The role of MCPIP1 in regulating NF-kB signaling and macrophage activation

MCPIP1 在调节 NF-kB 信号传导和巨噬细胞激活中的作用

• 批准号: 8047250
• 负责人: MINGUI FU
• 金额: $ 18.75万
• 依托单位: UNIVERSITY OF MISSOURI KANSAS CITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-01-01 至 2012-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8047250
• 关键词: Anemia; Arterial Fatty Streak; Atherosclerosis; Attention; Autoimmune Responses; Biochemical Genetics; Cell Survival; Cells; Complex; Cytokine Degradation; Data; Deubiquitinating Enzyme; Deubiquitination; Development; Disease; Family; Feedback; Gene Expression; Goals; Growth Factor; Human; Immunity; In Vitro; Inflammation; Inflammatory; Inflammatory Response; Interleukin-1; Knockout Mice; Ligands; MAPK8 gene; Macrophage Activation; Maps; Mediating; Messenger RNA; Molecular; Molecular Target; Monocyte Chemoattractant Protein-1; Mus; Mutagenesis; NF-kappa B; Pathogenesis; Peptide Hydrolases; Pharmacotherapy; Phenotype; Play; Polyubiquitination; Process; Proteins; Pulmonary Emphysema; Pulmonary Fibrosis; Regulation; Relative (related person); Reporting; Research; Rheumatoid Arthritis; Ribonucleases; Role; Septic Shock; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Stimulus; Syndrome; TNF gene; TNFRSF5 gene; TRAF2 gene; TRAF6 gene; Testing; Tissues; Toll-like receptors; Tumor Necrosis Factor Receptor; Ubiquitin; Ubiquitination; Up-Regulation; Work; Zinc Fingers; base; chronic pancreatitis; cytokine; human disease; in vitro Assay; in vivo; injured; macrophage; novel; oxidized low density lipoprotein; programs; repaired

DESCRIPTION (provided by applicant): Activated macrophages play an important role in many inflammatory diseases including septic shock and atherosclerosis. However, the molecular mechanisms limiting macrophage inflammation are not completely understood. MCP-induced protein 1 (MCPIP1) is a recently identified CCCH-zinc finger containing protein, which was significantly induced by monocyte chemotactic protein 1 (MCP-1) and thus designated as MCPIP1. In our previous works, we have identified MCPIP1 as a novel negative regulator of macrophage inflammatory activation. In our recent studies, we have further found that MCPIP1 acts as a deubiquitinating enzyme that may negatively regulates NF-:B signaling by removing ubiquitin moieties from critical proteins, such as TNF receptor adaptor factors (TRAFs), RIP and I:B1. Consistently, MCPIP1- deficient mice spontaneously developed inflammatory syndrome and died prematurely. Macrophages from MCPIP1-/- mice showed high up-regulation of inflammatory gene expression, together with a greatly increased NF-kB activation, as well as increased polyubiquitination of TRAF2 and TRAF6. Furthermore, in vitro assay directly demonstrated the deubiquitinating activity of purified MCPIP1. Based on these intriguing findings, we hypothesize that MCPIP1 represses NF-kB signaling and microphage activation mainly through deubiquitination of TRAF family. The overall objective of this proposal is to test the central hypothesis by using combined biochemical and genetic approaches. Specially, we will establish MCPIP1- mediated deubiquitination of TRAFs as an essential mechanism in the regulation of NF-:B signaling as well as macrophage activation both in vitro and in vivo. We will attain the objective of this aim by using following approaches: 1) determine the direct molecular targets of MCPIP1 deubiquitinase in vitro; 2) determine the in vivo targets of MCPIP1 deubiquitinase using the primary cells and tissues from MCPIP1 knockout mouse; 3) map the active domain of MCPIP1 deubiquitinase through serial mutagenesis analysis; 4) define the functional relationship between deubiquitinase domain and RNase domain; 5) determine the relative contribution of the deubiquitinase activity of MCPIP1 to the suppression of NF-:B signaling as well as macrophage activation; 6) determine the role of MCPIP1 deubiquitinase in human macrophages. Completion of the proposed studies will not only help to understand the molecular basis of macrophage activation, but also implicate in the development of novel drug therapies against inflammatory diseases such as atherosclerosis. PUBLIC HEALTH RELEVANCE: The goal of this proposal is to explore the role and molecular mechanisms of a newly identified zinc finger protein MCPIP1 in regulating NF-KB signaling and macrophage activation. Completion of this research program will not only help us understand the function and molecular mechanisms of MCPIP1 in macrophage activation, but also provide critical information for potential new therapies for inflammatory diseases including atherosclerosis.

描述（申请人提供）：活化的巨噬细胞在感染性休克和动脉粥样硬化等许多炎症性疾病中发挥重要作用。然而，限制巨噬细胞炎症的分子机制尚不完全清楚。MCP-induced protein 1 （MCPIP1）是最近发现的一种含有ccch -锌指的蛋白，它被单核细胞趋化蛋白1 （MCP-1）显著诱导，因此被命名为MCPIP1。在我们之前的工作中，我们已经确定MCPIP1是巨噬细胞炎症激活的一种新的负调节因子。在我们最近的研究中，我们进一步发现MCPIP1作为一种去泛素化酶，可能通过去除关键蛋白（如TNF受体适配因子（TRAFs）、RIP和I:B1）上的泛素片段负调控NF-:B信号传导。同样，MCPIP1缺陷小鼠自发地出现炎症综合征并过早死亡。MCPIP1-/-小鼠巨噬细胞炎症基因表达高上调，NF-kB活化显著增加，TRAF2和TRAF6多泛素化增加。此外，体外实验直接证明纯化的MCPIP1具有去泛素化活性。基于这些有趣的发现，我们假设MCPIP1主要通过TRAF家族的去泛素化来抑制NF-kB信号传导和微噬细胞激活。本提案的总体目标是通过结合生物化学和遗传方法来检验中心假设。特别是，我们将在体外和体内建立MCPIP1介导的TRAFs去泛素化是调节NF-:B信号传导和巨噬细胞活化的重要机制。我们将通过以下方法实现这一目标：1)确定MCPIP1脱泛素酶的直接分子靶点；2)利用MCPIP1基因敲除小鼠的原代细胞和组织确定MCPIP1去泛素酶的体内靶点；3)通过序列诱变分析绘制MCPIP1去泛素酶活性区域；4)明确去泛素酶结构域与RNase结构域的功能关系；5)确定MCPIP1去泛素酶活性对NF-:B信号传导抑制和巨噬细胞活化的相对贡献；6)确定MCPIP1去泛素酶在人巨噬细胞中的作用。这些研究的完成不仅有助于了解巨噬细胞活化的分子基础，而且对开发针对动脉粥样硬化等炎症性疾病的新型药物治疗具有重要意义。