Helicase Regulation of dsRNA Signaling and Innate Antiviral Immune Responses

描述（由申请人提供）：该项目的长期目标是了解先天抗病毒反应的调控机制和特定介质。宿主对病毒感染的免疫反应的主要贡献是I型干扰素（IFN）的产生和反应。病毒感染或积累的病毒复制中间体，如双链RNA (dsRNA)，无论是否存在toll样受体系统，都可以在大多数细胞中触发IFN的生物合成。胞质内dsRNA的检测是由一系列传感器蛋白介导的，包括rig - 1、MDA5和LGP2，这三种蛋白含有DEXD/H-box RNA解旋酶结构域。rig - 1和MDA5解旋酶结构域融合到与CARD结构域蛋白同源的n端区域，是导致IFN生物合成的正信号蛋白。LGP2缺乏CARD结构域，被认为是IFN合成的反馈抑制剂。尽管RNA解旋酶结构域相似，但对具体的酶活性以及催化活性如何影响信号传导知之甚少。初步结果和已发表的报告显示，解旋酶蛋白在RNA识别方面具有高度的特异性，并且对酶活性有不同的要求。天然抑制剂副粘病毒V蛋白选择性靶向MDA5和LGP2，但不靶向rig - 1，进一步揭示了特异性。该建议将确定RNA靶标偏好和酶专门化的作用，赋予解旋酶介导的抗病毒反应的特异性和选择性。将揭示选择性副粘病毒解旋酶干扰的分子基础，并与MDA5和LGP2对宿主应答的生物学评价相关联。调节机制和解旋酶介导的信号传导的细胞伙伴将被表征。这些目的将揭示细胞内先天抗病毒反应的正调控和负调控的特异性决定因素。麻疹、腮腺炎、人副流感和呼吸道合胞病毒等副黏液病毒，以及包括流感、艾滋病毒、埃博拉病毒和丙型肝炎病毒在内的其他RNA病毒，对全世界人类健康构成传染性威胁。提出的实验将揭示细胞抗病毒反应的基本机制，并确定细胞和病毒抑制机制的分子基础。这一策略将揭示抗病毒反应的优势和弱点，并确定治疗干预的目标。

DESCRIPTION (provided by applicant): The long term objective for this project is to understand the regulatory mechanisms and specific mediators of innate antiviral responses. A major contribution to host immune responses to virus infection is the production of and response to type I interferon (IFN). Virus infection or accumulated virus replication intermediates like double-stranded RNA (dsRNA) can trigger IFN biosynthesis in most cells irrespective of the presence of Toll-like receptor systems. Detection of cytosolic dsRNA is mediated by a family of sensor proteins that includes RIG-I, MDA5, and LGP2, three proteins that contain DEXD/H-box RNA helicase domains. RIG-I and MDA5 helicase domains are fused to an N-terminal region homologous to CARD domain proteins, and are positive signaling proteins leading to IFN biosynthesis. LGP2 lacks the CARD domain, and has been characterized as a feedback inhibitor for IFN synthesis. Despite the RNA helicase domain similarities, little is known about the specific enzymatic activities and how catalytic activity can influence the signaling. Preliminary results and published reports reveal a high degree of specificity in RNA recognition and differential requirements for enzymatic activity among the helicase proteins. Further specificity is revealed by natural inhibitors, paramyxovirus V proteins, that selectively target MDA5 and LGP2, but not RIG-I. This proposal will determine RNA target preferences and roles for enzymatic specialization that confer specificity and selectivity to helicase-mediated antiviral responses. The molecular basis for selective paramyxovirus helicase interference will be revealed and correlated with biological evaluation of the contributions of MDA5 and LGP2 to host responses. Regulatory mechanisms and cellular partners for helicase-mediated signaling will be characterized. These aims will expose specificity determinants for both positive and negative regulation of the intracellular innate antiviral response. Paramyxoviruses like measles, mumps human parainfluenza, and respiratory syncytial viruses, as well as other RNA viruses including influenza, HIV, Ebola and hepatitis C viruses are infectious threats to human health worldwide. The proposed experiments will reveal basic mechanisms of cellular antiviral responses and determine the molecular basis for both cellular and viral inhibition mechanisms. This strategy will reveal the strengths and vulnerabilities of the antiviral response and identify targets for therapeutic intervention.