ANTIVIRAL MECHANISMS OF 2-5A-DEPENDENT RNASE L

2-5A依赖性RNA酶L的抗病毒机制

• 批准号: 9761446
• 负责人: Robert H. Silverman
• 金额: $ 40.06万
• 依托单位: CLEVELAND CLINIC LERNER COM-CWRU
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-25 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9761446
• 关键词: 2-5A Synthetase; ABCE1 gene; ADAR1; Antiviral Agents; Antiviral Response; Apoptotic; Autophagocytosis; Binding; Biological Assay; Caspase; Cause of Death; Cell Death; Cells; Cleaved cell; Complex; Crystallization; DNA Viruses; Double-Stranded RNA; Enhancers; Enzymes; Equilibrium; Family; Health; Host Defense; Human; Immune system; Inflammasome; Inflammation; Inflammatory; Integration Host Factors; Interferons; Knockout Mice; Knowledge; Lead; Mediating; Mus; Oncolytic viruses; Pathogenicity; Pathway interactions; Pharmaceutical Preparations; Philadelphia; Population; Protein Kinase; Protocols documentation; RNA Viruses; Ramp; Reporting; Ribonucleases; Roentgen Rays; Role; Signal Transduction; Small RNA; Structure; System; Therapeutic; Toxic effect; Vertebrates; Viral; Virus; Virus Diseases; dimer; improved; in vivo; inhibitor/antagonist; knockout gene; member; novel; oligoadenylate; pathogenic virus; phosphodiesterase V; small molecule; small molecule inhibitor; viral RNA

The 2',5'-oligoadenylate (2-5A) synthetase (OAS)-RNase L system is a robust interferon (IFN)-inducible antiviral pathway that contributes to the survival of higher vertebrates against both RNA and DNA viruses. Accordingly, viruses have acquired and evolved a multitude of mechanisms to antagonize OAS-RNase L. During the past 5 years, we established roles for RNase L in IFN signaling, inflammasome activation and autophagy. Yet there remains much to be learned about how RNase L suppresses viral infections, how viruses evade these enzymes, and how the host recovers from RNase L activation. In particular, our knowledge of how RNase L is regulated through its protein kinase-like domain, how RNase L mediates cell death from ADAR1 deficiency, how the RNase L inhibitor (ABCE1 aka RLI) functions, and how RNase L activates the NLRP3 inflammasome is still fragmentary. Therefore, the proposed continuation of this project will systematically investigate the major outstanding questions about this critically important host antiviral pathway. The unifying theme of this application concerns how RNase L function is regulated beyond its activation by 2-5A, in particular through its protein kinase-like (PKL) domain, to control viral infections and inflammation. Our findings lead to the hypothesis that the OAS-RNase L system is an integral part of the host immune system that must be tightly regulated to eliminate viruses while also limiting its harmful pro-inflammatory effects. Our specific aims are: (1) To study the impact of the RNase L PKL domain on cells and viruses we will evaluate recently identified small molecule inhibitors, enhancers and their derivatives on RNase L activity, determine the ability of the small molecules to control viral infections, and investigate effects of the RNase L inhibitors on cell death caused by ADAR1 deficiency. (2) To investigate how the ATP-binding cassette member, ABCE1 (RLI), limits RNase L activity, we will determine precisely how ABCE1 interacts with RNase L to inhibit its activity, establish how the host cells ramps up RNase L activity during viral infections by caspase-mediated cleavage of ABCE1, and determine the ability of ABCE1 to counteract cellular toxicity of double stranded RNA by inhibiting RNase L. (3) To probe the mechanism of NLRP3 inflammasome activation by RNase L, we will identify and characterize RNase L-cleaved viral RNA activators of the NLRP3 inflammasome, determine effects of the small RNA cleavage products on inflammasome activation, and determine the impact of RNase L inhibitor drugs on viral mediated inflammation in vivo. Viral infections remain a serious threat to human health worldwide. Despite viral countermeasures, the OAS-RNase L system has a major impact against many types of pathogenic viruses. These studies emphasize how central and important it is to regulate RNase L in order to mediate the balance between antiviral activity and cell death. It is believed that a better understanding of the viral and host factors that regulate RNase L activity may eventually lead to novel and improved therapeutic approaches for treating established and emerging viral infections in the human population.

2 '，5'-寡腺苷酸（2-5A）合成酶（OAS）-RNase L系统是一种稳健的干扰素（IFN）诱导型 抗病毒途径，有助于高等脊椎动物抵抗RNA和DNA病毒的生存。 因此，病毒已经获得并进化了多种拮抗OAS-RNase L的机制。 在过去的5年中，我们确定了RNase L在IFN信号传导、炎性小体激活和细胞凋亡中的作用。 自噬然而，关于核糖核酸酶L如何抑制病毒感染，病毒如何抑制病毒感染， 逃避这些酶，以及宿主如何从RNase L激活中恢复。尤其是，我们对 RNase L通过其蛋白激酶样结构域调节，RNase L如何介导ADAR 1引起的细胞死亡 缺乏，RNase L抑制剂（ABCE 1 aka RLI）如何发挥作用，以及RNase L如何激活NLRP 3 炎性小体仍然是碎片。因此，拟议继续开展的这一项目将系统地 调查关于这一至关重要的宿主抗病毒途径的主要悬而未决的问题。的统一 本申请的主题涉及RNA酶L功能如何在其被2-5A激活之外被调节， 特别是通过其蛋白激酶样（PKL）结构域来控制病毒感染和炎症。我们的研究结果 导致OAS-RNase L系统是宿主免疫系统的组成部分的假设， 严格调控，以消除病毒，同时也限制其有害的促炎作用。我们的具体 目的：（1）研究RNase L PKL结构域对细胞和病毒的影响 鉴定了小分子抑制剂、增强剂及其衍生物对RNase L活性的影响，测定了它们对RNase L活性的抑制能力， 小分子控制病毒感染，并研究RNase L抑制剂对细胞死亡的影响 ADAR 1缺乏症。(2)研究ATP结合盒成员ABCE 1（RLI）如何限制 RNase L活性，我们将精确地确定ABCE 1如何与RNase L相互作用以抑制其活性， 宿主细胞如何在病毒感染期间通过半胱天冬酶介导的ABCE 1切割来提高RNase L活性， 并测定ABCE 1通过抑制RNase L来抵消双链RNA的细胞毒性的能力。 (3)为了探索RNase L激活NLRP 3炎性小体的机制，我们将鉴定和表征 NLRP 3炎性小体的RNase L切割病毒RNA激活剂，决定小RNA 裂解产物对炎性小体激活的影响，并确定RNase L抑制剂药物对病毒感染的影响。 介导的体内炎症。病毒感染仍然是对全世界人类健康的严重威胁。尽管病毒 作为对抗措施，OAS-RNase L系统对许多类型的病原性病毒具有重大影响。 这些研究强调了调节RNase L以调节平衡是多么的核心和重要 抗病毒活性和细胞死亡之间的联系人们相信，更好地了解病毒和宿主因素， 调节RNase L活性可能最终导致新的和改进的治疗方法，用于治疗 在人群中建立和出现病毒感染。