Clostridium Perfringens Epsilon Toxin Binds to Membrane Lipids and Its Cytotoxic Action Depends on Sulfatide
Clostridium Perfringens Epsilon Toxin Binds to Membrane Lipids and Its Cytotoxic Action Depends on Sulfatide
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DOI：
10.1371/journal.pone.0140321
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发表时间：
2015-10-09
期刊：
影响因子：
3.7
通讯作者：
Blasi, Juan
中科院分区：
文献类型：
--
作者：
Gil, Carles;Dorca-Arevalo, Jonatan;Blasi, Juan
Etx 毒素 (Etx) 是 B 型和 D 型产气荚膜梭菌产生的主要致命毒素之一，是动物（主要是绵羊和山羊）致命性肠毒血症的病原体。 Etx 被合成为非活性原毒素形式 (proEtx)，在蛋白水解激活后变得活跃。 Etx 通过在选定的细胞靶标的质膜上形成孔而表现出细胞毒性作用，其中 Etx 由于特定受体的存在而特异性结合。然而，Etx 宿主受体的身份和性质仍然存在争议。在本研究中，分析了 Etx 与大鼠脑突触体富集组分（P2 组分）和 MDCK 细胞质膜制剂的膜脂之间的相互作用。我们的研究结果表明，通过蛋白质-脂质叠加测定评估，Etx 和 proEtx 均与从两个不同模型的脂筏中提取的脂质结合。脂筏是富含胆固醇和鞘脂的膜微域。检测到 proEtx 与脑硫脂、磷脂酰丝氨酸、磷脂酰肌醇 (3)-磷酸和磷脂酰肌醇 (5)-磷酸的结合。通过硫酸酯酶处理去除硫酸基团导致 Etx 诱导的细胞毒性显着降低，但不会导致 proEtx-GFP 与 MDCK 细胞的结合或寡聚体形成的显着变化，这表明硫苷脂在筏中孔形成中发挥作用，但在与靶细胞膜结合的毒素中没有作用。这些结果首次显示了 Etx 与宿主组织膜脂之间的相互作用，并指出硫苷脂在产气荚膜梭菌毒素病理生理学中的主要作用。
Epsilon toxin (Etx) is one of the major lethal toxins produced by Clostridium perfringens types B and D, being the causal agent of fatal enterotoxemia in animals, mainly sheep and goats. Etx is synthesized as a non-active prototoxin form (proEtx) that becomes active upon proteolytic activation. Etx exhibits a cytotoxic effect through the formation of a pore in the plasma membrane of selected cell targets where Etx specifically binds due to the presence of specific receptors. However, the identity and nature of host receptors of Etx remain a matter of controversy. In the present study, the interactions between Etx and membrane lipids from the synaptosome-enriched fraction from rat brain (P2 fraction) and MDCK cell plasma membrane preparations were analyzed. Our findings show that both Etx and proEtx bind to lipids extracted from lipid rafts from the two different models as assessed by protein-lipid overlay assay. Lipid rafts are membrane microdomains enriched in cholesterol and sphingolipids. Binding of proEtx to sulfatide, phosphatidylserine, phosphatidylinositol (3)-phosphate and phosphatidylinositol (5)-phosphate was detected. Removal of the sulphate groups via sulfatase treatment led to a dramatic decrease in Etx-induced cytotoxicity, but not in proEtx-GFP binding to MDCK cells or a significant shift in oligomer formation, pointing to a role of sulfatide in pore formation in rafts but not in toxin binding to the target cell membrane. These results show for the first time the interaction between Etx and membrane lipids from host tissue and point to a major role for sulfatides in C. perfringens epsilon toxin pathophysiology.