Changes in ganglioside content affect the binding of clostridium perfringens epsilon-toxin to detergent-resistant membranes of Madin-Darby canine kidney cells
Changes in ganglioside content affect the binding of clostridium perfringens epsilon-toxin to detergent-resistant membranes of Madin-Darby canine kidney cells
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DOI：
10.1111/j.1348-0421.2005.tb03726.x
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发表时间：
2005-01-01
影响因子：
2.6
通讯作者：
Miyata, S
中科院分区：
文献类型：
--
作者：
Shimamoto, S;Tamai, E;Miyata, S
产气荚膜梭菌的Epsilon毒素(ET)在有蹄类动物中引起致命性肠毒素血症，先前被证明结合并在MDCK细胞的抗洗涤剂膜(DRM)内形成七聚体孔道。去除胆固醇还可以降低ET的细胞毒性和它的七聚体。在这项研究中，我们研究了鞘磷脂的变化，MDCK细胞的其他DRM成分，对细胞对ET易感性的影响。神经鞘糖脂合成的抑制剂伏马菌素B1和PDMP可增加敏感性，而神经鞘磷脂合成抑制剂D609则有相反的作用。外源性神经节苷脂G(M1)显著降低ET结合、七聚化和细胞毒性。这些作用不是由于ET与G(M1)结合或ET变性所致。我们还发现，ET对MDCK细胞的细胞毒作用随着培养时间的增加而降低。与观察到的长期培养细胞的耐药性相一致，神经节苷脂的主要成分G(M3)增加，唾液酸酶处理增加了细胞的敏感性。这些结果表明，DRM中膜锚定的G(M3)唾液酸抑制了ET的结合，从而阻止了ET的七聚化和细胞死亡。该菌产生的唾液酸酶有助于ET靶向MDCK细胞。
Epsilon-toxin (ET) of Clostridium perfringens, which causes fatal enterotoxemia in ungulates, was previously shown to bind to and form a heptameric pore within the detergent-resistant membranes (DRMs) of MDCK cells. Depletion of cholesterol has also been shown to decrease the cytotoxicity of ET and its heptamerization. In this study, we investigated the effects of changes in sphingolipids, other DRM components of MDCK cells, on the cells' susceptibility to ET. Treatment with fumonisin B1 and PDMP, inhibitors of sphingolipid and glycosphingolipid syntheses, respectively, increased the susceptibility, while D609, a sphingomyelin synthesis inhibitor, had the opposite effect. The exogenous addition of ganglioside G(M1) dramatically decreased the ET binding, heptamerization and cytotoxicity. These effects were shown not to be due to ET binding to G(M1) or to denaturation of ET. We also found that the ET cytotoxicity towards MDCK cells decreased with an increase in culture time. In accordance with the resistance observed for prolonged cultured cells, G(M3), a major ganglioside component, increased and sialidase treatment increased their susceptibility. These results suggest that membrane-anchored sialic acid of G(M3) within DRMs inhibits ET binding, leading to prevention of the heptamerization of ET and cell death. It is also suggested that sialidase produced by this organism aids the targeting of ET to MDCK cells.