Iserman Christiane, Roden Christine, Boerneke Mark, Sealfon Rachel, McLaughlin Grace, Jungreis Irwin, Park Chris, Boppana Avinash, Fritch Ethan, Hou Yixuan J, Theesfeld Chandra, Troyanskaya Olga G, Baric Ralph S, Sheahan Timothy P, Weeks Kevin, Gladfelter Amy S
Department of Biology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
bioRxiv. 2020 Jun 12:2020.06.11.147199. doi: 10.1101/2020.06.11.147199.
A mechanistic understanding of the SARS-CoV-2 viral replication cycle is essential to develop new therapies for the COVID-19 global health crisis. In this study, we show that the SARS-CoV-2 nucleocapsid protein (N-protein) undergoes liquid-liquid phase separation (LLPS) with the viral genome, and propose a model of viral packaging through LLPS. N-protein condenses with specific RNA sequences in the first 1000 nts (5'-End) under physiological conditions and is enhanced at human upper airway temperatures. N-protein condensates exclude non-packaged RNA sequences. We comprehensively map sites bound by N-protein in the 5'-End and find preferences for single-stranded RNA flanked by stable structured elements. Liquid-like N-protein condensates form in mammalian cells in a concentration-dependent manner and can be altered by small molecules. Condensation of N-protein is sequence and structure specific, sensitive to human body temperature, and manipulatable with small molecules thus presenting screenable processes for identifying antiviral compounds effective against SARS-CoV-2.
对严重急性呼吸综合征冠状病毒2（SARS-CoV-2）病毒复制周期的机制理解对于开发针对2019冠状病毒病（COVID-19）全球健康危机的新疗法至关重要。在本研究中，我们表明SARS-CoV-2核衣壳蛋白（N蛋白）与病毒基因组发生液-液相分离（LLPS），并提出了一种通过LLPS进行病毒包装的模型。在生理条件下，N蛋白与前1000个核苷酸（5'端）中的特定RNA序列凝聚，并且在人类上呼吸道温度下增强。N蛋白凝聚物排除未包装的RNA序列。我们全面绘制了N蛋白在5'端结合的位点，并发现对由稳定结构元件侧翼的单链RNA有偏好。类液体的N蛋白凝聚物以浓度依赖的方式在哺乳动物细胞中形成，并且可以被小分子改变。N蛋白的凝聚是序列和结构特异性的，对人体温度敏感，并且可以被小分子操纵，因此为鉴定对SARS-CoV-2有效的抗病毒化合物提供了可筛选的过程。
