Assessing the interplay between the mono-ADP-ribosyltransferase ARTD10 and the Chikungunya viral macrodomain in modulating the antiviral immune response.

评估单 ADP-核糖基转移酶 ARTD10 和基孔肯雅病毒大结构域在调节抗病毒免疫反应中的相互作用。

• 批准号: 401204753
• 负责人: Dr. Patricia Korn
• 金额: --
• 依托单位: Institut für Biochemie und Molekularbiologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2018
• 资助国家: 德国
• 起止时间: 2017-12-31 至 2021-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/401204753?language=en
• 关键词: Assessing; interplay; between; mono; ADP

Viruses entering host cells are recognized by the innate immune system. For efficient replication viruses evolved strategies to antagonize the immune system and to hijack the cellular processes. Recent findings suggest that mono-ADP-ribosylation (MARylation), a post-translational modification, is part of the innate immune response. In this proposal we plan to define the role of MARylation in the conflict between mammalian hosts and Chikungunya virus (CHIKV).CHIKV caused large epidemic outbreaks worldwide in recent years with severe health and economic burden. Presently neither a vaccine nor therapeutic treatments are available. CHIKV encodes only 4 non-structural proteins (nsPs), of which nsP3 harbors a macrodomain, a conserved protein fold, closely linked to MARylation. MARylation involves the transfer of ADP-ribose from NAD+ onto substrate proteins and is intracellularly mainly catalyzed by ADP-ribosyltransferases diphtheria toxin-like (ARTDs). We focus on ARTD10, which is induced in response to type I interferons (IFNs). We identified the CHIKV macrodomain as a de-MARylating enzyme, which is required for efficient viral replication. Moreover, preliminary work demonstrates that nsP1-nsP3 are substrates of ARTD10 and thus their activities potentially controlled by MARylation. These findings led us to hypothesize a role for MARylation in innate immunity, which we propose to function in two distinct ways: (I) Host-dependent MARylation of the CHIKV nsPs directly affects their function and thereby impacts on the viral life cycle. (II) MARylation of host-factors enables them to promote an antiviral state. Both hypotheses imply a critical role of the viral macrodomain and its MAR hydrolase function.Therefore, I propose to clarify the biochemical and physiological consequences of ARTD10-catalyzed MARylation of the nsPs. This will involve mapping the modification sites, addressing the impact of MARylation on specific functions of the individual nsPs, and in consequence on the viral life cycle. In addition, we aim at the identification of host factors regulated by MARylation. To determine those, we will identify common substrates of ARTD10 and the CHIKV nsP3 hydrolase. A further selection criterion will be whether the MARylation of a particular substrate is subject to IFN regulation. I assume that these substrates are important host factors that we plan to investigate regarding their relevance for viral replication. This will involve proteomic studies, the analysis of MARylation of individual cellular proteins, enzymatic MARylation and de-MARylation assays, the knock-down and knock-out of cellular host factors and the consequences on the viral life cycle. Together these studies will define how MARylation is used in antiviral defense, and how CHIKV antagonizes this innate immune strategy. The findings of these studies will be relevant to define novel entry points for antiviral therapies.

进入宿主细胞的病毒被先天免疫系统识别。为了有效复制，病毒进化出了对抗免疫系统和劫持细胞过程的策略。最近的研究结果表明，单ADP核糖基化（MARylation），一种翻译后修饰，是先天免疫反应的一部分。本研究旨在明确MARylation在哺乳动物宿主与基孔肯雅病毒（Chikungunya virus，CHIKV）冲突中的作用。目前既没有疫苗也没有治疗方法。CHIKV仅编码4种非结构蛋白（nsPs），其中nsP 3含有一个宏结构域，一种保守的蛋白折叠，与MARylation密切相关。MARylation涉及ADP-核糖从NAD+转移到底物蛋白上，并且在细胞内主要由ADP-核糖基转移酶白喉毒素样（ARTD）催化。我们专注于ARTD 10，这是诱导响应I型干扰素（IFN）。我们将CHIKV宏结构域鉴定为去MARylating酶，其是有效病毒复制所需的。此外，初步工作表明nsP 1-nsP 3是ARTD 10的底物，因此它们的活性可能受MAR化控制。这些发现使我们假设MARylation在先天免疫中的作用，我们提出以两种不同的方式发挥作用：（I）CHIKV nsP的宿主依赖性MARylation直接影响其功能，从而影响病毒生命周期。(II)宿主因子的MARylation使它们能够促进抗病毒状态。这两个假设都暗示了病毒宏结构域及其MAR水解酶功能的关键作用。因此，我建议澄清ARTD 10催化的nsPs的MAR化的生化和生理后果。这将涉及映射的修饰位点，解决MARylation对个别nsPs的特定功能的影响，并因此对病毒的生命周期。此外，我们的目标是确定由MARylation调节的宿主因子。为了确定这些，我们将鉴定ARTD 10和CHIKV nsP 3水解酶的共同底物。另一个选择标准是特定底物的MAR化是否受IFN调节。我假设这些底物是重要的宿主因子，我们计划研究它们与病毒复制的相关性。这将涉及蛋白质组学研究、单个细胞蛋白质的MARylation分析、酶MARylation和de-MARylation测定、细胞宿主因子的敲低和敲除以及对病毒生命周期的影响。这些研究将共同定义MARylation如何用于抗病毒防御，以及CHIKV如何拮抗这种先天免疫策略。这些研究的结果将有助于确定抗病毒治疗的新切入点。