The Tight Junction-Associated Protein Occludin Is Required for a Postbinding Step in Hepatitis C Virus Entry and Infection
The Tight Junction-Associated Protein Occludin Is Required for a Postbinding Step in Hepatitis C Virus Entry and Infection
复制标题
DOI：
10.1128/jvi.00038-09
复制
发表时间：
2009-08-15
影响因子：
5.4
通讯作者：
Majano, Pedro L.
中科院分区：
文献类型：
--
作者：
Benedicto, Ignacio;Molina-Jimenez, Francisca;Majano, Pedro L.
丙型肝炎病毒（HCV）进入肝细胞的确切机制尚不清楚。然而，几种细胞表面蛋白已被鉴定为该病毒的进入因子。在这些分子中，紧密连接（TJ）组分claudin-1被认为是HCV进入所需的辅助受体。最近，我们已经证明，HCV包膜糖蛋白（HCVgp）促进结构和功能的TJ改变。此外，我们已经表明，病毒E2糖蛋白和occludin，另一个TJ相关蛋白，细胞内的相互作用，可能是TJ蛋白的错误定位的原因。在此，我们证明，通过使用细胞培养衍生的HCV颗粒（HCV-DNA），封闭蛋白表达的干扰显着降低HCV感染。此外，我们的研究结果与HCV假型颗粒表明，occludin，而不是其他TJ相关蛋白，如连接粘附分子A或zonula occludens蛋白1，所需的HCV进入。使用HCCP，我们证明了闭合蛋白在HCV与靶细胞的初始附着中没有发挥重要作用。表面蛋白标记实验表明，表达水平和细胞表面定位的HCV（共）受体CD 81，清道夫受体类B I型，和claudin-1不受影响后occludin敲低。此外，免疫荧光共聚焦分析表明，occludin干扰不影响亚细胞分布的HCV（共）受体分析。然而，HCVgp融合相关的事件改变后occludin沉默。总之，我们认为闭合蛋白在HCV感染中起着重要作用，并可能影响晚期进入事件。这一观察结果可能为HCV感染及其相关发病机制提供新的见解。
The precise mechanisms regulating hepatitis C virus (HCV) entry into hepatic cells remain unknown. However, several cell surface proteins have been identified as entry factors for this virus. Of these molecules, claudin-1, a tight junction (TJ) component, is considered a coreceptor required for HCV entry. Recently, we have demonstrated that HCV envelope glycoproteins (HCVgp) promote structural and functional TJ alterations. Additionally, we have shown that the intracellular interaction between viral E2 glycoprotein and occludin, another TJ-associated protein, could be the cause of the mislocalization of TJ proteins. Herein we demonstrated, by using cell culture-derived HCV particles (HCVcc), that interference of occludin expression markedly reduced HCV infection. Furthermore, our results with HCV pseudotyped particles indicated that occludin, but not other TJ-associated proteins, such as junctional adhesion molecule A or zonula occludens protein 1, was required for HCV entry. Using HCVcc, we demonstrated that occludin did not play an essential role in the initial attachment of HCV to target cells. Surface protein labeling experiments showed that both expression levels and cell surface localization of HCV (co) receptors CD81, scavenger receptor class B type I, and claudin-1 were not affected upon occludin knockdown. In addition, immunofluorescence confocal analysis showed that occludin interference did not affect subcellular distribution of the HCV (co) receptors analyzed. However, HCVgp fusion-associated events were altered after occludin silencing. In summary, we propose that occludin plays an essential role in HCV infection and probably affects late entry events. This observation may provide new insights into HCV infection and related pathogenesis.