Molecular mechanisms of immune dysregulation by filoviral interferon

丝状病毒埃博拉病毒和马尔堡病毒(EBOV和MARV)是一种新出现的负链RNA病毒，可引起严重的病毒性出血热。这些人畜共患病病原体的毒力和新出现的性质使它们对人类健康构成重大威胁，是生物恐怖主义的潜在诱因，也是NIAID A类优先病原体。目前，还没有获得批准的抗丝状病毒疗法。丝状病毒出血热的特征是病毒复制失控，全身炎症和凝血过度，树突状细胞(DC)功能失调和淋巴细胞凋亡。我们先前鉴定并鉴定了丝状病毒干扰素拮抗剂蛋白，包括EBOV和MARV VP35，EBOV VP24和MARV VP40。VP35中和干扰素(干扰素)-α/β的产生，这是关键的抗病毒细胞因子。EBOV VP24和MARV VP40可阻断干扰素-α/β诱导的信号传导。我们推测，这些病毒干扰素拮抗剂的功能不仅将抵消干扰素-α/β的固有抗病毒作用，而且通过靶向模式识别、干扰素和细胞因子受体下游的信号通路，还将能够扰乱正常的DC成熟过程，促进失调的DC-T淋巴细胞相互作用。通过这种方式，丝状病毒干扰素拮抗剂将扰乱先天和获得性抗病毒免疫。在西奈山伊坎医学院巴斯勒实验室与华盛顿大学阿马拉辛格和格罗斯实验室之间持续合作的基础上，我们将表征丝状病毒干扰素拮抗剂扰乱免疫的分子机制。利用体外生化和结构研究，华盛顿大学的阿马拉辛哈和格罗斯实验室将确定丝状病毒干扰素拮抗剂如何与宿主免疫系统信号分子相互作用的结构和生化基础，并设计功能缺失突变体。巴斯勒实验室将确定每种丝状病毒干扰素拮抗剂在树突状细胞和巨噬细胞中影响的特定信号通路，并使用Amarasinghe和Gross实验室提供的结构和生化数据以及关键信号分子的特定敲除将确定丝状病毒干扰素拮抗剂对巨噬细胞功能和DC细胞成熟的影响。由此产生的数据将产生关于丝状病毒干扰素拮抗剂对EBOV和MARV抑制DC功能(由项目2测试)、促进病毒复制以及在非人类灵长类动物模型中诱导干扰素和获得性免疫反应(由项目3解决)的贡献的具体假设。

The filoviruses, Ebola and Marburg viruses (EBOV and MARV), are emerging, negative-strand RNA viruses that cause severe viral hemorrhagic fever. The virulence and emerging nature of these zoonotic pathogens makes them a significant threat to human health, potential agents of bioterrorism and NIAID category A priority pathogens. Currently, no approved anti-filovirus therapeutics are available. Filoviral hemorrhagic fever is characterized by uncontrolled, systemic virus replication, excessive inflammation and coagulation, dysregulated dendritic cell (DC) function and lymphocyte apoptosis. We previously identified and structurally characterized filoviral interferon antagonist proteins, including EBOV and MARV VP35, EBOV VP24 and MARV VP40. The VP35s counteract the production of interferon (IFN)-alpha/beta, critical antiviral cytokines. EBOV VP24 and MARV VP40 block the signaling induced by IFN-alpha/beta. We hypothesize that the function of these viral IFN-antagonists will not only counteract the innate antiviral effects of IFN-alpha/beta but, by targeting signaling downstream of pattern recognition, interferon and cytokine receptors, will also be able to disrupt the normal DC maturation process and promote dysregulated DC-T lymphocyte interactions. In this way, the filoviral IFN-antagonists will disrupt both innate and adaptive antiviral immunity. Building on ongoing collaborations between the Basler laboratory at the Icahn School of Medicine at Mt Sinai and the Amarasinghe and Gross laboratories at Washington University, we will characterize the molecular mechanisms of immune disruption by filoviral interferon antagonists. Using in vitro biochemical and structural studies, the Amarasinghe and Gross laboratories at Washington University will define the structural and biochemical basis for how filoviral interferon antagonists interact with host immune system signaling molecules and design loss of function mutants. The Basler laboratory will define the specific signaling pathways affected by each filoviral IFN-antagonist in dendritic cells and macrophages and using the structural and biochemical data provided by Amarasinghe and Gross labs as well as specific knockdown of key signaling molecules will define the impact of filoviral IFN-antagonists on macrophage function and DC cell maturation. The resulting data will generate specific hypotheses regarding the contribution of filoviral IFN-antagonists to EBOV and MARV suppression of DC function (to be tested by Project 2), to promotion of virus replication and induction of IFN and adaptive immune responses in non-human primate models (addressed by Project 3).