Department of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Japan.
Division of Biological Information Technology, Joint Research Center for Human Retrovirus Infection, Kagoshima University, Japan.
FEBS J. 2024 Mar;291(6):1119-1130. doi: 10.1111/febs.16980. Epub 2023 Oct 27.
During the replication of viral genomes, RNA viruses produce double-stranded RNA (dsRNA), through the activity of their RNA-dependent RNA polymerases (RdRps) as viral replication intermediates. Recognition of viral dsRNA by host pattern recognition receptors - such as retinoic acid-induced gene-I (RIG-I)-like receptors and Toll-like receptor 3 - triggers the production of interferon (IFN)-β via the activation of IFN regulatory factor (IRF)-3. It has been proposed that, during the replication of viral genomes, each of RIG-I and melanoma differentiation-associated gene 5 (MDA5) form homodimers for the efficient activation of a downstream signalling pathway in host cells. We previously reported that, in the non-neoplastic human hepatocyte line PH5CH8, the RdRp NS5B derived from hepatitis C virus (HCV) could induce IFN-β expression by its RdRp activity without the actual replication of viral genomes. However, the exact mechanism by which HCV NS5B produced IFN-β remained unknown. In the present study, we first showed that NS5B derived from another Flaviviridae family member, GB virus B (GBV-B), also possessed the ability to induce IFN-β in PH5CH8 cells. Similarly, HCV NS5B, but not its G317V mutant, which lacks RdRp activity, induced the dimerization of MDA5 and subsequently the activation of IRF-3. Interestingly, immunofluorescence analysis showed that HCV NS5B produced dsRNA. Like HCV NS5B, GBV-B NS5B also triggered the production of dsRNA and subsequently the dimerization of MDA5. Taken together, our results show that HCV NS5B triggers an MDA5-mediated innate immune response by producing dsRNA without the replication of viral genomes in human hepatocytes.
在病毒基因组复制过程中，RNA 病毒通过其 RNA 依赖性 RNA 聚合酶（RdRps）的活性产生双链 RNA（dsRNA），作为病毒复制中间体。宿主模式识别受体（如视黄酸诱导基因-I（RIG-I）样受体和 Toll 样受体 3）识别病毒 dsRNA 会触发干扰素（IFN）-β的产生通过干扰素调节因子（IRF）-3 的激活。有人提出，在病毒基因组复制过程中，RIG-I 和黑色素瘤分化相关基因 5（MDA5）各自形成同源二聚体，以便在宿主细胞中有效地激活下游信号通路。我们之前报道过，在非肿瘤性人肝细胞系 PH5CH8 中，来自丙型肝炎病毒（HCV）的 RdRp NS5B 可以通过其 RdRp 活性诱导 IFN-β表达，而无需病毒基因组的实际复制。然而，HCV NS5B 产生 IFN-β的确切机制仍不清楚。在本研究中，我们首先表明，来自另一种黄病毒科家族成员 GB 病毒 B（GBV-B）的 NS5B 也具有在 PH5CH8 细胞中诱导 IFN-β的能力。同样，HCV NS5B，但不是缺乏 RdRp 活性的 G317V 突变体，诱导 MDA5 二聚化，随后激活 IRF-3。有趣的是，免疫荧光分析表明 HCV NS5B 产生 dsRNA。与 HCV NS5B 一样，GBV-B NS5B 也触发 dsRNA 的产生，随后 MDA5 的二聚化。总之，我们的研究结果表明，HCV NS5B 通过产生 dsRNA 在人肝细胞中触发 MDA5 介导的先天免疫反应，而无需病毒基因组的复制。
