Study on infection mechanisms of hepatitis C virus

由于缺乏支持丙型肝炎病毒（HCV）复制的常规细胞培养系统，阻碍了对HCV感染机制的研究。为了克服这个问题，我们已经建立了一个敏感的细胞融合检测系统，并构建了假型水泡性口炎病毒（VSV）具有HCV包膜蛋白的病毒粒子的表面上，而不是自己的包膜G糖蛋白。由E1或E2包膜蛋白的胞外域和VSV G糖蛋白的跨膜和胞浆区组成的嵌合HCV E1和E2蛋白在细胞表面表达。细胞融合的诱导需要具有低pH依赖性的嵌合E1和E2蛋白。具有嵌合E1和E2蛋白的假型病毒表现出显着更高的易感性比单独具有任一糖蛋白。这些结果表明，HCV在感染过程中需要E1和E2蛋白，并进入靶细胞。 ...更多信息 T细胞通过内体途径。在内体中，E1或/和E2蛋白发生低pH依赖性构象变化，然后触发膜融合和核衣壳进入细胞质。用链霉蛋白酶、肝素酶或肝素酶处理HepG 2细胞降低了假型VSV的细胞融合活性和感染性水平，表明细胞表面的某些蛋白分子和糖胺聚糖在HCV感染中起重要作用。最近，人CD 81（hCD 81）已被证明是E2蛋白的结合受体。然而，表达hCD 81的小鼠细胞系和亲本细胞系之间的细胞融合活性和对假型VSV的易感性没有差异。这些结果提示，hCD 81本身不足以使HCV感染，可能需要另一种辅助因子，或者HCV感染可能以非hCD 81依赖的方式发生，本研究揭示的感染机制可能为进一步研究HCV细胞受体和开发丙型肝炎的抗病毒和治疗药物提供重要信息。少

Studies of infection mechanisms of hepatitis C virus (HCV) have been hampered by the lack of conventional cell culture system which support replication of HCV. To overcome this problem, we have established a sensitive cell fusion assay system and also constructed pseudotype vesicular stomatitis virus (VSV) possessing HCV envelope protein on the surface of the virion instead of its own envelope G glycoprotein. The chimeric HCV E1 and E2 proteins consisting of the ectodomain of E1 or E2 envelope proteins and the transmembrane and cytoplasmic domains of VSV G glycoprotein were expressed on the cell surface. The induction of cell fusion requires both of the chimeric E1 and E2 proteins with low pH-dependent manner. The pseudotype virus possessing both of the chimeric E1 and E2 proteins exhibited significantly higher susceptibility than that possessing either of the glycoproteins individually. These results suggest that HCV requires both E1 and E2 proteins in the infection and enters a targe … More t cell via an endosomal pathway. In the endosome, a low pH-dependent conformation change of the E1 or/and E2 proteins occurs, which then triggers membrane fusion and the entry of the nucleocapsid into the cytoplasm. Treatment of HepG2 cells with pronase, heparinase, or heparitinase reduced the levels of cell fusion activity and infectivity of the pseudotype VSV suggesting that certain protein molecules and glycosaminoglycans on the cell surface play an important role in the infection with HCV. Recently, human CD81 (hCD81) has been shown to be a binding receptor of the E2 protein. However, there was no difference in cell fusion activity and susceptibility to pseudotype VSV between the mouse cell line expressing hCD81 and the parental cell line. These results suggest that hCD81 atone is not sufficient to allow infection of HCV and that another cofactor(s) might be required or that HCV infection may occur in an hCD81-independent manner.The infection mechanisms revealed in this study might offer an important information for future studies on cellular receptors for HCV and for the development of prophylactics and therapeutics for hepatitis C. Less