Exploring Determinants of Intrinsic DNA Load Towards Inflammaging

探索导致炎症的内在 DNA 负荷的决定因素

• 批准号: 10373598
• 负责人: Yuk Yuen Lan
• 金额: $ 24.66万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-03-01 至 2024-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10373598
• 关键词: Abate; Aging; Algorithms; Ataxia; Ataxia Telangiectasia; Autoimmune; Autoimmunity; Automobile Driving; Autophagocytosis; Biological; Biological Assay; Biological Process; Biology; Cell Aging; Cell Nucleus; Cells; Cellular Morphology; Chromatin; Chronic; Clustered Regularly Interspaced Short Palindromic Repeats; Cyclic GMP; Cytosol; DNA; DNA Damage; DNA Virus Infections; Diabetes Mellitus; Disease; Exhibits; Family; Foundations; Future; Genomic Instability; Heart Diseases; Homeostasis; Human; Immune; Immune response; Immunity; Immunofluorescence Immunologic; Impaired cognition; Infection; Inflammaging; Inflammation; Inflammatory; Interferon Type I; Interferons; Intrinsic factor; Knock-out; Knockout Mice; Ligands; Malignant Neoplasms; Mediating; Mitochondrial DNA; Modeling; Mus; Nuclear; Optics; Pathologic; Pathology; Pathway interactions; Patients; Pattern; Phenotype; Play; Ploidies; Polyarthritides; Premature aging syndrome; Production; Progeria; Property; Retrotransposon; Risk; Role; Signal Transduction; Sterility; Stimulator of Interferon Genes; System; Tissues; base; cell growth; clinically relevant; ds-DNA; endonuclease; extranuclear DNA; immune activation; immunogenic; improved; insight; member; novel; novel strategies; novel therapeutic intervention; nuclease; overexpression; response; senescence; sensor; systemic inflammatory response; theories; transcriptomics

PROJECT SUMMARY Sterile inflammation is exhibited in aging‐related conditions, including heart disease, cancer, diabetes, and cognitive decline. Such hyperinflammation typically displays an altered immune response comprising a type I interferon response and downstream senescence-associated secretory phenotype (SASP). The intrinsic factors that induce this chronic low-grade inflammation are not well understood. In studying the initiation of autoimmune polyarthritis in mice lacking the lysosomal endonuclease Dnase2a, we discovered a cell‐autonomous pathway through which damaged nuclear DNA is trafficked to the cytosol where it is recognized by the innate cytosolic DNA sensing cGAS-STING axis, triggering an inflammatory cascade. In healthy cells, nuclear DNA found in the cytosol is removed via autophagy for lysosomal degradation by DNASE2A, which degrades double-stranded DNA (dsDNA)—ligand for the DNA sensor cGAS. Consistent with this notion that damaged or irreparable nuclear DNA is a trigger of immunity when mis- localized and accumulated in the cytosol, elevated loads of extranuclear DNA in replicative senescent cells and cells from ataxia (AT) and progeria (HGPS) patients engage the same DNA sensing pathway leading to persistent inflammation. Controlling the intrinsic DNA load is thus critical to avoid sterile inflammation. While nucleases can clear DNA, unknown DNA export factors may facilitate DNA exit to the cytosol. By targeting curated sets of potential nucleases and DNA export factors, we propose to identify unknown intrinsic DNA controlling factors in a single-cell based CRISPR knockout optical screen by assaying dsDNA content using immunofluorescence. Hits will be validated for their DNA export or degradative function and ranked for their involvement in senescence and STING activation downstream of cGAS. Selected tops hits, along with DNASE2A, will be evaluated for their capacity in suppressing inflammation and SASP in senescent cells and AT and HGPS patient cells. In Dnase2a knockout mice, we will investigate immune cell phenotype in tissues to understand the mechanisms underlying the systemic senescent pathology. The identification and characterization of new determinants in inflammation and senescence will add new insights to the basic biology of DNA-mediated immunity and open up novel strategies in modulating inflammation to benefit aging-associated pathologies, including interferonopathy and laminopathy, autoimmunity and cancer.

项目摘要 无菌性炎症在与衰老相关的疾病中表现出来，包括心脏病、癌症、 糖尿病和认知能力下降这种过度炎症通常表现为免疫功能改变， 包括I型干扰素应答和下游衰老相关的干扰素应答 分泌表型（SASP）。诱发这种慢性低度炎症的内在因素 并没有得到很好的理解。在研究小鼠自身免疫性多发性关节炎的启动时， 溶酶体核酸内切酶Dnase 2a，我们发现了一个细胞自主途径， 受损的核DNA被运输到胞质溶胶，在那里它被先天性胞质溶胶识别。 DNA传感cGAS-STING轴，触发炎症级联反应。在健康细胞中， 在细胞溶质中发现的DNA通过自噬被去除，用于通过DNASE 2A的溶酶体降解， 其降解DNA传感器cGAS的双链DNA（dsDNA）-配体。一致 这种观念认为，受损或不可修复的核DNA是免疫力的触发器，当错误的- 在细胞质中定位和积累，在复制细胞中增加核内DNA的负荷， 衰老细胞和来自共济失调（AT）和早衰症（HGPS）患者的细胞接合相同的DNA 导致持续性炎症的传感通路。因此，控制固有DNA负载是 对于避免无菌炎症至关重要。虽然核酸酶可以清除DNA，但未知的DNA输出因子 可以促进DNA离开细胞质。通过靶向潜在的核酸酶和DNA 出口因素，我们建议确定未知的内在DNA控制因素，在一个单一的细胞 基于CRISPR敲除的光学筛选通过使用免疫荧光测定dsDNA含量来进行。 将验证命中的DNA输出或降解功能，并根据其参与程度进行排名 在cGAS下游的衰老和STING激活中。选定的顶部命中，沿着与 将评价它们在小鼠中抑制炎症和SASP的能力。 衰老细胞以及AT和HGPS患者细胞。在Dnase 2a敲除小鼠中，我们将研究 组织中的免疫细胞表型，以了解系统性 衰老病理学炎症中新决定因素的鉴定和表征 和衰老将为DNA介导的免疫和开放的基础生物学增加新的见解。 提出调节炎症的新策略，以有益于衰老相关的病理学，包括 干扰素病和核纤层蛋白病、自身免疫和癌症。