SARS-CoV-2 Mac1 is required for IFN antagonism and efficient virus replication in cell culture and in mice.
SARS-CoV-2 Mac1 is required for IFN antagonism and efficient virus replication in cell culture and in mice.
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DOI：
10.1073/pnas.2302083120
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发表时间：
2023-08-29
影响因子：
11.1
通讯作者：
Fehr, Anthony R.
中科院分区：
文献类型：
--
作者：
Alhammad, Yousef M.;Parthasarathy, Srivatsan;Ghimire, Roshan;Kerr, Catherine M.;O'Connor, Joseph J.;Pfannenstiel, Jessica J.;Chanda, Debarati;Miller, Caden A.;Baumlin, Nathalie;Salathe, Matthias;Unckless, Robert L.;Zuniga, Sonia;Enjuanes, Luis;More, Sunil;Channappanavar, Rudragouda;Fehr, Anthony R.
所有 CoV，包括 SARS-CoV-2，都编码一个保守的宏结构域 (Mac1)，可对抗宿主 ADP 核糖基化。之前对 SARS-CoV 和 MHV 的研究发现 Mac1 是发病机制所必需的，这促使了 SARS-CoV-2 Mac1 抑制剂的开发。然而，将这些化合物开发成抗病毒药物需要我们了解缺乏 Mac1 的 SARS-CoV-2 在体外和体内复制和引起疾病的程度。在这里，我们发现含有完整 Mac1 缺失的 SARS-CoV-2 在细胞培养物中正常复制，但诱导 IFN 反应升高，降低了体内病毒载量，并且不会在小鼠中引起明显的疾病。这些结果将为测试 Mac1 抑制剂提供路线图，帮助识别 Mac1 功能，并为冠状病毒治疗开辟更多途径。几种冠状病毒 (CoV) 编码的蛋白质正在被评估为 COVID-19 抗病毒治疗的靶标。这些药物靶点包括保守的大结构域（Mac1），一种 ADP-核糖基水解酶和 ADP-核糖结合蛋白，编码为非结构蛋白 3 N 末端的小结构域。利用点突变重组病毒，Mac1 被证明对鼠肝炎病毒 (MHV) 和严重急性呼吸综合征 (SARS)-CoV 毒力至关重要。然而，作为潜在的药物靶点，必须了解 Mac1 的完全缺失如何影响不同 CoV 的复制和发病机制。为此，我们创建了重组细菌人工染色体 (BAC)，其中包含 MHV、MERS-CoV 和 SARS-CoV-2 中 Mac1 的完整缺失 (ΔMac1)。虽然我们无法从 MHV 或 MERS-CoV ΔMac1 BAC 中回收感染性病毒，但 SARS-CoV-2 ΔMac1 很容易从 BAC 转染中回收，这表明不同 CoV 之间对 Mac1 的需求存在明显差异。此外，SARS-CoV-2 ΔMac1 在多个易受感染的细胞系中以野生型水平或接近野生型水平复制。然而，在严重感染的小鼠模型中，ΔMac1 很快被清除，导致最小的病理变化而没有任何发病率。 ΔMac1 SARS-CoV-2 诱导细胞培养物和小鼠中干扰素 (IFN) 和 IFN 刺激的基因表达水平升高，表明 Mac1 阻断 IFN 反应，这可能有助于其减弱。 ΔMac1 感染还导致炎症单核细胞和中性粒细胞明显减少。这些结果表明，与 MHV 和 MERS-CoV 不同，Mac1 对 SARS-CoV-2 复制的影响极小，但它是 SARS-CoV-2 发病机制所必需的，并且是独特的抗病毒药物靶点。
All CoVs, including SARS-CoV-2, encode for a conserved macrodomain (Mac1) that counters host ADP-ribosylation. Prior studies with SARS-CoV and MHV found that Mac1 is required for pathogenesis, which has prompted the development of SARS-CoV-2 Mac1 inhibitors. However, development of these compounds into antivirals requires that we understand the degree to which SARS-CoV-2 lacking Mac1 replicates and causes disease in vitro and in vivo. Here, we found that SARS-CoV-2 containing a complete Mac1 deletion replicates normally in cell culture but induces an elevated IFN response, has reduced viral loads in vivo, and does not cause significant disease in mice. These results will provide a roadmap for testing Mac1 inhibitors, help identify Mac1 functions, and open additional avenues for coronavirus therapies. Several coronavirus (CoV) encoded proteins are being evaluated as targets for antiviral therapies for COVID-19. Included in these drug targets is the conserved macrodomain, or Mac1, an ADP-ribosylhydrolase and ADP-ribose binding protein encoded as a small domain at the N terminus of nonstructural protein 3. Utilizing point mutant recombinant viruses, Mac1 was shown to be critical for both murine hepatitis virus (MHV) and severe acute respiratory syndrome (SARS)-CoV virulence. However, as a potential drug target, it is imperative to understand how a complete Mac1 deletion impacts the replication and pathogenesis of different CoVs. To this end, we created recombinant bacterial artificial chromosomes (BACs) containing complete Mac1 deletions (ΔMac1) in MHV, MERS-CoV, and SARS-CoV-2. While we were unable to recover infectious virus from MHV or MERS-CoV ΔMac1 BACs, SARS-CoV-2 ΔMac1 was readily recovered from BAC transfection, indicating a stark difference in the requirement for Mac1 between different CoVs. Furthermore, SARS-CoV-2 ΔMac1 replicated at or near wild-type levels in multiple cell lines susceptible to infection. However, in a mouse model of severe infection, ΔMac1 was quickly cleared causing minimal pathology without any morbidity. ΔMac1 SARS-CoV-2 induced increased levels of interferon (IFN) and IFN-stimulated gene expression in cell culture and mice, indicating that Mac1 blocks IFN responses which may contribute to its attenuation. ΔMac1 infection also led to a stark reduction in inflammatory monocytes and neutrophils. These results demonstrate that Mac1 only minimally impacts SARS-CoV-2 replication, unlike MHV and MERS-CoV, but is required for SARS-CoV-2 pathogenesis and is a unique antiviral drug target.
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影响因子：
6.4
作者：
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通讯作者：
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DOI：
10.1016/j.bbrc.2007.10.081
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2007-12-28
影响因子：
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