Identification of host factors required for HCV-IRES activity and their physiological roles in liver disease

背景与目的：丙型肝炎病毒（HCV）的翻译是由内部核糖体进入位点（IRES）介导的。许多翻译起始因子与HCV IRES相互作用;然而，这些因子对HCV翻译和复制的功能相关性尚未阐明。方法：使用两种不同的亚基因组HCV复制子，NNeo/3- 5 B和MH 14 C。NNeo/3- 5 B由具有标准结构的HCV复制子组成，其中NS 3-NS 5由脑心肌炎（EMCV）IRES翻译。MH 14 C是一种具有修饰结构的HCV复制子，其中EMCV IRES被HCV IRES取代。在14个被反义寡核苷酸或siRNA抑制的翻译起始因子中，La蛋白、PTB、eIF 3 p170、eIF 2 γ和PSMA 7影响HCV IRES活性和MH 14 C的复制。然而，这些因素并不影响EMCV IRES活性和NNeo/3- 5 B的复制。37例慢性丙型肝炎（CH-C）患者肝组织中La蛋白、eIF 2 γ和PSMA 7与HCV RNA显著相关。使用cDNA微阵列的基因表达谱显示，这些起始因子在CH-C中被诱导，并聚集在一组代表炎症、DNA修复和应激反应的基因中。HCV编码蛋白之一NS 5A激活La蛋白启动子，NS 5A中的干扰素敏感性决定区负责激活。结论：HCV复制高度依赖于CH-C组织病变中诱导的翻译起始因子。HCV NS 5A蛋白对La蛋白的诱导作用可能支持HCV的复制和持续感染。

Background and aim : Translation of the hepatitis C virus (HCV) is mediated by an internal ribosome entry site (IRES). Many translation initiation factors interact with HCV IRES; however, the functional relevance of these factors for the translation and replication of HCV has not been clarified. Methods : Two different subgenomic HCV replicons, NNeo/3-5B and MH14C, were used. NNeo/3-5B consists of an HCV replicon with a standard structure in which NS3-NS5 was translated by encephalomyocarditis (EMCV) IRES. MH14C is an HCV replicon with a modified structure in which EMCV IRES was replaced with HCV IRES. Of 14 translation initiation factors suppressed by antisense oligodeoxynucleotides or siRNA, La protein, PTB, eIF3 p170, eIF2 gamma, and PSMA 7 affected HCV IRES activity and the replication of MH14C. However, these factors did not affect EMCV IRES activity and the replication of NNeo/3-5B. La protein, eIF2 gamma, and PSMA7 were significantly correlated with HCV RNA in the liver of 37 patients with chronic hepatitis C (CH-C). Gene expression profiling using a cDNA microarray revealed these initiation factors were induced in CH-C and clustered in a group of genes representing inflammation, DNA repair and stress responses. One of the HCV coding proteins, NS5A, activated the La protein promoter and the interferon sensitivity-determining region in NS5A was responsible for the activation. Conclusions: HCV replication was highly dependent on translation initiation factors that were induced in tissue lesions of CH-C. The induction of La protein by the HCV protein NS5A might support the replication and persistent infection of HCV.