The innate immune antagonism of SARS-CoV-2 ORF3a, ORF6 and ORF7a

人类的先天免疫系统是抵御入侵病原体的强大屏障。例如，I型干扰素系统的诱导导致抗病毒干扰素刺激基因(ISGs)的表达，自噬针对病毒成分进行破坏。要确定感染，病原体，如新型大流行严重急性呼吸综合征冠状病毒-2(SARS-CoV-2)需要操纵人类的先天免疫防御系统。事实上，在冠状病毒病(COVID19)期间观察到的许多临床症状与免疫激活紊乱有关。然而，SARS-CoV-2的致病性低于以往大流行的人类冠状病毒，导致病毒的有效传播。此外，对抗人类免疫防御机制的能力很可能是SARS-CoV-2通过人畜传播给人类的先决条件。我们的数据表明，SARS-CoV-2的Nsp1、ORF3a、ORF6和ORF7a蛋白是干扰素系统或自噬的有效拮抗剂。然而，大多数潜在的分子机制仍有待确定。我们最近可以证明，Nsp1通过阻断核糖体mRNA进入隧道从而关闭I型干扰素反应来抑制细胞翻译。为了阐明ORF3a、ORF6和ORF7a的分子机制(S)，我们对SARS-CoV-2感染细胞和只表达单个蛋白的细胞进行了比较蛋白质组分析。我们的初步数据确定GNG5、LYRM4、Rab5a或RILPL1是分子机制的极有可能的靶点。因此，我们的目标是探索这些因子是如何被ORF3a、ORF6或ORF7a靶向和利用来操纵天然免疫的。接下来，为了确定干扰素和自噬拮抗是否可能在SARS-CoV-2的快速传播中发挥了作用，我们将比较所有七种人类冠状病毒的ORF3a、ORF6或ORF7a的活性。最后，我们打算通过评估相关非人类冠状病毒的辅助蛋白对人类先天免疫系统的活性，来评估它们是否促进了SARS-CoV-2的人畜传播。该项目将为今后深入探讨SARS-CoV-2与人类先天免疫系统的相互作用奠定基础。这可能有助于解释COVID19的症状，从而为新的治疗靶点提供线索。保护分析将为SARS-CoV-2的快速传播和人畜传播提供洞察力。最后，识别SARS-CoV-2编码的免疫拮抗剂可能有助于故意构建一种有效激活我们免疫系统的减毒株。这些毒株可能诱导长期免疫，有助于合理的疫苗设计。

The human innate immune system is a powerful barrier against invading pathogens. For example, induction of the type-I interferon system leads to the expression of anti-viral interferon-stimulated genes (ISGs) and autophagy targets viral components for destruction. To establish an infection, pathogens such as the novel pandemic severe-acute respiratory syndrome Coronavirus-2 (SARS-CoV-2) need to manipulate the human innate immune defense systems. Indeed, many clinical symptoms observed during the Coronavirus disease (COVID19) are related to perturbed immune activation. However, the pathogenicity of SARS-CoV-2 is lower than previous pandemic human CoVs, resulting in an efficient spread of the virus. In addition, the ability to antagonize human immune defense mechanisms was most likely a prerequisite for zoonotic transmission of SARS-CoV-2 to humans. Our data show that the proteins Nsp1, ORF3a, ORF6 and ORF7a of SARS-CoV-2 are potent antagonists of the interferon system or autophagy. However, most of the underlying molecular mechanisms remain to be determined. We could recently show that Nsp1 inhibits cellular translation by blocking the ribosome mRNA entry tunnel thus shutting down type-I interferon responses. To elucidate the molecular mechanism(s) of ORF3a, ORF6 and ORF7a, we conducted comparative proteome analysis of SARS-CoV-2 infected cells and cells expressing only the individual proteins. Our preliminary data identified GNG5, LYRM4, Rab5a or RILPL1 as highly likely targets of the molecular mechanism. Thus, we aim to explore how these factors are targeted and exploited by ORF3a, ORF6 or ORF7a to manipulate innate immunity. Next, to determine whether interferon and autophagy antagonism may have played a role in the rapid spread of SARS-CoV-2, we will compare the activities of ORF3a, ORF6 or ORF7a of all seven human CoVs. Finally, we intend to assess whether they facilitated zoonotic transmission of SARS-CoV-2 by assessing the activity of the accessory proteins of related non-human CoVs against the human innate immune system. This project will be the basis of future in-depth exploration of of the interplay between SARS-CoV-2 and the human innate immune system. This may help explain symptoms of COVID19 and thus give hints for new therapeutic targets. Conservation analysis will yield insights on the rapid spread and zoonotic transmission of SARS-CoV-2. Finally, identification of immune antagonists encoded by SARS-CoV-2 may help deliberately constructing an attenuated strain, that effectively activate our immune system. These strains may induce long-term immunity, aiding rational vaccine design.