Definition of Serum Ribonucleoprotein Composition and its Regulation and Function

描述（由申请人提供）： 来自病原体的胞外RNA（exRNA）作为天然免疫的激活剂在哺乳动物中起着关键作用。然而，宿主exRNA在血清和其他体液中的存在挑战了目前基于RNA的免疫识别模型。为了理解其特异性的基础，重要的是要对血清中的宿主exRNA及其相关蛋白进行编目，研究导致循环核糖核蛋白（RNP）稳态的机制，并鉴定有助于细胞RNA释放的蛋白质因子。RNA靶点或相互作用蛋白的遗传改变可能导致RNP正常与应激触发释放的不平衡，并将适应性长期病原体导向的免疫推向自身免疫。ExRNA也可能在细胞外信号传导中发挥更广泛的作用，类似于肽激素，这也将被这种实验方法捕获。该应用程序汇集了一个由多名研究人员组成的团队，他们具有互补的专业知识和密切合作的历史，为细胞外RNA的识别，机制和功能以及参与其生物发生，出口，识别和营业额的蛋白质奠定了坚实的基础。拟议项目的具体目标是：1。使用各种已建立和新型RNA seq方法对正常受试者的人血清中的所有类别的细胞外RNA进行分类和定量，并检查个体内、个体之间循环RNA谱的正常变异性以及性别、年龄、种族和疾病的影响。建立处理和归档临床材料的核心设施（威廉姆斯、普特曼、图斯基）。2.确定系统性红斑狼疮（SLE）患者的exRNA组成，对他们来说，针对不同类型RBP的抗体是一个标志。考虑到这些RNP在应激时改变其亚细胞定位，并出现在具有未成熟RNA和/或靶向周转的RNA的应激颗粒中，我们将尽可能多地评估从正常和SLE受试者的永生化B细胞收获的应激颗粒中RNP的组成具有免疫刺激功能，以及这些RNP颗粒是否也在应激期间释放（Tuschi，Putterman，威廉姆斯）。3.发展应激颗粒形成和RNA/RNP介导的先天免疫反应的分子和机制的理解。确定tRNA应激反应中自身抗原的RNA靶点和RNP结构，它们的周转率和它们的免疫受体。

DESCRIPTION (provided by applicant): Extracellular RNA (exRNA) from pathogens plays a key role as activator of innate immunity in mammals. However, the presence of host exRNA in serum and other body fluids challenges the current models of RNA based immune recognition. To understand its basis of specificity, it is important to catalog host exRNAs and their associated proteins in serum, investigate mechanisms leading to circulating ribonucleoprotein (RNP) homeostasis, and identify protein factors contributing to cellular RNA release. Genetic alterations in RNA targets or interacting proteins may contribute to imbalances in normal versus stress-triggered release of RNPs and push adaptive long-term pathogen-directed immunity towards autoimmunity. ExRNAs may also play a broader role in extracellular signaling similar to peptide hormones, which would also be captured by this experimental approach. This application brings together a team of multiple investigators with complementary expertise and history of close collaboration to build a solid foundation regarding identification, mechanism and function of extracellular RNAs and the proteins involved in their biogenesis, export, recognition, and turnover. The specific aims of the proposed project are: 1. Catalogue and quantify all classes of extracellular RNAs in human serum from normal subjects using various established and novel RNA seq approaches and examine normal variability of circulating RNA profiles within an individual, between individuals and the influence of gender, age, race, and disease. Establish a core facility for processing and archiving clinical materials (Williams, Putterman, Tuschi). 2. Determine exRNA composition in patients suffering from systemic lupus erythematosus (SLE), for whom antibodies against different classes of RBPs is a hallmark. Considering that these RBPs alter their subcellular localization upon stress and appear in stress granules with immature RNA and/or RNA targeted for turnover, we will evaluate in as much the composition of RNPs in stress granules harvested from immortalized B cells of normal and SLE subjects possesses immunostimulatory function and if these RNP granules are also released during stress (Tuschi, Putterman, Williams). 3. Develop a molecular and mechanistic understanding of stress granule formation and RNA/RNP mediated innate immune responses. Identify the RNA targets and RNP structures of autoantigens in tRNA stress responses, their turnover, and their immune receptors.