Department of Pathology, Galveston National Laboratory and Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, Texas, USA.
Department of Pathology, Galveston National Laboratory and Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, Texas, USA
J Virol. 2020 Apr 16;94(9). doi: 10.1128/JVI.02006-19.
The arenaviruses Lassa virus (LASV), Junín virus (JUNV), and Machupo virus (MACV) can cause severe and fatal diseases in humans. Although these pathogens are closely related, the host immune responses to these virus infections differ remarkably, with direct implications for viral pathogenesis. LASV infection is immunosuppressive, with a very low-level interferon response. In contrast, JUNV and MACV infections stimulate a robust interferon (IFN) response in a retinoic acid-inducible gene I (RIG-I)-dependent manner and readily activate protein kinase R (PKR), a known host double-stranded RNA (dsRNA) sensor. In response to infection with RNA viruses, host nonself RNA sensors recognize virus-derived dsRNA as danger signals and initiate innate immune responses. Arenavirus nucleoproteins (NPs) contain a highly conserved exoribonuclease (ExoN) motif, through which LASV NP has been shown to degrade virus-derived immunostimulatory dsRNA in biochemical assays. In this study, we for the first time present evidence that LASV restricts dsRNA accumulation during infection. Although JUNV and MACV NPs also have the ExoN motif, dsRNA readily accumulated in infected cells and often colocalized with dsRNA sensors. Moreover, LASV coinfection diminished the accumulation of dsRNA and the IFN response in JUNV-infected cells. The disruption of LASV NP ExoN with a mutation led to dsRNA accumulation and impaired LASV replication in minigenome systems. Importantly, both LASV NP and RNA polymerase L protein were required to diminish the accumulation of dsRNA and the IFN response in JUNV infection. For the first time, we discovered a collaboration between LASV NP ExoN and L protein in limiting dsRNA accumulation. Our new findings provide mechanistic insights into the differential host innate immune responses to highly pathogenic arenavirus infections. Arenavirus NPs contain a highly conserved DEDDh ExoN motif, through which LASV NP degrades virus-derived, immunostimulatory dsRNA in biochemical assays to eliminate the danger signal and inhibit the innate immune response. Nevertheless, the function of NP ExoN in arenavirus infection remains to be defined. In this study, we discovered that LASV potently restricts dsRNA accumulation during infection and minigenome replication. In contrast, although the NPs of JUNV and MACV also harbor the ExoN motif, dsRNA readily formed during JUNV and MACV infections, accompanied by IFN and PKR responses. Interestingly, LASV NP alone was not sufficient to limit dsRNA accumulation. Instead, both LASV NP and L protein were required to restrict immunostimulatory dsRNA accumulation. Our findings provide novel and important insights into the mechanism for the distinct innate immune response to these highly pathogenic arenaviruses and open new directions for future studies.
沙粒病毒科的拉沙病毒（LASV）、胡宁病毒（JUNV）和马丘波病毒（MACV）均可引起人类严重且致命的疾病。尽管这些病原体密切相关，但宿主对这些病毒感染的免疫反应却大不相同，这对病毒发病机制具有直接影响。LASV 感染具有免疫抑制作用，干扰素反应水平很低。相比之下，JUNV 和 MACV 感染以依赖视黄酸诱导基因 I（RIG-I）的方式刺激强烈的干扰素（IFN）反应，并容易激活蛋白激酶 R（PKR），PKR 是一种已知的宿主双链 RNA（dsRNA）传感器。在受到 RNA 病毒感染时，宿主非自身 RNA 传感器将病毒衍生的 dsRNA 识别为危险信号，并启动先天免疫反应。沙粒病毒核蛋白（NP）含有高度保守的核糖核酸外切酶（ExoN）基序，LASV NP 通过该基序在生化分析中已显示出降解病毒衍生的免疫刺激性 dsRNA。在这项研究中，我们首次提供证据表明 LASV 在感染过程中限制 dsRNA 的积累。尽管 JUNV 和 MACV 的 NP 也具有 ExoN 基序，但 dsRNA 在受感染的细胞中很容易积累，并且经常与 dsRNA 传感器共定位。此外，LASV 共感染会减少 JUNV 感染细胞中 dsRNA 的积累和 IFN 反应。用突变破坏 LASV NP 的 ExoN 会导致 dsRNA 积累，并在 minigenome 系统中损害 LASV 复制。重要的是，LASV NP 和 RNA 聚合酶 L 蛋白都需要减少 JUNV 感染中 dsRNA 的积累和 IFN 反应。我们首次发现 LASV NP ExoN 与 L 蛋白之间的协同作用限制了 dsRNA 的积累。我们的新发现为理解高致病性沙粒病毒感染引起的宿主先天免疫反应的差异提供了机制上的见解。沙粒病毒 NP 含有高度保守的 DEDDh ExoN 基序，通过该基序，LASV NP 在生化分析中降解病毒衍生的免疫刺激性 dsRNA，消除危险信号并抑制先天免疫反应。然而，NP ExoN 在沙粒病毒感染中的功能仍有待确定。在这项研究中，我们发现 LASV 在感染和 minigenome 复制过程中有效地限制 dsRNA 的积累。相比之下，尽管 JUNV 和 MACV 的 NP 也具有 ExoN 基序，但在 JUNV 和 MACV 感染过程中很容易形成 dsRNA，并伴有 IFN 和 PKR 反应。有趣的是，LASV NP 本身不足以限制 dsRNA 的积累。相反，LASV NP 和 L 蛋白都需要限制免疫刺激性 dsRNA 的积累。我们的发现为这些高致病性沙粒病毒引起的不同先天免疫反应的机制提供了新的和重要的见解，并为未来的研究开辟了新的方向。
