and parasites by a fungal defence system.
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DOI：
10.1371/journal.ppat.1002706
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发表时间：
2012
期刊：
影响因子：
6.7
通讯作者：
Künzler M
中科院分区：
文献类型：
--
作者：
Schubert M;Bleuler-Martinez S;Butschi A;Wälti MA;Egloff P;Stutz K;Yan S;Collot M;Mallet JM;Wilson IB;Hengartner MO;Aebi M;Allain FH;Künzler M
区分自我和非自我是任何防御机制的先决条件；在天然防御中，这种区分通常是由识别非自我碳水化合物结构的凝集素介导的，因此依赖于对目标生物体碳水化合物结构具有不同特异性的大量宿主凝集素。最近，从真菌子实体中分离出的细胞质凝集素被证明在防御多细胞真菌抵抗捕食者和寄生虫方面发挥了作用。在这里，我们提出了一种新的子实体凝集素，CCL2，从墨帽蘑菇中提取。我们证明了凝集素对秀丽线虫和果蝇的毒性，并给出了它与三糖GlcNAcβ1，4[Fucα1，3]GlcNAc的核磁共振溶液结构，该三糖在体外具有很高的特异性和亲和力。结构表明，单体CCL2采用β-三叶折叠，并通过单一的、拓扑上新的碳水化合物结合部位识别三糖。CCl_2的定点突变和线虫抗凝集素突变体的鉴定表明，CCl_2的线虫毒性是通过与线虫糖蛋白的α1，3-岩藻糖基化N-糖核结构结合而介导的；用荧光标记的CCl_2取食表明这些靶向糖蛋白定位于线虫的肠道。由于已鉴定的糖表位是无脊椎动物特有的，而真菌中没有，我们的数据表明，子实体凝集素的防御功能是基于对非自身碳水化合物结构的特异性识别。CCL2特异性识别的三糖是花粉和昆虫毒液变应原的关键碳水化合物决定因素，这意味着这种特殊的糖表位是真菌防御和哺乳动物免疫系统的靶标。综上所述，我们的结果表明，一个共同的蛋白质折叠的可塑性如何通过一种天然的防御机制来帮助识别和控制拮抗剂，即凝集素对其配体的单价意味着一种新的凝集素介导的毒性机制。所有多细胞生物都已发展出防御捕食者、寄生虫和病原体的机制。作为一种常见的机制，动物、植物和真菌使用大量的碳水化合物结合蛋白(凝集素)来保护自己免受捕食和寄生。这种天然防御机制的成功关键取决于宿主凝集素对外来碳水化合物结构的特异性识别的多样性。在本研究中，我们使用核磁共振结构测定来证明这种多样性的一部分是由常见蛋白质折叠的可塑性造成的。已鉴定的真菌凝集素对线虫和昆虫具有毒性，采用共同的凝集素折叠，但在识别的外源碳水化合物结构、蛋白质上碳水化合物结合部位的数量和位置以及寡聚化程度方面具有显著的特异性和亲和力。由于真菌凝集素的体内靶标是无脊椎动物特有的，我们的结果可能被用来开发控制动物和人类寄生虫的新方法。
Discrimination between self and non-self is a prerequisite for any defence mechanism; in innate defence, this discrimination is often mediated by lectins recognizing non-self carbohydrate structures and so relies on an arsenal of host lectins with different specificities towards target organism carbohydrate structures. Recently, cytoplasmic lectins isolated from fungal fruiting bodies have been shown to play a role in the defence of multicellular fungi against predators and parasites. Here, we present a novel fruiting body lectin, CCL2, from the ink cap mushroom Coprinopsis cinerea. We demonstrate the toxicity of the lectin towards Caenorhabditis elegans and Drosophila melanogaster and present its NMR solution structure in complex with the trisaccharide, GlcNAcβ1,4[Fucα1,3]GlcNAc, to which it binds with high specificity and affinity in vitro. The structure reveals that the monomeric CCL2 adopts a β-trefoil fold and recognizes the trisaccharide by a single, topologically novel carbohydrate-binding site. Site-directed mutagenesis of CCL2 and identification of C. elegans mutants resistant to this lectin show that its nematotoxicity is mediated by binding to α1,3-fucosylated N-glycan core structures of nematode glycoproteins; feeding with fluorescently labeled CCL2 demonstrates that these target glycoproteins localize to the C. elegans intestine. Since the identified glycoepitope is characteristic for invertebrates but absent from fungi, our data show that the defence function of fruiting body lectins is based on the specific recognition of non-self carbohydrate structures. The trisaccharide specifically recognized by CCL2 is a key carbohydrate determinant of pollen and insect venom allergens implying this particular glycoepitope is targeted by both fungal defence and mammalian immune systems. In summary, our results demonstrate how the plasticity of a common protein fold can contribute to the recognition and control of antagonists by an innate defence mechanism, whereby the monovalency of the lectin for its ligand implies a novel mechanism of lectin-mediated toxicity. All multicellular organisms have developed mechanisms to defend themselves against predators, parasites and pathogens. As a common mechanism, animals, plants and fungi use a large arsenal of carbohydrate-binding proteins (lectins) to protect themselves from predation and parasitism. The success of this type of innate defence mechanism critically depends on the diversity of specific recognition of foreign carbohydrate structures by the host lectins. In this study, we use NMR structure determination to show that part of this diversity is created by the plasticity of common protein folds. The identified fungal lectin that is toxic to nematodes and insects, adopts a common lectin fold but is remarkable in terms of its specificity and affinity for the recognized foreign carbohydrate structure, the number and location of the carbohydrate binding sites on the protein and the degree of oligomerization. Since the identified in vivo target of the fungal lectin is characteristic for invertebrates, our results may be exploited to develop novel approaches for the control of animal and human parasites.
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