The nonstructural protein 1 (nsp1) of severe acute respiratory syndrome coronavirus and severe acute respiratory syndrome coronavirus 2 is a critical viral protein that suppresses host gene expression by blocking the assembly of the ribosome on host mRNAs. To understand the mechanism of inhibition of host gene expression, we sought to identify cellular proteins that interact with nsp1. Using proximity-dependent biotinylation followed by proteomic analyses of biotinylated proteins, here we captured multiple dynamic interactions of nsp1 with host cell proteins. In addition to ribosomal proteins, we identified several pre-mRNA processing proteins that interact with nsp1, including splicing factors and transcription termination proteins, as well as exosome, and stress granule (SG)-associated proteins. We found that the interactions with transcription termination factors are primarily governed by the C-terminal region of nsp1 and are disrupted by the mutation of amino acids K164 and H165 that are essential for its host shutoff function. We further show that nsp1 interacts with Ras GTPase-activating protein SH3 domain-binding protein 1 (G3BP1) and colocalizes with G3BP1 in SGs under sodium arsenite-induced stress. Finally, we observe that the presence of nsp1 disrupts the maturation of SGs over a long period. Isolation of SG core at different times shows a gradual loss of G3BP1 in the presence of nsp1.

中文翻译：

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邻近依赖性生物素化检测 SARS 冠状病毒非结构蛋白 1 和应激颗粒相关蛋白之间的关联。


严重急性呼吸综合征冠状病毒和严重急性呼吸综合征冠状病毒2的非结构蛋白1 (nsp1)是一种关键的病毒蛋白，它通过阻断宿主mRNA上核糖体的组装来抑制宿主基因表达。为了了解抑制宿主基因表达的机制，我们试图鉴定与 nsp1 相互作用的细胞蛋白。使用邻近依赖性生物素化，然后对生物素化蛋白质进行蛋白质组学分析，我们捕获了 nsp1 与宿主细胞蛋白质的多种动态相互作用。除了核糖体蛋白之外，我们还鉴定了几种与 nsp1 相互作用的前 mRNA 加工蛋白，包括剪接因子和转录终止蛋白，以及外泌体和应激颗粒 (SG) 相关蛋白。我们发现与转录终止因子的相互作用主要由 nsp1 的 C 端区域控制，并被对其宿主关闭功能至关重要的氨基酸 K164 和 H165 的突变所破坏。我们进一步表明，在亚砷酸钠诱导的应激下，nsp1 与 Ras GTP 酶激活蛋白 SH3 结构域结合蛋白 1 (G3BP1) 相互作用，并与 SG 中的 G3BP1 共定位。最后，我们观察到 nsp1 的存在会长期破坏 SG 的成熟。不同时间SG核心的分离显示在nsp1存在的情况下G3BP1逐渐丢失。