Biochemical and structural characterization of RNA turnover processes and resolu

RNA 周转过程和解析的生化和结构表征

基本信息

批准号：
8590500
负责人：
THOMAS TUSCHL
金额：
$ 27.78万
依托单位：
ROCKEFELLER UNIVERSITY
依托单位国家：
美国
项目类别：
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/8590500
关键词：
Amyotrophic Lateral Sclerosis Antinuclear Antibodies Autoantibodies Autoantigens Autoimmune Diseases Autoimmunity Binding Biochemical Biological Assay Cells Cellular Stress Complex Cytoplasmic Granules DNA Polymerase I DNA Polymerase II DNA Polymerase III DNA-Directed RNA Polymerase Disease Endoribonucleases Event Frontotemporal Dementia Functional RNA Genes Human Immune response Immune system Inclusion Bodies Lead Link Mammalian Cell Messenger RNA Molecular Myopathy Nucleic Acids Pathologic Pattern Pattern recognition receptor Phagocytes Process Production Property Proteins RNA RNA Binding RNA Processing RNA Viruses RNA-Binding Proteins Recombinant Proteins Regulation Research Resolution Ribonucleoproteins Ribosomal RNA Role Serum Small Nuclear RNA Source Stress Structure Systemic Lupus Erythematosus Transcript Transfer RNA cell type endoribonuclease extracellular in vivo polyglutamine programs protein complex receptor uptake

项目摘要

Project Leader: Dinshaw Patel. The broad aim of this project is the biochemical and structural characterization of the natural function of a class of autoantigens that are released upon stress and may trigger strong innate immune responses that ultimately result in autoantibody production. Extracellular RNA is a potent activator of the innate immune system sensed by binding to endosomal and intracellular pattern recognition receptors present in most cells including phagocytic cells. The most well known sources of immunostimulatory extracellular RNA are viruses. However, RNA sensing receptors by themselves cannot discriminate stringently against self-nucleic acids. Thus, events of cellular stress that lead to the uptake of host cell RNA into neighboring cells, which then activates pattern recognition receptors, may represent a plausible mechanism by which autoimmunity is initiated. Autoimmune disorders are characterized by the presence of high titers of antinuclear antibodies (ANA), and may be the pathologic manifestations of inappropriate activation of sensing mechanisms by stress-induced RNA-containing granules. We propose to: (1) Identify the RNA binding targets of RBP autoantigens including SSB/La, TROVE2/SSA2/Ro60 and TRIM21/SSA1/Ro52 under normal and stress conditions using PAR-CLIP. Develop new biochemical assays that recapitulate the formation of RNA granules or their resolution using recombinant proteins. (2) Study the role of pol III RNA transcript binding factors during stress and the specific structure and function of SSB/La, TROVE2/SSA2/RO60 and TRIM21/SSA1/Ro52 with their in vivo RNA targets. Investigate how stress conditions influence the processing patterns of tRNAs, e.g. polyuridylation. (3) Determine the mechanism and structure of pol III RNA transcript processing involving the tRNA-specific endoribonuclease TSN-TSNAX/C3P0 and pattern recognition receptor RIG-1 and MDA5 complexes with their in vivo RNA targets, and investigate their perturbation under stress conditions

项目负责人：Dinshaw Patel。该项目的广泛目的是在压力下释放的一类自身抗原的自然功能的生化和结构表征，并可能触发强烈的先天免疫反应，最终导致自身抗体的产生。细胞外RNA是先天免疫的有效激活剂通过与大多数细胞中存在的内体和细胞内模式识别受体结合而感受的系统包括吞噬细胞。免疫刺激性细胞外RNA的最著名来源是病毒。但是，RNA感应受体本身不能严格歧视自我核酸酸。因此，导致宿主细胞RNA摄入邻近细胞的细胞应激事件，然后激活模式识别受体，可能代表自身免疫性的合理机制发起。自身免疫性疾病的特征是存在抗核抗体的高滴度（ANA），并且可能是通过应力诱导的含RNA的颗粒。我们建议：（1）确定RBP自动抗原的RNA结合靶标，包括SSB/LA，TROVE2/SSA2/RO60和 TRIM21/SSA1/RO52在正常和应力条件下使用PAR-CLIP。开发新的生化测定这概括了使用重组蛋白的RNA颗粒形成或它们的分辨率。（2）研究Pol III RNA转录因子在压力和特定结构和功能中的作用 SSB/LA，TROVE2/SSA2/RO60和TRIM21/SSA1/RO52的体内RNA靶标。调查如何压力条件会影响TRNA的加工模式，例如多尿酰化。（3）确定涉及tRNA特异性的POL III RNA转录处理的机理和结构内切核酸酶TSN-TSNAX/C3P0以及模式识别受体RIG-1和MDA5复合物及其体内RNA靶标，并在应力条件下研究其扰动