Chemical Biology of the E3 Ubiqutin Ligase Nedd4

E3 泛素连接酶 Nedd4 的化学生物学

• 批准号: 10568123
• 负责人: Dewey G McCafferty
• 金额: $ 31.87万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-01 至 2027-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10568123
• 关键词: Address; Binding; Binding Sites; Biology; Brain; Cell model; Cell physiology; Chemicals; Communication; Development; Disease; Enzymes; Family; Future; Glues; Golgi Apparatus; Knowledge; Label; Lewy Body Dementia; Ligase; Location; Malignant Neoplasms; Methods; Modification; Molecular; Multienzyme Complexes; Neurodegenerative Disorders; Neuroglia; Neurons; Parkinson Disease; Pathway interactions; Phenotype; Play; Post-Translational Protein Processing; Process; Proteins; Role; Signal Transduction; Structure; Substrate Specificity; Testing; Toxic effect; Ubiquitin; Ubiquitination; alpha synuclein; enzyme activity; gain of function; insight; member; novel therapeutics; precursor cell; proteostasis; proteotoxicity; response; screening; small molecule; synucleinopathy; therapeutic development; trafficking; ubiquitin-protein ligase

Ubiquitylation is a covalent posttranslational modification that influences a tremendous number of normal and disease-related cellular functions. The neuronal precursor cell-expressed developmentally downregulated 4 enzyme (Nedd4) is the founding member of a family of HECT-type E3 ubiquitin ligases that regulate proteostasis in various conditions including cancers and neurodegenerative disorders. In addition to its importance in cancer, Nedd4 has also recently emerged as a significant regulator of a-synuclein proteotoxicity, and may be a target for treating synucleinopathies, neurodegenerative disorders characterized by the accumulation of aggregated forms of the protein a-synuclein in both neuronal and non-neuronal cells in the brain. NAB2 is an N- arylbenzimidazole small molecule discovered from phenotypic screening that reverses a-synuclein associated toxicity in cellular models of a-synuclein-associated PD, including ER-to-Golgi trafficking disruption. Recently we elucidated the mode of action of NAB2, finding that it bound tightly to the E3 ubiquitin ligase Nedd4 (Kdapp = 42 nM) and expands the substrate specificity of Nedd4. We determined that NAB2 binding to Nedd4 triggers the ubiquitylation of new substrates, such as the trafficking-associated proteins such as TFG, TRAPP1 subunits, and RER1. We hypothesize that Nedd4 signaling influences a new pathway that regulates trafficking processes in response to a-synuclein-associated toxicity. The proteoform state of Nedd4 that responds to a-synuclein toxicity and binds NAB2 is also not known. In addition, the location of the NAB2 binding site within Nedd4 also is not known. In this proposal we wish to test the hypothesis that NAB2 acts on Nedd4 alone or as ‘molecular glue’ to facilitate communication between Nedd4 and potential substrates. We wish to understand how NAB2 interacts with NEDD4, and how this interaction affords Nedd4 a gain of function activity to ubiquitin modifiy non PxY-containing substrates like TFG. If successful, these studies will better illuminate the role Nedd4 plays in proteotoxicity signaling. In addition, the degree to which Nedd4 activity can be modulated by small molecules is important, given the important role Nedd4 ubiquitination plays in cancer pathobiology and neurodegenerative disorders. These studies will hopefully add to our fundamental knowledge of the role that E3 ligases play in proteotoxicity signaling, and may inspire the future discovery and development of therapeutics that may address some of the causes of synucleinopathies.

泛素化是一种共价翻译后修饰，影响大量正常细胞 和疾病相关的细胞功能。神经元前体细胞表达的发育下调4。 酶 (Nedd4) 是调节蛋白质稳态的 HECT 型 E3 泛素连接酶家族的创始成员 在包括癌症和神经退行性疾病在内的各种疾病中除了它在癌症中的重要性之外， Nedd4 最近也成为 a-突触核蛋白蛋白毒性的重要调节因子，并且可能是一个靶点 用于治疗突触核蛋白病、以聚集体积累为特征的神经退行性疾病 NAB2 蛋白在大脑神经元和非神经元细胞中的形式都是 N-。 通过表型筛选发现的芳基苯并咪唑小分子可逆转α-突触核蛋白相关 α-突触核蛋白相关 PD 细胞模型的毒性，包括 ER 至高尔基体运输中断。 阐明了 NAB2 的作用模式，发现它与 E3 泛素连接酶 Nedd4 紧密结合（Kdapp = 42 nM）并扩展了 Nedd4 的底物特异性，我们确定 NAB2 与 Nedd4 的结合会触发 新底物的泛素化，例如 TFG、TRAPP1 亚基等运输相关蛋白， 我们认为 Nedd4 信号传导影响调节贩运过程的新途径。 回应 α-突触核蛋白相关毒性。 Nedd4 的蛋白质状态对 α-突触核蛋白有反应。 毒性和结合 NAB2 也是未知的。此外，NAB2 结合位点在 Nedd4 内的位置也是未知的。 在此提议中，我们希望检验 NAB2 单独作用于 Nedd4 或作为“分子”作用的假设。 胶”以促进 Nedd4 和潜在底物之间的通信我们希望了解 NAB2 是如何进行的。 与 NEDD4 相互作用，以及这种相互作用如何使 Nedd4 获得泛素修饰非功能活性 含有 PxY 的底物如 TFG 如果成功，这些研究将更好地阐明 Nedd4 在其中发挥的作用。 此外，小分子可以调节 Nedd4 活性的程度。 鉴于 Nedd4 泛素化在癌症病理学和神经退行性疾病中发挥的重要作用，这一点很重要 这些研究有望增加我们对 E3 连接酶在疾病中的作用的基础知识。 蛋白质毒性信号传导，可能会激发未来治疗方法的发现和开发，从而解决 突触核蛋白病的一些原因。