Two conserved histidines (His490 and His621) on the E2 glycoprotein of hepatitis C virus are critical for CD81-mediated cell entry.
Two conserved histidines (His490 and His621) on the E2 glycoprotein of hepatitis C virus are critical for CD81-mediated cell entry.
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DOI：
10.1099/vir.0.000091
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发表时间：
2015-06
期刊：
影响因子：
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通讯作者：
Zhao-Ling Qin;He-peng Ju;Ting-ting Gao;Wen-bo Wang;H. Ren;P. Zhao;Z. Qi
中科院分区：
文献类型：
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作者：
Zhao-Ling Qin;He-peng Ju;Ting-ting Gao;Wen-bo Wang;H. Ren;P. Zhao;Z. Qi
丙型肝炎病毒 (HCV) 进入是一个连续的多步骤过程，包括受体相互作用和 pH 依赖性膜融合。病毒包膜蛋白上特定且保守的组氨酸残基参与大多数 pH 诱导的病毒进入。就 HCV 而言，E1 和 E2 蛋白上的一些保守组氨酸已在 HCV 假型颗粒 (HCVpp) 系统中进行了研究。然而，由于这两个系统强调的病毒生命周期的不同方面，这些组氨酸在细胞培养衍生的 HCV 颗粒 (HCVcc) 系统中的作用仍不清楚。在本研究中，在基于 JFH-1 的 HCVcc 中使用丙氨酸替代评估了两个保守组氨酸（His490 和 His621，分别位于 E2 的 II 和 III 域）在 HCV 感染中的作用。尽管 H490A 突变体的初始 RNA 复制、蛋白质表达和组装效率与 WT 病毒相当，但其感染性却下降。 H621A突变体不影响病毒蛋白表达，但没有表现出明显的感染性；与WT病毒相比，培养上清液中的核心蛋白较少，表明病毒组装存在部分缺陷。使用酶免疫测定进一步评估了两种突变体 E2 的 HCV 受体 CD81 结合能力。 H490A-E2的CD81结合活性降低，并且H621A-E2不能与CD81结合。这些数据揭示了 His490 和 His621 在 HCV 感染中发挥的关键作用，特别是在细胞进入过程中 CD81 结合期间。这些结果也有助于对参与 pH 依赖性 HCV 进入的组氨酸进行机械鉴定。
Hepatitis C virus (HCV) entry is a sequential and multi-step process that includes receptor interactions followed by pH-dependent membrane fusion. Specific and conserved histidine residues on the viral envelope proteins are involved in most pH-induced virus entries. In the case of HCV, some conserved histidines on the E1 and E2 proteins have been investigated in HCV pseudotype particle (HCVpp) systems. However, the roles of these histidines in cell-culture-derived HCV particle (HCVcc) systems remain unclear due to the different aspects of the viral life cycle emphasized by the two systems. In this study, the role of two conserved histidines (His490 and His621, located in domains II and III of E2, respectively) in HCV infection was evaluated in the context of JFH-1-based HCVcc using alanine substitutions. The infectivity of the H490A mutant decreased in spite of comparable initial RNA replication, protein expression and assembly efficiency as WT virus. The H621A mutant did not affect viral protein expression, but exhibited no obvious infectivity; there were fewer core proteins in the culture supernatant compared with WT virus, indicating the partially deficient virus assembly. The HCV receptor CD81-binding ability of the two mutant E2s was assessed further using enzyme immunoassays. The CD81-binding activity of H490A-E2 was reduced, and H621A-E2 was unable to bind to CD81. These data revealed the crucial role played by His490 and His621 in HCV infection, particularly during CD81 binding in cell entry. These results also contributed to the mechanical identification of the histidines involved in pH-dependent HCV entry.