Nelson Emily V, Schmidt Kristina M, Deflubé Laure R, Doğanay Sultan, Banadyga Logan, Olejnik Judith, Hume Adam J, Ryabchikova Elena, Ebihara Hideki, Kedersha Nancy, Ha Taekjip, Mühlberger Elke
Department of Microbiology, Boston University School of Medicine, Boston University, Boston, Massachusetts, USA.
National Emerging Infectious Diseases Laboratories, Boston University, Boston, Massachusetts, USA.
J Virol. 2016 Jul 27;90(16):7268-7284. doi: 10.1128/JVI.00459-16. Print 2016 Aug 15.
A hallmark of Ebola virus (EBOV) infection is the formation of viral inclusions in the cytoplasm of infected cells. These viral inclusions contain the EBOV nucleocapsids and are sites of viral replication and nucleocapsid maturation. Although there is growing evidence that viral inclusions create a protected environment that fosters EBOV replication, little is known about their role in the host response to infection. The cellular stress response is an effective antiviral strategy that leads to stress granule (SG) formation and translational arrest mediated by the phosphorylation of a translation initiation factor, the α subunit of eukaryotic initiation factor 2 (eIF2α). Here, we show that selected SG proteins are sequestered within EBOV inclusions, where they form distinct granules that colocalize with viral RNA. These inclusion-bound (IB) granules are functionally and structurally different from canonical SGs. Formation of IB granules does not indicate translational arrest in the infected cells. We further show that EBOV does not induce formation of canonical SGs or eIF2α phosphorylation at any time postinfection but is unable to fully inhibit SG formation induced by different exogenous stressors, including sodium arsenite, heat, and hippuristanol. Despite the sequestration of SG marker proteins into IB granules, canonical SGs are unable to form within inclusions, which we propose might be mediated by a novel function of VP35, which disrupts SG formation. This function is independent of VP35's RNA binding activity. Further studies aim to reveal the mechanism for SG protein sequestration and precise function within inclusions.
Although progress has been made developing antiviral therapeutics and vaccines against the highly pathogenic Ebola virus (EBOV), the cellular mechanisms involved in EBOV infection are still largely unknown. To better understand these intracellular events, we investigated the cellular stress response, an antiviral pathway manipulated by many viruses. We show that EBOV does not induce formation of stress granules (SGs) in infected cells and is therefore unrestricted by their concomitant translational arrest. We identified SG proteins sequestered within viral inclusions, which did not impair protein translation. We further show that EBOV is unable to block SG formation triggered by exogenous stress early in infection. These findings provide insight into potential targets of therapeutic intervention. Additionally, we identified a novel function of the interferon antagonist VP35, which is able to disrupt SG formation.
埃博拉病毒（EBOV）感染的一个标志是在受感染细胞的细胞质中形成病毒包涵体。这些病毒包涵体包含EBOV核衣壳，是病毒复制和核衣壳成熟的场所。尽管越来越多的证据表明病毒包涵体创造了一个促进EBOV复制的保护环境，但对于它们在宿主对感染的反应中的作用却知之甚少。细胞应激反应是一种有效的抗病毒策略，可导致应激颗粒（SG）形成以及由翻译起始因子真核起始因子2（eIF2α）的α亚基磷酸化介导的翻译停滞。在这里，我们表明选定的SG蛋白被隔离在EBOV包涵体内，在那里它们形成与病毒RNA共定位的独特颗粒。这些包涵体结合（IB）颗粒在功能和结构上与典型的SG不同。IB颗粒的形成并不表明受感染细胞中的翻译停滞。我们进一步表明，EBOV在感染后的任何时间都不会诱导典型SG的形成或eIF2α磷酸化，但无法完全抑制由不同外源性应激源（包括亚砷酸钠、热和马桑毒素）诱导的SG形成。尽管SG标记蛋白被隔离到IB颗粒中，但典型的SG无法在包涵体内形成，我们认为这可能是由VP35的一种新功能介导的，该功能会破坏SG的形成。此功能独立于VP35的RNA结合活性。进一步的研究旨在揭示SG蛋白被隔离的机制及其在包涵体内的确切功能。
尽管在开发针对高致病性埃博拉病毒（EBOV）的抗病毒疗法和疫苗方面取得了进展，但EBOV感染所涉及的细胞机制仍 largely未知。为了更好地理解这些细胞内事件，我们研究了细胞应激反应，这是许多病毒操纵的一种抗病毒途径。我们表明EBOV不会在受感染细胞中诱导应激颗粒（SG）的形成，因此不受其伴随的翻译停滞的限制。我们鉴定出隔离在病毒包涵体内的SG蛋白，这并不损害蛋白质翻译。我们进一步表明，EBOV在感染早期无法阻断由外源性应激触发的SG形成。这些发现为治疗干预的潜在靶点提供了见解。此外，我们鉴定出干扰素拮抗剂VP35的一种新功能，它能够破坏SG的形成。
