Laboratory of Cell Engineering, Research Unit of Cell Death Mechanism, Beijing Institute of Biotechnology, Chinese Academy of Medical Sciences (2021RU008), 20 Dongda Street, Beijing, 100071, China.
The Experimental High School Attached to Beijing Normal University, Beijing, China.
Biol Direct. 2022 Jun 5;17(1):14. doi: 10.1186/s13062-022-00325-x.
Rodents, such as mice, are vulnerable targets, and potential intermediate hosts, of SARS-CoV-2 variants of concern, including Alpha, Beta, Gamma, and Omicron. N501Y in the receptor-binding domain (RBD) of Spike protein is the key mutation dictating the mouse infectivity, on which the neighboring mutations within RBD have profound impacts. However, the impacts of mutations outside RBD on N501Y-mediated mouse infectivity remain to be explored.
Herein, we report that two non-RBD mutations derived from mouse-adapted strain, Ins215KLRS in the N-terminal domain (NTD) and H655Y in the subdomain linking S1 to S2, enhance mouse infectivity in the presence of N501Y mutation, either alone or together. This is associated with increased interaction of Spike with mouse ACE2 and mutations-induced local conformation changes in Spike protein. Mechanistically, the H655Y mutation disrupts interaction with N657, resulting in a less tight loop that wraps the furin-cleavage finger; and the insertion of 215KLRS in NTD increases its intramolecular interaction with a peptide chain that interfaced with the RBD-proximal region of the neighboring protomer, leading to a more flexible RBD that facilitates receptor binding. Moreover, the Omicron Spike that contains Ins214EPE and H655Y mutations confer mouse infectivity > 50 times over the N501Y mutant, which could be effectively suppressed by mutating them back to wild type.
Collectively, our study sheds light on the cooperation between distant Spike mutations in promoting virus infectivity, which may undermine the high infectiousness of Omicron variants towards mice.
啮齿动物（如老鼠）是 SARS-CoV-2 变体的易感目标和潜在中间宿主，包括 Alpha、Beta、Gamma 和 Omicron。 Spike 蛋白受体结合域（RBD）中的 N501Y 突变是决定病毒对老鼠感染性的关键突变，而 RBD 内的相邻突变对其具有深远影响。然而，RBD 之外的突变对 N501Y 介导的老鼠感染性的影响仍有待探索。
在此，我们报告两个源自鼠适应株的非 RBD 突变，即 N 端结构域（NTD）中的 Ins215KLRS 和 S1 与 S2 连接子中的 H655Y，在 N501Y 突变存在的情况下，单独或共同增强了老鼠的感染性。这与 Spike 与老鼠 ACE2 的相互作用增加以及突变诱导的 Spike 蛋白局部构象变化有关。在机制上，H655Y 突变破坏了与 N657 的相互作用，导致包裹弗林切割指的环变松；而 NTD 中的 215KLRS 插入增加了其与界面与相邻亚基 RBD 近端区域的肽链的分子内相互作用，导致更灵活的 RBD，从而促进受体结合。此外，含有 Ins214EPE 和 H655Y 突变的 Omicron Spike 赋予了比 N501Y 突变体高出 50 倍的老鼠感染性，而通过将这些突变体突变为野生型则可以有效地抑制其感染性。
总的来说，我们的研究揭示了 Spike 突变之间的远距离合作在促进病毒感染性方面的作用，这可能削弱了 Omicron 变体对老鼠的高传染性。
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