Regulation of Vascular Inflammatory Signaling by the Deubiquitinase USP20

去泛素酶 USP20 对血管炎症信号的调节

• 批准号: 9893026
• 负责人: NEIL J. FREEDMAN
• 金额: $ 53.46万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-03-15 至 2023-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9893026
• 关键词: Adaptor Signaling Protein; Adenoviruses; Amino Acids; Anti-Inflammatory Agents; Antiatherogenic; Arrestins; Arterial Injury; Atherosclerosis; Attenuated; Binding; Biological Assay; Blood Vessels; CCL2 gene; CRISPR/Cas technology; Cause of Death; Cell Adhesion Molecules; Chronic; Cytokine Receptors; Deubiquitination; Dominant-Negative Mutation; Endothelial Cells; Endothelium; Family; Goals; Human; Hyperplasia; I Kappa B-Alpha; IRAK1 gene; In Vitro; Industrialization; Inflammation; Inflammatory; Interleukin 4 Receptor; Interleukin-1 Receptors; LOX gene; Link; Lipopolysaccharides; Mediating; Molecular; Mus; Myocardial Infarction; Partner in relationship; Phosphorylation; Phosphotransferases; Polyubiquitination; Post-Translational Protein Processing; Protein Kinase; Proteins; RIPK1 gene; ROCK1 gene; Regulation; Role; Signal Pathway; Signal Transduction; Signaling Protein; Smooth Muscle Myocytes; Stroke; TLR2 gene; TLR4 gene; TNF gene; TNFRSF1A gene; TRAF2 gene; TRAF6 gene; Tamoxifen; Testing; Transgenic Mice; Transgenic Organisms; Tumor Necrosis Factor Receptor; Ubiquitin; Ubiquitination; atherogenesis; cellular transduction; congenic; cytokine; desensitization; in vivo; interleukin-1 receptor-associated kinase; loss of function; lysophosphatidic acid; macrophage; mutant; new therapeutic target; novel; protein B; response; scaffold; transcription factor; ubiquitin-protein ligase; ubiquitin-specific protease; vascular inflammation; western diet

PROJECT SUMMARY We recently demonstrated that ubiquitin-specific protease-20 (USP20) is scaffolded by the adaptor protein known as β-arrestin2 (βarr2), and that USP20 desensitizes ubiquitin-dependent signaling from Toll-like receptor-4 (TLR4) to NFκB activation by deubiquitinating TRAF6 and βarr2. Using transgenic mice expressing dominant-negative USP20 in smooth muscle cells, we found that USP20 reduces neointimal hyperplasia after arterial injury and that USP20 activity in SMCs reduces atherosclerosis in Ldlr-/- mice. To establish anti-atherogenic effects of systemically expressed USP20, and to elucidate further molecular mechanisms by which USP20 protects against atherosclerosis, this project will test the hypothesis that USP20 attenuates atherosclerosis by deubiquitinating several substrate proteins that were previously unassociated with USP20 but that are important in signaling pathways that activate NFκB: βarr1, TRAF6, TRAF2, and RIPK1. Furthermore, because USP20 employs βarr2 as a scaffold to facilitate association with distinct proteins, and because βarr1 reduces vascular inflammation, this project will test whether USP20’s anti-atherogenic activity involves βarr1-mediated scaffolding. To these ends, this project will study systemic effects of USP20 on atherosclerosis by comparing Usp20-/- /Ldlr-/- versus Ldlr-/- mice, on a background of βarr1+/+ or βarr1-/+. To determine the effects of endothelial USP20 on atherosclerosis, this project will compare atherosclerosis among VECad-Cre- ERT2/Usp20flox/flox/Ldlr-/- vs. Usp20flox/flox/Ldlr-/- mice treated ± tamoxifen; furthermore, we will investigate cytokine secretion, and dynamic ubiquitination of signaling proteins in primary aortic endothelial cells that are WT, Usp20-/-, Usp20-/-/βarr1-/+, or βarr1-/-. To determine what kinase in endothelial cells phosphorylates USP20 on Ser333 (and thereby abrogates USP20 deubiquitinase activity), this project will test IRAK1, PAK1, and ROCK1 with several loss-of function approaches, including a USP20 minigene, in primary endothelial cells. These studies collectively may identify USP20 phosphorylation as novel therapeutic target for atherosclerosis.

项目总结 我们最近证明泛素特异性蛋白水解酶-20(USP20)是以接头为骨架的 被称为β-arrestin2(βarr2)的蛋白质，并且USP20使泛素依赖的信号从 Toll样受体4通过去泛素化TRAF6和κArr2激活核因子βB。使用转基因技术 在血管细胞中表达显性阴性USP20的小鼠，我们发现USP20减少了 动脉损伤后新生内膜增生及SMC中USP20活性降低动脉粥样硬化 LDLR-/-小鼠。建立系统表达的USP20的抗动脉粥样硬化作用，并阐明 该项目将测试USP20预防动脉粥样硬化的进一步分子机制 USP20通过去泛素化几种底物蛋白减轻动脉粥样硬化的假说 它们以前与USP20无关，但在激活的信号通路中很重要 核因子κB：βARR1、TRAF6、TRAF2和RIPK1。此外，由于USP20使用βARR2作为脚手架 为了促进与不同蛋白质的联系，并且因为βarr1减少血管炎症，这 该项目将测试USP20的S抗动脉粥样硬化活性是否涉及βArr1介导的支架。至 最后，本项目将通过比较USP20-/-来研究USP20在动脉粥样硬化中的系统作用 /Ldlr-/-对Ldlr-/-小鼠，背景为βarr1+/+或βarr1-/+。确定血管内皮细胞的作用 USP20关于动脉粥样硬化，本项目将比较VECad-CRE- ERT2/Usp20Flox/Flox/Ldlr-/-vs.Usp20Flox/Flox/Ldlr-/-治疗的小鼠±他莫昔芬；此外，我们将研究 原代主动脉内皮细胞细胞因子分泌与信号蛋白的动态泛素化 它们是WT、USP20-/-、USP20-/-/βARR1-/+或βARR1-/-。要确定内皮细胞中的哪种激酶 使Ser333上的USP20磷酸化(从而取消USP20去泛素酶活性)，该项目 将使用几种功能丧失方法测试IRAK1、PAK1和ROCK1，包括USP20 微基因，在原代内皮细胞中。这些研究可能共同确定USP20的磷酸化 作为动脉粥样硬化的新治疗靶点。