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

人类先天免疫系统是抵抗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（ISG）的表达，并且自噬靶向病毒组分以进行破坏。为了建立感染，病原体如新型大流行性严重急性呼吸道综合征冠状病毒2（SARS-CoV-2）需要操纵人类先天免疫防御系统。事实上，在冠状病毒疾病（COVID 19）期间观察到的许多临床症状与干扰的免疫激活有关。然而，SARS-CoV-2的致病性低于以前的大流行人类CoV，导致病毒的有效传播。此外，拮抗人类免疫防御机制的能力很可能是SARS-CoV-2向人类传播的先决条件。我们的数据表明，蛋白Nsp 1，ORF 3a，ORF 6和ORF 7a的SARS-CoV-2是有效的拮抗剂的干扰素系统或自噬。然而，大多数潜在的分子机制仍有待确定。我们最近发现Nsp 1通过阻断核糖体mRNA进入通道从而关闭I型干扰素反应来抑制细胞翻译。为了阐明ORF 3a、ORF 6和ORF 7a的分子机制，我们对SARS-CoV-2感染细胞和仅表达单个蛋白的细胞进行了比较蛋白质组学分析。我们的初步数据确定GNG 5，LYRM 4，Rab 5a或RILPL 1是分子机制的高度可能的靶点。因此，我们的目标是探索这些因子是如何被ORF 3a，ORF 6或ORF 7a靶向和利用来操纵先天免疫的。接下来，为了确定干扰素和自噬拮抗作用是否可能在SARS-CoV-2的快速传播中发挥作用，我们将比较所有七种人类CoV的ORF 3a，ORF 6或ORF 7a的活性。最后，我们打算通过评估相关非人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的人畜共患传播。该项目将为未来深入探索SARS-CoV-2与人类先天免疫系统之间的相互作用奠定基础。这可能有助于解释COVID 19的症状，从而为新的治疗靶点提供线索。保护性分析将有助于了解SARS-CoV-2的快速传播和人畜共患传播。最后，SARS-CoV-2编码的免疫拮抗剂的鉴定可能有助于故意构建减毒株，有效激活我们的免疫系统。这些菌株可诱导长期免疫，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.