Highly permissive cell lines for subgenomic and genomic hepatitis C virus RNA replication
Highly permissive cell lines for subgenomic and genomic hepatitis C virus RNA replication
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DOI：
10.1128/jvi.76.24.13001-13014.2002
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发表时间：
2002-12-01
影响因子：
5.4
通讯作者：
Rice, CM
中科院分区：
文献类型：
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作者：
Blight, KJ;McKeating, JA;Rice, CM
丙型肝炎病毒（HCV）的复制似乎仅限于人肝癌细胞系Huh-7，表明这些细胞内存在有利的细胞环境。尽管HCV非结构蛋白的适应性突变通常增强Huh-7细胞中亚基因组复制子的复制能力，但只能在这些细胞的亚群中检测到复制。在这里，我们表明，自我复制的亚基因组RNA可以从Huh-7克隆消除长期治疗α干扰素（IFN-α）和治愈细胞的频率更高，可以支持亚基因组和全长HCV复制。治愈细胞系之一的增加的容许性使我们能够在RNA转染后早期容易地检测HCV RNA和抗原，从而消除了选择复制阳性细胞的需要。我们还表明，NS 5A中的单个氨基酸取代足以在大多数治愈细胞中建立HCV复制，并且亚型1b NS 5A的主要磷酸受体位点对HCV复制不是必需的。
Hepatitis C virus (HCV) replication appears to be restricted to the human hepatoma cell line Huh-7, indicating that a favorable cellular environment exists within these cells. Although adaptive mutations in the HCV nonstructural proteins typically enhance the replicative capacity of subgenomic replicons in Huh-7 cells, replication can only be detected in a subpopulation of these cells. Here we show that self-replicating subgenomic RNA could be eliminated from Huh-7 clones by prolonged treatment with alpha interferon (IFN-alpha) and that a higher frequency of cured cells could support both subgenomic and full-length HCV replication. The increased permissiveness of one of the cured cell lines allowed us to readily detect HCV RNA and antigens early after RNA transfection, eliminating the need for selection of replication-positive cells. We also demonstrate that a single amino acid substitution in NS5A is sufficient for establishing HCV replication in a majority of cured cells and that the major phosphate acceptor site of subtype 1b NS5A is not essential for HCV replication.