Replication-competent recombinant vesicular stomatitis virus encoding hepatitis C virus envelope proteins
Replication-competent recombinant vesicular stomatitis virus encoding hepatitis C virus envelope proteins
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DOI：
10.1128/jvi.00608-07
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发表时间：
2007-08-01
影响因子：
5.4
通讯作者：
Matsuura, Yoshiharu
中科院分区：
文献类型：
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作者：
Tani, Hideki;Komoda, Yasumasa;Matsuura, Yoshiharu
尽管已经开发出基因型 2a (HCVcc) 丙型肝炎病毒 (HCV) JFH1 克隆的体外复制，但尚未建立针对 la 和 1b 基因型（世界上最流行的病毒且对干扰素治疗具有抗性）的强大细胞培养系统。作为替代病毒系统，基于水泡性口炎病毒（VSV）和逆转录病毒，已开发出暂时携带 HCV 包膜蛋白的假型病毒。在此，我们开发了具有复制能力的重组VSV，其具有编码未修饰的HCV E1和E2蛋白的基因组，代替人细胞系中的VSV包膜蛋白(HCVrv)。 HCVrv 和在 293T 或 Huh7 细胞中产生的带有未修饰的 HCV 包膜蛋白 (HCVpv) 的假型 VSV 在 Huh7 细胞中表现出高感染性。在某些检查的细胞系中，感染性 HCVrv 的产生受到限制。此外，HCVrv（而非HCVpv）能够在Huh7细胞中增殖并形成病灶。 Huh7细胞被HCVpv和HCVrv感染可被抗hCD81和抗E2抗体以及来自慢性HCV患者的血清中和。 HCVrv 的感染性可被内质网 α-葡萄糖苷酶抑制剂 N-(n-壬基) 脱氧野尻霉素 (Nn-DNJ) 抑制，但不能被高尔基体甘露糖苷酶抑制剂脱氧甘露尻霉素抑制。 Nn-DNJ 处理会损害 Huh7 细胞中 HCVrv 病灶的形成。这些结果表明，本研究中开发的HCVrv可用于研究HCV包膜蛋白，不仅涉及进入过程中的生物学功能，而且还涉及其成熟步骤。
Although in vitro replication of the hepatitis C virus (HCV) JFH1 clone of genotype 2a (HCVcc) has been developed, a robust cell culture system for the la and 1b genotypes, which are the most prevalent viruses in the world and resistant to interferon therapy, has not yet been established. As a surrogate virus system, pseudotype viruses transiently bearing HCV envelope proteins based on the vesicular stomatitis virus (VSV) and retrovirus have been developed. Here, we have developed a replication-competent recombinant VSV with a genome encoding unmodified HCV El and E2 proteins in place of the VSV envelope protein (HCVrv) in human cell lines. HCVrv and a pseudotype VSV bearing the unmodified HCV envelope proteins (HCVpv) generated in 293T or Huh7 cells exhibited high infectivity in Huh7 cells. Generation of infectious HCVrv was limited in some cell lines examined. Furthermore, HCVrv but not HCVpv was able to propagate and form foci in Huh7 cells. The infection of Huh7 cells with HCVpv and HCVrv was neutralized by anti-hCD81 and anti-E2 antibodies and by sera from chronic HCV patients. The infectivity of HCVrv was inhibited by an endoplasmic reticulum alpha-glucosidase inhibitor, N-(n-nonyl) deoxynojirimycin (Nn-DNJ), but not by a Golgi mannosidase inhibitor, deoxymannojirimycin. Focus formation of HCVrv in Huh7 cells was impaired by Nn-DNJ treatment. These results indicate that the HCVrv developed in this study can be used to study HCV envelope proteins with respect to not only the biological functions in the entry process but also their maturation step.