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: 10576391
• 负责人: Yuk Yuen Lan
• 金额: $ 20.46万
• 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 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10576391
• 关键词: Abate; Acceleration; 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; Ploidies; Polyarthritides; Premature aging syndrome; Production; Progeria; Property; Retrotransposon; Risk; Role; Signal Transduction; Sterility; Stimulator of Interferon Genes; System; Tissues; cell growth; clinically relevant; ds-DNA; endonuclease; extranuclear DNA; immune activation; immunogenic; immunoregulation; improved; insight; member; novel; novel strategies; novel therapeutic intervention; nuclease; overexpression; patient engagement; 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.

项目总结 无菌炎症在与衰老相关的疾病中表现出来，包括心脏病、癌症、 糖尿病和认知功能减退。这种过度炎症通常表现为免疫功能的改变。 包括I型干扰素应答和下游衰老相关的应答 分泌表型(SASP)。导致这种慢性低度炎症的内在因素 都没有被很好地理解。在研究自身免疫性多发性关节炎小鼠缺乏 溶酶体内切酶Dnase2a，我们发现了一条细胞自主途径，通过它 受损的核DNA被运送到胞浆，在那里它被固有的胞浆识别 DNA感应cGAS刺轴，触发炎症级联反应。在健康细胞中，有核 在胞浆中发现的DNA通过自噬被DNASE 2A降解溶酶体而被移除， 它降解DNA传感器cGAS的双链DNA(DsDNA)配体。一致 认为受损或无法修复的核DNA是免疫的触发因素，当- 在胞浆中定位和积聚，复制体核外DNA负荷增加 共济失调(AT)和早衰症(HGPS)患者的衰老细胞和细胞具有相同的DNA 导致持续性炎症的感知途径。因此，控制固有的DNA载量 对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输出因素 可能会促进DNA进入胞浆。通过靶向精选的潜在核酸酶和DNA 输出因子，我们建议在单细胞中识别未知的内在DNA控制因子 用免疫荧光法测定dsDNA含量，建立CRISPR基因敲除光学屏幕。 HITS将根据其DNA输出或降解功能进行验证，并根据其参与程度进行排名 在cGAS下游的衰老和刺痛激活中。精选TOPS点击量，以及 将评估其在抑制炎症和SASP方面的能力 衰老细胞、AT和HGPS患者细胞。在Dnase2a基因敲除小鼠中，我们将研究 组织中免疫细胞表型以了解系统性红斑狼疮的发病机制 衰老的病理学。炎症中新决定因素的鉴定和表征 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 调整炎症以利于衰老相关病理的新策略，包括 干扰素病和椎板病、自身免疫和癌症。