RT-PCR and qPCR Primer sequences.

Viruses face selective pressure to evade cellular antiviral responses to control the outcome of an infection. However, due to their limited genome size, viruses must adopt unique strategies to confront cellular sensors. Since its emergence in humans, SARS-CoV-2 accrued many mutations; however, the functional consequence of many such genetic changes remains unexplored. Here, we show that SARS-CoV-2 produces a truncated form of the nucleocapsid protein, called N*M210. Due to the acquisition of a viral transcription regulatory sequence (TRS) in the N gene, certain variants like Omicron produce a new viral mRNA that markedly increases N*M210 production. We show that N*M210 is a double-stranded RNA (dsRNA)-binding protein. Using its dsRNA binding motif, N*M210 inhibits multiple antiviral responses, supressing interferon, triggering processing body disassembly, and potently blocking G3BP1 foci, including stress granules and RNase L-dependent bodies. Using a panel of recombinant SARS-CoV-2 viruses (rSARS-2), we show that enhanced N*M210 production increases virus fitness in primary human cells and in mice. Furthermore, we show that during infection N*M210 improves virus fitness, in part, due to its ability to potently block G3BP1 foci. We propose a model where, to evade the cellular antiviral response, SARS-CoV-2 has evolved a mechanism to increase the production of a truncated form of the N protein, which limits activation of dsRNA-induced antiviral responses, tipping the balance in favor of the virus in the battle for control of the cell.病毒为掌控感染结局，需逃避宿主细胞的抗病毒应答，因此面临选择压力。然而，由于病毒基因组规模有限，它们必须演化出独特策略以应对宿主细胞的抗病毒感受器。自严重急性呼吸综合征冠状病毒2型（SARS-CoV-2）在人群中出现以来，其积累了众多突变；然而，多数此类遗传变异的功能效应仍未被阐明。本研究发现，SARS-CoV-2可产生一种截短型核衣壳蛋白，命名为N*M210。由于在N基因中获得了病毒转录调控序列（transcription regulatory sequence, TRS），奥密克戎等部分变异株可产生一种新型病毒mRNA，显著提升N*M210的表达水平。研究表明，N*M210是一种双链RNA（double-stranded RNA, dsRNA）结合蛋白。借助其dsRNA结合基序，N*M210可抑制多种抗病毒应答：包括抑制干扰素产生、诱导加工小体（processing body）解体，以及强效阻断G3BP1聚集灶，其中涵盖应激颗粒与核糖核酸酶L（RNase L）依赖性小体。本研究利用一组重组SARS-CoV-2病毒（recombinant SARS-CoV-2, rSARS-2）证实，N*M210表达水平提升可增强病毒在原代人细胞与小鼠体内的适合度。此外，研究证实，在感染过程中N*M210可提升病毒适合度，这部分归因于其强效阻断G3BP1聚集灶的能力。本研究提出一种模型：为逃避宿主细胞的抗病毒应答，SARS-CoV-2演化出一种可提升截短型N蛋白表达水平的机制，该机制可抑制dsRNA诱导的抗病毒应答激活，从而在宿主细胞控制权的博弈中扭转局势，助力病毒占据优势。