Comprehensive analysis of hepatitis Cvirus protein that disturb host interferon system

在7种丙型肝炎病毒（HCV）蛋白（核心、NS 2、NS 3、NS 4A、NS 4 B、NS 5A和NS 5 B）中，只有病毒RNA依赖性RNA聚合酶NS 5 B激活干扰素（IFN）-β启动子，而没有模板/引物结合的RNA依赖性RNA聚合酶活性的突变体NS 5 B不激活IFN-β启动子。通过NS 5 B激活IFN-β启动子需要正调控结构域III，即IFN调节因子（IRF）-3的结合序列。此外，IRF-3被NS 5 B磷酸化。通过小干扰RNA抑制Toll样受体（TLR）3表达和表达显性阴性形式的Toll/IL-1受体结构域-含有衔接子诱导IFN-β（TRIF）均显著降低了NS 5 B诱导的IFN-β活化。在其他六种HCV蛋白中，NS 4A、NS 4 B和NS 5A有效地抑制这种激活。HCV NS 5 B可能通过TLR 3/TRIF和dsRNA的合成而激活宿主先天免疫系统。 ...更多信息 通过NS 5 B阻断IFN-β诱导，这可能有助于这种病毒的持久性。p53可能在细胞对HCV的先天防御中起关键作用。在Huh 7细胞中，当p53表达被敲低时，观察到HCV RNA复制和病毒蛋白表达的显著更高水平。此外，IFN治疗在抑制p53敲低（p53 kd）Huh 7细胞中HCV RNA复制方面不太有效。事实上，IFN刺激的反应元件（ISRE）的激活和IFN刺激的基因（ISG）的诱导在p53 kd Huh 7细胞中显著减弱，并且发现p53与IFN调节因子（IRF）9直接相互作用。这些结果强调了肿瘤抑制蛋白p53在细胞抗HCV免疫中的潜在作用，并可能具有治疗意义。双链RNA激活蛋白激酶（PKR）是ISG之一。我们建立PKR敲低Huh 7细胞使用RNA干扰和PKR对HCV复制的影响进行了研究。在稳定的PKRkd细胞中，HCV复制高于对照细胞。此外，稳定的PKRkd细胞分泌的HCV颗粒明显多于对照细胞。通过在两种细胞中加入IFN-α抑制了HCV的复制，甚至10 U/ml的IFN-α在两种细胞中抑制HCV的复制超过98%。PKR在先天状态下抑制HCV复制中起重要作用，但在IFN治疗中不是必需的。2 '-5'-寡腺苷酸合成酶1（OAS-1）是ISG之一。我们评估了OAS-1单核苷酸多态性（SNPs）及其与慢性HCV感染分期的关系。选择OAS-1的6个SNPs，并在409例日本慢性HCV感染患者中进行检测。在OAS-1编码序列的外显子3处具有SNP的A/A、A/G和G//G基因型的患者患有较高血清丙氨酸转氨酶、较高程度肝纤维化和较高存在肝硬化的风险呈梯度增加。此外，具有G等位基因的OAS-1抑制病毒复制的能力低于具有A等位基因的OAS-1。总之，OAS-1编码序列外显子3的SNP与日本HCV感染患者的疾病进展相关。少

Among 7 hepatitis C virus (HCV) proteins(core, NS2, NS3, NS4A, NS4B, NS5A, and NS5B), only NS5B, a viral RNA-dependent RNA polymerase, activated the interferon (IFN)-beta promoter However, mutant NS5B without RNA dependent RNA polymerase activity of template/primer association did not activate the IFN-beta promoter. Activation of the IFN-beta promoter by NS5Brequired the positive regulatory domain III, a binding sequence for IFN regulatory factor (IRF)-3. Moreover IRF-3 was phosphorylated by NS5B. Both inhibition of Toll-like receptor (TLR)3 expression by small interfering RNA and expression of the dominant negative form of Toll/IL-1 receptor domain- containing adapter inducing IFN-beta (TRIF) significantly reduced NS5B-induced activation of IFN-beta. Of the six other HCV proteins, NS4A, NS4B, and NS5A efficiently inhibited this activation. HCV NS5B is a potent activator of the host innate immune system, possibly through TLR3/TRIF and synthesis of dsRNA Meanwhile, NS4A, NS4B, and NS5A … More block IFN-beta induction by NS5B, which may contribute toward the persistence of this virus.p53 could have a crucial role in the cellular innate defense against HCV. Significantly higher levels of HCV RNA replication and viral protein expression in the Huh7 cells were observed when their p53 expressions were knocked down. Moreover, IFN treatment was less effective in inhibiting the HCV RNA replication in the p53-knocked-down (p53kd) Huh7 cells. In fact, the activation of the IFN-stimulated response elements (ISRE) and the induction of IFN-stimulated genes (ISGs) were significantly attenuated in the p53kd Huh7 cells and p53 was found to directly interact with IFN regulatory factor (IRF)9. These observations underscore the potential contributions of the tumor suppresser p53 in cellular antiviral immunity against HCV with possible therapeutic implications.Double-stranded-RNA activated protein kinase (PKR) is one of ISGs. We established PKR knockdown Huh7 cells using RNA interference and investigated the effect of PKR against HCV replication. In stable PKRkd cells, HCV replication was higher than that of control cells. Furthermore, stable PKRkd cells secreted significantly more HCV particles than did control cells. The replication of HCV was suppressed by the addition of IFN-alpha in both cells, and even 10 U/ml of IFN-alpha suppressed the replication of HCV more than 98% in both cells. PKR plays an important role in suppressing HCV replication in an innate state, but is not essential in IFN therapy.2'-5'-oligoadenylate syntetase 1(OAS-1) is one of ISGs. We evaluated single nucleotide polymorphisms (SNPs) of OAS-1 and its relationship with stage of chronic HCV infection. 6 SNPs of OAS-1 were selected and examined in 409 Japanese patients with chronic HCV infection. Patients with genotypes A/A, A/G, and G//G of a SNP of OAS-1 at the exon 3 of its coding sequence were at gradient increased risks of suffering from higher serum alanine aminotransferase, higher degree of liver fibrosis, and higher presence of liver cirrhosis. Moreover, OAS-1 with G allele showed lower ability of inhibiting virus replication compared to OAS-1 with A allele. In conclusion, the SNP of OAS-1 at the exon 3 of its coding sequence was associated with progression of disease in Japanese patients with HCV infection. Less