Iwasaki Masaharu, Ng Cherie T, Cubitt Beatrice, de la Torre Juan C
Department of Immunology and Microbial Science, Scripps Research Institute, La Jolla, California, USA.
Department of Immunology and Microbial Science, Scripps Research Institute, La Jolla, California, USA
J Virol. 2016 Oct 28;90(22):10102-10112. doi: 10.1128/JVI.01303-16. Print 2016 Nov 15.
Several arenaviruses, chiefly Lassa virus (LASV), cause hemorrhagic fever disease in humans and pose serious public health concerns in their regions of endemicity. Moreover, mounting evidence indicates that the worldwide-distributed prototypic arenavirus, lymphocytic choriomeningitis virus (LCMV), is a neglected human pathogen of clinical significance. We have documented that a recombinant LCMV containing the glycoprotein (GPC) gene of LASV within the backbone of the immunosuppressive clone 13 (Cl-13) variant of the Armstrong strain of LCMV (rCl-13/LASV-GPC) exhibited Cl-13-like growth properties in cultured cells, but in contrast to Cl-13, rCl-13/LASV-GPC was unable to establish persistence in immunocompetent adult mice, which prevented its use for some in vivo experiments. Recently, V459K and K461G mutations within the GP2 cytoplasmic domain (CD) of rCl-13/LASV-GPC were shown to increase rCl-13/LASV-GPC infectivity in mice. Here, we generated rCl-13(GPC/VGKS) by introducing the corresponding revertant mutations K465V and G467K within GP2 of rCl-13 and we show that rCl-13(GPC/VGKS) was unable to persist in mice. K465V and G467K mutations did not affect GPC processing, virus RNA replication, or gene expression. In addition, rCl-13(GPC/VGKS) grew to high titers in cultured cell lines and in immunodeficient mice. Further analysis revealed that rCl-13(GPC/VGKS) infected fewer splenic plasmacytoid dendritic cells than rCl-13, yet the two viruses induced similar type I interferon responses in mice. Our findings have identified novel viral determinants of Cl-13 persistence and also revealed that virus GPC-host interactions yet to be elucidated critically contribute to Cl-13 persistence.
The prototypic arenavirus, lymphocytic choriomeningitis virus (LCMV), provides investigators with a superb experimental model system to investigate virus-host interactions. The Armstrong strain (ARM) of LCMV causes an acute infection, whereas its derivative, clone 13 (Cl-13), causes a persistent infection. Mutations F260L and K1079Q within GP1 and L polymerase, respectively, have been shown to play critical roles in Cl-13's ability to persist in mice. However, there is an overall lack of knowledge about other viral determinants required for Cl-13's persistence. Here, we report that mutations K465V and G467K within the cytoplasmic domain of Cl-13 GP2 resulted in a virus, rCl-13(GPC/VGKS), that failed to persist in mice despite exhibiting Cl-13 wild-type-like fitness in cultured cells and immunocompromised mice. This finding has uncovered novel viral determinants of viral persistence, and a detailed characterization of rCl-13(GPC/VGKS) can provide novel insights into the mechanisms underlying persistent viral infection.
几种沙粒病毒，主要是拉沙病毒（LASV），可导致人类出血热疾病，并在其流行地区引起严重的公共卫生问题。此外，越来越多的证据表明，全球分布的原型沙粒病毒——淋巴细胞性脉络丛脑膜炎病毒（LCMV）是一种被忽视的具有临床意义的人类病原体。我们已证明，一种重组LCMV，其在LCMV阿姆斯特朗株免疫抑制性克隆13（Cl-13）变体的主干内包含LASV的糖蛋白（GPC）基因（rCl-13/LASV-GPC），在培养细胞中表现出类似Cl-13的生长特性，但与Cl-13不同的是，rCl-13/LASV-GPC无法在免疫功能正常的成年小鼠中建立持续性感染，这使其无法用于某些体内实验。最近，rCl-13/LASV-GPC GP2胞质结构域（CD）内的V459K和K461G突变被证明可增加rCl-13/LASV-GPC在小鼠中的感染性。在此，我们通过在rCl-13的GP2中引入相应的回复突变K465V和G467K，构建了rCl-13(GPC/VGKS)，并表明rCl-13(GPC/VGKS)无法在小鼠中持续存在。K465V和G467K突变不影响GPC加工、病毒RNA复制或基因表达。此外，rCl-13(GPC/VGKS)在培养细胞系和免疫缺陷小鼠中生长至高滴度。进一步分析表明，rCl-13(GPC/VGKS)感染的脾脏浆细胞样树突状细胞比rCl-13少，但这两种病毒在小鼠中诱导的I型干扰素反应相似。我们的研究结果确定了Cl-13持续性的新病毒决定因素，还揭示了尚未阐明的病毒GPC与宿主的相互作用对Cl-13持续性起关键作用。
原型沙粒病毒——淋巴细胞性脉络丛脑膜炎病毒（LCMV）为研究人员提供了一个极好的实验模型系统，用于研究病毒与宿主的相互作用。LCMV的阿姆斯特朗株（ARM）引起急性感染，而其衍生物克隆13（Cl-13）则引起持续性感染。已证明GP1和L聚合酶内的F260L和K1079Q突变在Cl-13在小鼠中持续存在的能力中起关键作用。然而，对于Cl-13持续存在所需的其他病毒决定因素，总体上缺乏了解。在此，我们报告Cl-13 GP2胞质结构域内的K465V和G467K突变产生了一种病毒rCl-13(GPC/VGKS)，尽管其在培养细胞和免疫受损小鼠中表现出类似Cl-13野生型的适应性，但仍无法在小鼠中持续存在。这一发现揭示了病毒持续性的新病毒决定因素，对rCl-13(GPC/VGKS)的详细表征可为持续性病毒感染的潜在机制提供新的见解。
