Regulation and Function of Nuclear cGAS

核cGAS的调节和功能

• 批准号: 10686032
• 负责人: Daniel B Stetson
• 金额: $ 62.39万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-22 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10686032
• 关键词: Amino Acids; Applications Grants; Autoimmune Diseases; Base Pairing; Biology; Biotinylation; Cell Cycle; Cells; Chimeric Proteins; Chromatin; Cytosol; DNA; DNA Virus Infections; DNA Viruses; Detection; Discrimination; Genomic DNA; Host Defense; Human; Human Cell Line; Infection; Knock-in Mouse; Label; Ligands; Mediating; Mus; Mutation; Nature; Nuclear; Nuclear Protein; Nucleic Acids; Pathway interactions; Phase; Proteins; RNA; Regulation; Resistance; Rest; Sodium Chloride; Stimulator of Interferon Genes; Surface; Tumor Immunity; Untranslated RNA; Viral Proteins; Virus Diseases; autoreactivity; innovation; insight; mouse model; new therapeutic target; prevent; response; sensor; temporal measurement; tool

Project Summary/Abstract Detection of intracellular DNA by the cGAS-STING pathway mediates host defense against DNA viruses, contributes to specific human autoimmune diseases, and can be harnessed to promote anti-tumor immunity. The presence of billions of base pairs of genomic DNA in all nucleated cells raises the question of how cGAS is not constitutively activated. A widely accepted explanation for this is the sequestration of cGAS in the cytosol, which is thought to prevent cGAS from accessing nuclear DNA. We have found that endogenous cGAS is predominantly a nuclear protein, regardless of cell cycle phase or cGAS activation status. We show that nuclear cGAS is tethered tightly by a salt-resistant interaction. This tight tethering is independent of the domains required for cGAS activation, and it requires intact nuclear chromatin. We identify the evolutionarily conserved tethering surface on cGAS and we show that mutation of single amino acids within this surface renders cGAS massively and constitutively active against self-DNA. Thus, tight nuclear tethering maintains the resting state of cGAS and prevents autoreactivity, revealing a fundamental mechanism that controls the resting state of cGAS. In this grant application, we have developed innovative new tools to define the cGAS nuclear tethering compartment, quantitate changes in cGAS disposition during DNA virus infection, and explore cGAS- associated nucleic acids before and after its activation in the context of infection and autoimmune disease. Our studies will uncover fundamental new aspects of cGAS biology, will identify new targets for therapeutic modulation of cGAS activity, and will provide new insights into the principles of self/non-self discrimination by intracellular nucleic acid sensors.

项目概要/摘要 通过 cGAS-STING 途径检测细胞内 DNA 介导宿主对 DNA 病毒的防御， 有助于特定的人类自身免疫性疾病，并可用于促进抗肿瘤免疫。 所有有核细胞中都存在数十亿个基因组 DNA 碱基对，这引发了 cGAS 如何实现这一问题的问题。 没有被组成性激活。对此的一个广泛接受的解释是 cGAS 被隔离在细胞质中， 这被认为可以阻止 cGAS 接触核 DNA。我们发现内源性cGAS 主要是核蛋白，无论细胞周期阶段或 cGAS 激活状态如何。我们表明 核 cGAS 通过耐盐相互作用紧密束缚。这种紧密的束缚独立于 cGAS 激活所需的结构域，并且需要完整的核染色质。我们从进化角度识别 cGAS 上的保守束缚表面，我们表明该表面内单个氨基酸的突变 使 cGAS 对自身 DNA 具有大规模和组成性的活性。因此，紧密的核束缚维持了 cGAS 的静息状态并防止自身反应，揭示了控制静息状态的基本机制 cGAS 状态。 在本次拨款申请中，我们开发了创新的新工具来定义 cGAS 核束缚 区室，定量 DNA 病毒感染期间 cGAS 处置的变化，并探索 cGAS- 在感染和自身免疫性疾病的情况下激活前后的相关核酸。我们的 研究将揭示 cGAS 生物学的基本新方面，并将确定治疗的新靶点 cGAS 活动的调节，并将为自我/非自我歧视的原则提供新的见解 细胞内核酸传感器。