K63-linked ubiquitination regulates cGAS activation upon DNA damage

K63 连接的泛素化在 DNA 损伤时调节 cGAS 激活

• 批准号: 10348290
• 负责人: Shitao Li
• 金额: $ 22.8万
• 依托单位: TULANE UNIVERSITY OF LOUISIANA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-24 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10348290
• 关键词: Antiviral Agents; Ataxia Telangiectasia; Ataxia Telangiectasia Patients; Autoimmune Diseases; Binding; Cell Nucleus; Chromatin; Communicable Diseases; Cyclic GMP; Cytosol; DNA; DNA Binding; DNA Damage; DNA Viruses; DNA-Binding Proteins; Data; Defect; Disease; Fibroblasts; Foundations; Gene Expression; Goals; Histones; Host Defense; Immobilization; Interferon Type I; Interferons; Knock-out; Ligation; Link; Literature; Mediating; Modeling; Natural Immunity; Nuclear; Nucleic Acids; Nucleosomes; Play; Polyubiquitin; Proteomics; Role; Second Messenger Systems; Signal Pathway; Signal Transduction; Site; Structure; Systemic Lupus Erythematosus; Testing; Therapeutic; Ubiquitination; Virus Diseases; autoinflammatory; base; cell injury; novel therapeutic intervention; pathogen; protein complex; response; sensor; ubiquitin-protein ligase; viral DNA

Project summary Excessive DNA either from pathogen or host activates the cytosolic DNA sensor, cyclic GMP-AMP synthase (cGAS) that produces the second messenger, cyclic GMP-AMP (cGAMP). cGAMP triggers a signal cascade leading to type I interferon (IFN) expression. It has been thought that cGAS is sequestered in the cytosol and cannot access host DNA normally restricted in cellular compartments, such as the nucleus. The recent discovery of nuclear cGAS has dramatically changed this model. Structural studies showed cGAS binds to the H2A-H2B heterodimer of the nucleosome and the binding immobilizes cGAS on the chromatin; thus, cGAS cannot access the nearby DNA. However, the micronuclei caused by DNA damage are immunostimulatory, and the chromatin fragments in micronuclei activate cGAS. Currently, we don’t know whether cGAS is still immobilized on the nucleosomes in the micronuclei and how cGAS is activated by the interaction with chromatin upon DNA damage. Endogenous defects in the DNA damage response (DDR) induce autoinflammatory diseases, such as Ataxia-Telangiectasia (A-T). Thus, there is a critical need to elucidate the mechanisms controlling cGAS activation upon DNA damage. Our objective is to define the role of K63-linked ubiquitination in DNA damage-induced cGAS activation and elucidate the underlying mechanisms by which K63-linked polyubiquitin regulates cGAS activity. Our preliminary data found that cGAS was subject to K63-linked ubiquitination upon DNA damage and inhibition K63-linked ubiquitination abrogated DNA damage-induced cGAS signaling pathway and interferon-stimulated gene expression. Based on the existing literature and our preliminary data, we hypothesize that K63-linked polyubiquitin facilitates cGAS activation in the micronuclei. We will examine two aims: First, we will determine the role of K63-linked ubiquitination in cGAS activation. Second, we will determine the mechanisms by which K63-linked polyubiquitin facilitates DNA damage-induced cGAS activation. The chromatin fragments in the micronuclei are a strong immunostimulatory agent for cGAS, but how cGAS overcome the histone sequestration and competes with other DNA binding proteins to access the DNA has yet to be determined. This exploratory R21 application proposes that K63-linked ubiquitination facilitates cGAS activation upon DNA damage. Overall, this study elucidates the regulatory mechanisms of nuclear cGAS activation and provides foundations for developing novel therapeutic strategies for infectious and autoimmune diseases.

项目摘要 病原体或宿主的过量DNA激活胞质DNA传感器，环状GMP-AMP合酶 （CGA）产生第二个信使GMP-AMP（CGAMP）。 CGAMP触发信号级联 导致I型I Interferon（IFN）表达式。人们认为CGA被隔离在细胞质中， 无法访问通常在细胞室中受到限制的宿主DNA，例如核。最近 核CGA的发现已极大地改变了该模型。结构研究表明CGA与 核小体的H2A-H2B异二聚体和结合将CGA固定在染色质上；因此，CGA 无法访问附近的DNA。然而，由DNA损伤引起的微核是免疫刺激性的， 微核中的染色质片段激活CGA。目前，我们不知道CGA是否还在 固定在微核中的核小体上，以及如何通过与 DNA损伤时染色质。 DNA损伤反应（DDR）影响的内源性缺陷 自身炎症性疾病，例如共济失调 - 舌张症（A-T）。那是阐明的迫切需要 DNA损伤后控制CGA激活的机制。 我们的目的是定义K63连接的泛素化在DNA损伤引起的CGAS激活中的作用和 阐明了K63连接多泛素调节CGAS活性的基本机制。我们的 初步数据发现，在DNA损伤和抑制时，CGA受到K63连接的泛素化 K63连接的泛素化废除了DNA损伤诱导的CGAS信号通路和干扰素刺激 基因表达。根据现有文献和我们的初步数据，我们假设K63链接 多泛素在微核中宿主CGAS活化。我们将研究两个目标：首先，我们将确定 K63连接的泛素化在CGAS激活中的作用。第二，我们将确定该机制 K63连接的多泛素托管DNA损伤诱导的CGAS活化。 微核中的染色质片段是CGA的强烈免疫刺激剂，但CGA如何 克服组蛋白隔离并与其他DNA结合蛋白竞争以访问DNA 要确定。这项探索性R21应用程序提案提出了K63连接的泛素化设施CGA DNA损伤激活。总体而言，这项研究阐明了核CGA的调节机制 激活并为开发新型治疗策略的基础，以进行感染和自身免疫 疾病。