Molecular Bases of Inflammasome Regulation Mediated by ASC Isoforms

ASC 异构体介导的炎症小体调节的分子基础

基本信息

批准号：
9810959
负责人：
Eva de Alba Bastarrechea
金额：
$ 44.34万
依托单位：
UNIVERSITY OF CALIFORNIA, MERCED
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-06-01 至 2023-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9810959
关键词：
Adaptor Signaling Protein Address Adopted Adult Respiratory Distress Syndrome Affect Amino Acid Sequence Amino Acids Atherosclerosis Autoimmune Diseases Behavior Binding CASP1 gene Cardiovascular Diseases Cells Characteristics Chemicals Chronic Complement Complex Cone Cytokine Activation Data Death Domain Disease Engineering Enzyme-Linked Immunosorbent Assay Fluorescence Anisotropy Fluorescence Spectroscopy Glues Goals Grant Homo Immune In Vitro Infection Inflammasome Inflammation Inflammatory Inflammatory Response Injury Innate Immune Response Innate Immune System Interleukin-18 Interleukins Kinetics Knowledge Length Life Macromolecular Complexes Mediating Mental Depression Mission Modeling Molecular Molecular Conformation Molecular Sieve Chromatography Molecular Structure Monitor Motion Movement Multiprotein Complexes Natural Immunity Nuclear Magnetic Resonance Nuclear Structure Peptides Play Property Protein Isoforms Proteins Publishing Regulation Relaxation Relaxation Techniques Resolution Rheumatoid Arthritis Role Signal Transduction Site Spectrometry, Mass, Electrospray Ionization Structure Testing Time Tissues Transmission Electron Microscopy United States National Institutes of Health Vertebral column base cancer type cytokine design experience experimental study fighting flexibility human disease innovation nervous system disorder organizational structure pathogen protein complex protein function recruit self assembly sensor stoichiometry therapeutic development therapeutic target

项目摘要

PROJECT SUMMARY/ABSTRACT. Inflammation is our primary response from the innate immune system to fight infection and self-protect from damage. However, dysfunctional regulation of inflammation results in disease, including certain types of cancer, autoimmune, cardiovascular and neurological disorders, rheumatoid arthritis, and even depression. The onset of inflammation depends on the assembly of a multiprotein complex known as the inflammasome. The main players in inflammasome assembly are; - sensor proteins that react upon danger signals derived from pathogens or damaged tissue; - procaspase-1 that activates inflammatory cytokines as a result of inflammasome assembly; - the adapter ASC that functions like a molecular glue by connecting sensor and procaspase-1 molecules. Canonical ASC has an isoform, which shows different self-assembly capabilities and modulates the intensity of inflammation in the cellular context. The presence of protein isoforms is a well-known, natural mechanism for the regulation of protein function. Both proteins are bimodular with two Death Domains connected by a linker, and their amino acid sequences differ solely in the linker length. Canonical ASC has a 23 amino acid-long linker, whereas its isoform has a shorter linker of 3 amino acids (ASC_short). We demonstrated that ASC linker plays a key role in defining the orientation of its domains, and show in this proposal that both domains actively participate in ASC self-assembly to form filamentous macrostructures. Evidence indicate that inflammasomes assemble into different supramolecular structures. However, little is known on the factors controlling these structural arrangements, or how the different assemblies impact inflammation. This proposal aims at addressing a knowledge gap in the role of ASC isoforms on inflammasome structural organization and inflammatory response. The long-term goal of this proposal is to gain in-depth knowledge on the interplay between the inflammasome components to understand its function and regulation, which is of great significance to set the grounds for the development of therapeutics to control inflammation. The objective of this grant is innovative because it will decipher the unknown molecular bases for inflammasome regulation mediated by ASC isoforms. Our hypothesis is that; 1) the linker length has important implications in the interaction properties of ASC and ASC_short at the molecular level, which can account for the observed alteration of the inflammatory response, 2) the linker length leads to differences in the interdomain dynamics of the two isoforms and in the resulting macrostructures. To test this hypothesis we propose to; 1) determine the inflammatory activity, precise self-association and interacting capabilities of ASC isoforms, including ASC, ASC_short and an artificial form of ASC engineered with a long linker (3 times the canonical linker length: ASC_long) as a reference for independent domains, 2) determine the interdomain dynamics of the three isoforms using Nuclear Magnetic Resonance; 3) discern with Transmission Electron Microscopy the potentially different characteristics of the macrostructures resulting from ASC isoforms self-association, and their implication in inflammatory activity.

项目摘要/摘要。炎症是我们先天免疫系统的主要反应对抗感染并自我保护免受伤害。然而，炎症调节功能失调会导致疾病，包括某些类型的癌症、自身免疫性疾病、心血管和神经系统疾病、类风湿病关节炎，甚至抑郁症。炎症的发生取决于多蛋白复合物的组装称为炎症小体。炎症小体组装的主要参与者是： - 反应的传感器蛋白来自病原体或受损组织的危险信号； - 激活炎症细胞因子的 procaspase-1 由于炎性小体组装的结果； - 适配器 ASC 通过连接发挥类似分子胶的作用传感器和 procaspase-1 分子。 Canonical ASC 有一个异构体，显示出不同的自组装能力并调节细胞环境中炎症的强度。蛋白质亚型的存在是一种众所周知的蛋白质功能调节天然机制。两种蛋白质都是双模块的，有两个死亡结构域通过接头连接，其氨基酸序列仅在接头长度上有所不同。典范 ASC 具有 23 个氨基酸长的接头，而其同种型具有较短的 3 个氨基酸的接头（ASC_short）。我们证明 ASC 链接器在定义其域的方向方面发挥着关键作用，并在本提案中显示这两个结构域都积极参与 ASC 自组装形成丝状宏观结构。证据表明炎症小体组装成不同的超分子结构。然而，人们对此知之甚少控制这些结构排列的因素，或不同组件如何影响炎症。这该提案旨在解决 ASC 亚型对炎症小体结构的作用的知识差距组织和炎症反应。该提案的长期目标是深入了解炎症小体成分之间的相互作用，对于了解其功能和调节具有重要意义为开发控制炎症的疗法奠定基础具有重要意义。此举的目的该资助具有创新性，因为它将破译炎症小体调节介导的未知分子基础由 ASC 亚型。我们的假设是； 1）接头长度对相互作用有重要影响 ASC 和 ASC_short 在分子水平上的特性，这可以解释观察到的炎症反应，2) 连接子长度导致两种亚型的域间动力学差异以及由此产生的宏观结构。为了检验这个假设，我们建议： 1）确定炎症 ASC 同种型的活性、精确自关联和相互作用能力，包括 ASC、ASC_short 和使用长链接器（规范链接器长度的 3 倍：ASC_long）作为参考设计的 ASC 的人工形式对于独立域，2) 使用核磁确定三种亚型的域间动力学谐振; 3）用透射电子显微镜辨别潜在的不同特征 ASC 亚型自关联产生的宏观结构及其在炎症活动中的意义。