Toll-like receptor 9: trafficking and signaling

Toll 样受体 9：运输和信号转导

基本信息

批准号：
8384885
负责人：
CYNTHIA A LEIFER
金额：
$ 31.92万
依托单位：
CORNELL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-12-01 至 2014-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8384885
关键词：
Accounting Address Adjuvant Affect Allergic Disease Autoimmune Diseases Autoimmune Process Autoimmunity B-Lymphocytes Biological Assay Bypass Carbohydrates Cell Fractionation Cells Communicable Diseases Cytoplasmic Tail DNA Data Dendritic Cells Development Discrimination Dominant-Negative Mutation Endocytosis Endoplasmic Reticulum Endosomes Event Goals Golgi Apparatus Immune response Immune system In Vitro Interphase Cell Knowledge Lysine Lysosomes Malignant Neoplasms Mass Spectrum Analysis Methods Modeling Modification Molecular Movement Mutate Nucleic Acids Pathology Pathway interactions Pattern Peptide Hydrolases Phosphorylation Play Post-Translational Protein Processing Proteins Receptors, Antigen, B-Cell Regulation Role Secretory Cell Signal Transduction Site Small Interfering RNA Systemic Lupus Erythematosus TLR9 gene Testing Therapeutic Tyrosine Ubiquitin Ubiquitination Vaccines base microbial microorganism mutant novel receptor response synergism trafficking

项目摘要

ABSTRACT DNA containing CpG motifs (CpG DNA) has incredible potential to treat cancer, infectious and allergic diseases. Despite this potential, response to our own nucleic acids, including DNA, triggers autoimmune diseases such as systemic lupus erythematosus. Localization and trafficking of the specific receptor, TLR9, may play a key role in self/foreign DNA discrimination. Uncovering the molecular mechanisms of TLR9 trafficking is the first step towards our long term goal of manipulating TLR9 trafficking to achieve modulation of CpG DNA response. We have shown that TLR9 is localized intracellularly, predominantly in the endoplasmic reticulum (ER), prior to CpG DNA stimulation. TLR9 traffics from the ER to endosomes and lysosomes where it co-localizes with endocytosed CpG DNA. These data raise the fundamental questions of what regulates TLR9 access to CpG DNA and how does access affect response to self and microbial DNA? Using new TLR9 trafficking assays, we have accumulated evidence that TLR9 constitutively exits the ER, that CpG DNA induces a secondary TLR9 trafficking event, and that both events occur through traditional cell trafficking pathways. Therefore, we hypothesize that TLR9 constitutively traffics in a highly regulated fashion through the cell secretory pathway from the Golgi complex to endolysosomes and that Golgi transit is a prerequisite for TLR9 response to CpG DNA. We believe that TLR9 trafficking may account for synergy with other TLRs and the autoimmune B cell receptor, and is regulated by post-translational modification of the TLR9 cytoplasmic tail. This hypothesis will be tested in three Specific Aims. First, transfected and endogenous TLR9 trafficking through the Golgi complex will be examined using multiple in vitro approaches including a novel protease cleavage assay. Second, we will examine the mechanism of TLR9-autoimmune B cell receptor synergism. Third, we will determine the role of TLR9 post-translational modification in regulating intracellular trafficking and in TLR-autoimmune B cell receptor synergy. By understanding the mechanism of TLR9 trafficking we can uncover how regulation of response to foreign DNA and lack of response to self-DNA is achieved. This knowledge will provide the basis for the further development of CpG DNA as an adjuvant and therapeutic as well as develop mechanisms to interrupt the cycle of autoimmune pathology.

抽象的含有CPG基序（CpG DNA）的DNA具有难以置信的治疗癌症，感染性和过敏性的潜力疾病。尽管有潜力，但对我们自己的核酸的反应，包括DNA，触发自身免疫性诸如全身性红斑狼疮等疾病。特定受体TLR9的定位和运输可能在自我/外国DNA歧视中起关键作用。发现TLR9的分子机制贩运是朝着操纵TLR9贩运以实现的长期目标的第一步 CpG DNA响应的调节。我们已经证明TLR9是细胞内局部的，主要是在在CpG DNA刺激之前，内质网（ER）。 TLR9从急诊室到内体的运输者和它与内吞CpG DNA共定位的溶酶体。这些数据提出了是什么调节TLR9访问CpG DNA的访问，访问如何影响对自我和微生物DNA的反应？使用新的TLR9贩运分析，我们积累了证据，表明TLR9组成型退出ER，表明 CpG DNA诱导了次级TLR9运输事件，并且这两个事件都通过传统细胞发生贩运途径。因此，我们假设TLR9在高度贩运中构成了运输通过细胞分泌途径从高尔基综合体到内侧化体和高尔基体过境是TLR9对CpG DNA响应的先决条件。我们相信TLR9的贩运可能会与其他TLR和自身免疫性造成协同作用细胞受体，并由TLR9细胞质尾巴的翻译后修饰调节。该假设将以三个特定目的进行检验。首先，转染和内源性TLR9运输通过多种体外方法（包括新型蛋白酶）将通过高尔基体复合物进行检查切割测定法。其次，我们将研究TLR9-Automune B细胞受体协同作用的机制。第三，我们将确定TLR9翻译后修饰在调节细胞内的作用贩运和TLR-AutoMune B细胞受体协同作用。通过了解TLR9的机制贩运我们可以发现对外国DNA反应的调节以及对自我DNA的缺乏的调节是如何的成就了。这些知识将为CpG DNA作为辅助者的进一步发展提供基础治疗和开发机制，以中断自身免疫性病理的周期。