ExRNA composition in SLE patients and factors influencing exRNP abundance and th

SLE患者ExRNA组成及影响exRNP丰度和th的因素

• 批准号: 8590490
• 负责人: THOMAS TUSCHL
• 金额: $ 43.64万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-01 至 2018-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8590490
• 关键词: Affect; Amyotrophic Lateral Sclerosis; Antibodies; Antinuclear Antibodies; Arginine; Autoantibodies; Autoantigens; Autoimmune Diseases; B-Lymphocytes; Biological Assay; Cell Culture Techniques; Cells; Cellular Stress; Clinical; Cultured Cells; Cytokine Activation; Cytoplasmic Granules; DNA Polymerase II; DNA Polymerase III; Development; Disease; Fluorescence; Fractionation; Frontotemporal Dementia; Gene Mutation; Genetic Predisposition to Disease; Heat-Shock Response; Human; Immune response; Inclusion Bodies; Interferon Activation; Lead; Lupus; Monitor; Myopathy; Natural Immunity; Nuclear; Nuclear RNA; Organ; Oxidative Stress; Pathogenesis; Pathologic; Patients; Peptide Hydrolases; Pharmaceutical Preparations; Phosphorylation; Play; Post-Translational Protein Processing; Process; Production; Property; Protein Family; Proteins; RNA; RNA Processing; RNA Splicing; RNA-Binding Proteins; Regulation; Resolution; Ribonucleases; Ribonucleoproteins; Role; Serological; Serum; Small Nuclear Ribonucleoproteins; Staging; Stress; Structure; Systemic Lupus Erythematosus; Testing; Tissues; Transcript; Ubiquitination; Variant; Virus Diseases; base; biochemical model; body system; extracellular; immunogenic; mouse model; novel; polyglutamine; research study; tissue culture; ultraviolet irradiation

The broad aim of this project is to identify possible triggers of systemic lupus erythematosus (SLE) and evaluate if extracellular RNA (exRNA) and ribonucleoproteins (RNPs), which accumulate under stress conditions in cytoplasmic granules, can stimulate innate immune responses and result in the formation of autoantibodies. Genetic mutations in factors facilitating RNP granule formation or inhibiting their resolution may further influence their immunostimulatory properties, including size, structure, or local concentration in tissues. Posttranslational modification of nuclear RBPs and RNA-processing factors including phosphorylation, ubiquitination, arginine dimethylation, and protease cleavage, as well as the associated RNAs and their specific sequences and structures accompanying various maturation stages, may be critical to distinguish normal from pathologic and immunogenic RNPs. The specific aims are: (1) Characterize the exRNA composition in SLE patients with distinct classes of antibodies. Examine if exRNA composition of cultured immortalized B cells from SLE patients mirrors variations detected in serum exRNA composition. Assay the effect of cellular stress on the immunostimulatory properties of immortalized B cell lysates or tissue culture supernatants obtained under normal and stress conditions, and identify relevant RNPs by fractionation experiments. (2) Examine the influence of extracellular and intracellular RNases on exRNA abundance and composition, in existing mouse models, and investigate if any of these profiles resembles the exRNA composition of SLE patients, arguing for a role of RNases in the pathogenesis of SLE. (3) Develop human cell culture models for the biochemical isolation and characterization of stress-related RNA granules. Test various drugs to reproducibly induce specific stress conditions (e.g. oxidative stress, heat shock or translational arrest) and use fluorescence-based assays to monitor dynamic effects on RNA granule formation. Establish assays for interferon and cytokine activation upon application of the aforementioned stress conditions using cultured cells as well as primary cells.

该项目的主要目标是确定系统性红斑狼疮(SLE)和 评估细胞外RNA(ExRNA)和核糖核蛋白(RNPs)在应激状态下是否积累 在细胞质颗粒的条件下，可以刺激先天免疫反应并导致 自身抗体。促进RNP颗粒形成或抑制其分解的因素的基因突变 可能进一步影响它们的免疫刺激特性，包括大小、结构或局部浓度。 纸巾。核限制性商业惯例和RNA加工因子的翻译后修饰，包括 磷酸化、泛素化、精氨酸二甲基化和蛋白酶裂解，以及相关的 伴随着不同成熟阶段的RNA及其特定的序列和结构可能是关键的 区分正常、病理和免疫原性RNPs。 具体目标是： (1)用不同类型的抗体表征SLE患者的exRNA组成。检查是否 培养的SLE患者永生化B细胞的exRNA组成反映了血清中检测到的变化 ExRNA组成。细胞应激对永生化细胞免疫刺激特性的影响 在正常和应激条件下获得的B细胞裂解物或组织培养上清，并鉴定 通过分馏实验得到相应的RNPs。 (2)检测胞外和胞内核糖核酸酶对外源RNA丰度和组成的影响 现有的小鼠模型，并调查这些图谱中是否有任何类似于SLE的exRNA组成 患者，主张核糖核酸酶在SLE发病机制中的作用。 (3)建立与应激相关的生物化学分离和表征的人细胞培养模型 核糖核酸颗粒。测试各种药物以重复地诱导特定的应激条件(例如，氧化应激， 热休克或翻译停滞)，并使用基于荧光的分析来监测对RNA的动态影响 颗粒形成。建立干扰素和细胞因子激活的检测方法 利用培养细胞和原代细胞进行上述应激条件的研究。