Viral inhibition of cell death in host immune responses

工作摘要/摘要 细胞死亡是一种强大的免疫防御机制，可以限制病毒在受感染宿主内的复制。在……里面 病毒通常编码抑制物来对抗宿主细胞的死亡。例如，许多疱疹病毒 痘病毒编码半胱氨酸天冬氨酸酶的抑制物，半胱氨酸酶是执行细胞凋亡的关键酶。通过颠覆主机 细胞死亡和相关的炎症反应，病毒细胞死亡抑制剂也可以戏剧性地改变 病毒性疾病的后果。 坏死性下垂是一种细胞死亡的裂解形式，在caspase8抑制的情况下诱导效果最佳。 因此，坏死性下垂在控制编码半胱氨酸天冬氨酸酶抑制剂的病毒方面起着关键作用。此外， 从坏死性细胞中释放“损伤相关分子模式(DAMP)”可以进一步刺激抗- 病毒性炎症。这些抗病毒作用表明，坏死性下垂也可能是病毒抑制的目标。至 为了验证这一假设，我们进行了siRNA筛选，发现了我们已经拥有的牛痘病毒的病毒抑制物 被称为“RIPK3降解的病毒诱导剂(VIRD)”。在初步研究中，我们发现vIRD通过以下方式发挥作用 针对蛋白酶体降解的必需的坏死下垂激酶RIPK3。 在这些观察的基础上，我们提出了三个目标来进一步阐明其功能和机制 VIRD的。在第一个目标中，我们将评估VIRD感染牛痘病毒和其他相关疾病的机制。 正痘病毒促进RIPK3的降解。研究将包括绘制结构域和残基的地图 VIRD和RIPK3负责它们的物理相互作用和泛素化。在第二个目标中，我们 将询问宿主Skp1-Cullin-F盒(SCF)复合体在vIRD介导的RIPK3降解中的作用。 我们还将评估vIRD对受感染细胞的全球泛素化格局的影响。在目标3中，我们 将以小鼠感染为模型评价vIRD的生物学功能。我们将用牛痘感染老鼠 表达vIRD不同的病毒或痘苗病毒。VIRD对细胞死亡、炎症、 将检查病毒复制和组织病理学。总而言之，这些研究将揭示医学上的相关性 有关病毒如何操纵免疫系统和改变病毒疾病结局的信息。我们预计 从拟议研究中获得的知识将为更好和更多的设计提供重要的见解 有效的疫苗。

Abstract/Summary of Work Cell death is a powerful immune defense mechanism to limit viral replication within the infected host. In response, viruses often encode inhibitors that counteract host cell death. For example, many herpesviruses and poxviruses encode inhibitors of caspases, the key enzymes that execute apoptosis. By subverting host cell death and the associated inflammatory response, viral cell death inhibitors can also dramatically alter the outcome of viral diseases. Necroptosis is a lytic form of cell death that is optimally induced in the presence of caspase 8 inhibition. As such, necroptosis has critical roles in controlling viruses that encode caspase inhibitors. Moreover, the release of “damage-associated molecular patterns (DAMPs)” from necroptotic cells can further stimulate anti- viral inflammation. These anti-viral effects suggest that necroptosis may also be a target of viral inhibition. To test this hypothesis, we conducted a siRNA screen and found a viral inhibitor from cowpox virus that we have termed “viral inducer of RIPK3 degradation (vIRD)”. In preliminary studies, we found that vIRD functions by targeting the essential necroptosis kinase RIPK3 for proteasomal degradation. Based on these observations, we propose three aims to further elucidate the function and mechanism of vIRD. In the first aim, we will evaluate the mechanism by which vIRD from cowpox virus and other related orthopoxviruses promotes RIPK3 degradation. Studies will include mapping the domain and residues within vIRD and RIPK3 that are responsible for their physical interaction and ubiquitination. In the second aim, we will interrogate the role of the host SKP1-Cullin-F box (SCF) complex in vIRD-mediated RIPK3 degradation. We will also evaluate the effect of vIRD on the global ubiquitination landscape of the infected cell. In Aim 3, we will evaluate the biological function of vIRD using mouse infection as model. We will infect mice with cowpox virus or vaccinia virus that differ in their expression of vIRD. The effect of vIRD on cell death, inflammation, viral replication and tissue pathology will be examined. Collectively, these studies will reveal medically relevant information on how viruses manipulate the immune system and alter the outcome of viral disease. We expect the knowledge gained from the proposed study will provide important insight in the design of better and more efficacious vaccines.