Di-ubiquitin modification of ubiquitin ligase adaptors in membrane protein downregulation

泛素连接酶接头的双泛素修饰在膜蛋白下调中的作用

• 批准号: 10521677
• 负责人: SCOTT D EMR
• 金额: $ 41.07万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-01 至 2026-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10521677
• 关键词: Adaptor Signaling Protein; Amino Acids; Architecture; Arrestins; Autophagocytosis; Binding; Binding Sites; Biological Assay; C-terminal; C2 Domain; Cell Cycle; Cell membrane; Cell physiology; Cells; Collaborations; Complex; Cryoelectron Microscopy; Crystallography; DNA Repair; Data; Defect; Distal; Down-Regulation; Endocytosis; Enzymes; Eukaryota; Evolution; Family; Family member; Fostering; Genetic Transcription; Goals; Immune response; In Vitro; Ligase; Light; Link; Lysine; Maintenance; Malignant Neoplasms; Mammals; Mediating; Membrane; Membrane Proteins; Methionine; Modeling; Modification; Molecular; N-terminal; Nature; Neurodegenerative Disorders; PHEMX gene; Physiological; Play; Post-Translational Protein Processing; Process; Proteins; Quality Control; Research; Role; Signal Transduction; Site; Specificity; System; Testing; Ubiquitin; Ubiquitin family; Ubiquitin-Activating Enzymes; Ubiquitination; Yeasts; base; human disease; in vivo; interest; member; protein degradation; protein function; proteostasis; recruit; therapeutic development; therapeutic target; trafficking; ubiquitin ligase; ubiquitin-protein ligase

PROJECT SUMMARY/ABSTRACT Post-translational modification by ubiquitin is an essential mechanism to alter protein function in eukaryotes. Ubiquitin, a 76 amino acid protein, is attached to specific proteins via a cascade of ubiquitin activating enzyme E1, conjugating enzyme E2, and ubiquitin ligase E3. Ubiquitination plays an essential role in a broad aspect of cellular processes, including transcription, DNA repair, signal transduction, autophagy, cell cycle, immune response, and membrane trafficking. Aberration in the ubiquitination system leads to a number of human diseases, such as neurodegenerative diseases and cancers. Within the ubiquitination cascade, E3s primarily dictate the specificity of the ubiquitination system and thus are often the focal points of research and attractive therapeutic targets. The Nedd4 family E3s are an essential family of HECT-type E3s, members of which contain an N-terminal C2 domain followed by 2-4 WW domains and the C-terminal HECT domain. Nedd4 E3s recognize substrates carrying a “PPxY” motif through their WW domains. However, most substrates lack such a motif but engage with the ligase through “PPxY” motif-containing adaptors. We recently discovered that in yeast, the ubiquitin E3 ligase adaptor protein Art1 is primed with di-ubiquitination and the attachment of the di-Ub chain to a specific lysine residue in Art1 is warranted for its full activity. In this proposal, we plan to investigate the physiological function of adaptor di-ubiquitination and to elucidate the molecular mechanisms of the modular ubiquitination platform form with Nedd4 E3 ligase and di-ubiquitinated adaptors. Specifically, we will pursue the following aims: Aim 1: To investigate the mechanism of Nedd4 E3 adaptor di-ubiquitination. Aim 2: To determine the role of di-ubiquitinated adaptor-E3 complexes in substrate ubiquitination. Aim 3: To elucidate the molecular architecture of di-ubiquitinated adaptors with the Nedd4 E3 ligases. Uncovering the physiological role of Need4 E3 adaptors di-ubiquitination will be of critical importance to understand the molecular basis of how E3 adaptors specifically recognize substrate proteins and efficiently present the substrates for ubiquitination by HECT E3 ligases. We expect the successful implementation of this proposal will not only make significant contributions to the understanding of the molecular mechanisms underlying the Nedd4 E3 ligase/adaptor mediated ubiquitination, but also shed light on the mechanism of protein quality control governed by targeted ubiquitination.

项目摘要/摘要 泛素翻译后修饰是改变蛋白质功能的基本机制 真核生物。泛素是一种76个氨基酸蛋白 激活酶E1，共轭酶E2和泛素连接酶E3。泛素化起着必不可少的 在细胞过程的广泛方面的作用，包括转录，DNA修复，信号转导， 自噬，细胞周期，免疫反应和膜运输。泛滥 系统导致许多人类疾病，例如神经退行性疾病和癌症。之内 泛素化级联E3S主要决定了泛素化系统的特异性，因此是 通常是研究的焦点和有吸引力的治疗靶标。 NEDD4家族E3是必不可少的 Hect型E3的家族，其成员包含N末端C2域，然后是2-4 ww 域和C末端HECT域。 NEDD4 E3S识别携带“ PPXY”主题的底物 通过他们的WW域。但是，大多数底​​物缺乏这样的主题，但通过连接酶通过 “ PPXY”含有主题的适配器。我们最近发现，在酵母中，泛素E3连接酶适配器 蛋白质Art1涂有二氨液和Di-Ub链的附着在特定歌词上 Art1中的居住有必要的全部活动。在此提案中，我们计划研究生理 适配器二泛素化的功能并阐明模块化的分子机制 NEDD4 E3连接酶和Di泛素化适配器的泛素化平台形式。具体来说，我们会的 追求以下目的：目标1：研究NEDD4 E3适配器Di-upiquitation的机制。 目的2：确定二磷化适应器-E3复合物在底物泛素化中的作用。目标3： 为了阐明具有NEDD4 E3连接酶的Di泛素化适配器的分子结构。发现 需要4 e3适配器的物理作用di泛素化对于理解至关重要 E3适配器如何特别识别底物蛋白并有效地呈现的分子基础 Hect E3连接酶泛素化的底物。我们希望成功实施 提案不仅会为理解分子机制做出重大贡献 NEDD4 E3连接酶/衔接子介导的泛素化的基础，但也阐明了 蛋白质质量控​​制受靶向泛素化的控制。