Serine/Threonine Phosphatases in Neurological Diseases

神经系统疾病中的丝氨酸/苏氨酸磷酸酶

• 批准号: 10583671
• 负责人: Wolfgang Peti
• 金额: $ 53.63万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-01-01 至 2027-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10583671
• 关键词: A kinase anchoring protein; Active Sites; Affinity Chromatography; Alzheimer' s Disease; Binding; Biochemical; Biological; Biological Process; Brain; Calcineurin; Cell Line; Cells; Cellular biology; Collaborations; Complex; Coupled; Cyclic AMP-Dependent Protein Kinases; Cyclosporine; Data; Development; Disease; Environment; FK506; Foundations; Funding; Immunosuppressive Agents; Knowledge; Lead; Long-Term Depression; Long-Term Potentiation; MAP3K7 gene; MAP3K7IP1 gene; Mass Spectrum Analysis; Mediating; Memory; Molecular; Molecular Target; Nervous System Physiology; Neurons; PPP3CA gene; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphotransferases; Physiological; Physiology; Population; Post-Translational Protein Processing; Process; Protein Dephosphorylation; Protein phosphatase; Proteins; Proteome; Recording of previous events; Regulation; Research; Research Personnel; Resolution; Role; Scaffolding Protein; Serine; Shapes; Signal Pathway; Signal Transduction; Stimulus; Stroke; Structure; Substrate Interaction; System; Techniques; Threonine; Work; calcineurin phosphatase; cell type; experimental study; human disease; inhibitor; innovation; insight; nervous system disorder; neuron development; neurotransmission; novel; palmitoylation; receptor; recruit; response; success; therapeutic target; tool; voltage

ABSTRACT. Phosphorylation is one of the most ubiquitous, reversible posttranslational modifications in cells, and is a critical component of most signaling cascades. Strict temporal and spatial control are essential for the fidelity of this process, as derailed signaling cascades lead to disease. Here, we continue our long-standing effort to investigate signaling in neurons. If neuronal signaling goes awry, the most prominent results are well known diseases, such as Alzheimer's disease and stroke. Recently, we successfully determined how the most ubiquitous neuronal ser/thr protein phosphatase (PPP) calcineurin (CN) recruits its substrates. Namely, CN binds regulators and substrates via CN-specific recruitment motifs (PxIxIT and LxVP). Further, we also discovered that CN uses an active site recognition sequence (TxxP) to target substrate phosphosites, which, in turn, drives vital biological functions. This is the first defined active site recognition sequence for any PPP, transforming our ability to identify novel CN-specific phosphosites. Here, we will leverage our newly established tools and discoveries to achieve three aims. First, we will establish the CN interactome and substratome in distinct neuronal populations. This will enable us to demonstrate the diversity of CN functions in neurons, define if they differ in response to stimuli as well as identify the molecular substrates that are necessary for these changes to occur. Building further on the success of the previous funding period, we will also advance our molecular understanding of CN substrate recruitment by studying two critical CN substrate signaling platforms: CN-AKAP5 and CN-TAK1. Specifically, we will show that the these signaling platforms utilize multiple, competing PxIxIT/LxVP motifs to recruit CN via different proteins and show how these distinct mechanisms define CN substrate dephosphorylation efficacy. Critically, our preliminary data suggest that posttranslational modification of AKAP5 modulates CN control of PKA activity and ultimately receptor regulation. Finally, we have also recently confirmed our prediction that the transforming growth factor-β activated kinase 1 (TAK1) binds directly to CN. However, unexpectedly the TAK1 regulator TAB2 also binds directly to CN via different LxVP and PxIxIT motifs. We will investigate the mechanisms and consequences of this interaction on CN recruitment and TAK1 function. The modes of action and regulation of CN in the AKAP5 and TAK1 signaling platforms are unexpected and highlight the broad variety of mechanisms used to regulate CN activity. Taken together, the proposed studies leverage a powerful integrated approach that combines atomic resolution techniques with biochemical and cell biology experiments to obtain novel insights into the molecular mechanisms used to direct CN activity. Because CN has critical roles in human diseases generally, and in the brain specifically, and because CN is the only successfully therapeutically targeted PPP (CN is the target of the blockbuster immunosuppressants cyclosporin A and FK-506), our proposed work will provide a critically needed molecular and cellular understanding of CN activity and regulation in neuronal function.

抽象的。磷酸化是细胞中最普遍、可逆的翻译后修饰之一， 并且是大多数信号级联的关键组成部分。严格的时间和空间控制对于 这个过程的保真度，因为脱轨的信号级联会导致疾病。在这里，我们继续我们的长期坚持 努力研究神经元信号传导。如果神经元信号传导出错，最显着的结果是好的 已知的疾病，例如阿尔茨海默病和中风。最近，我们成功地确定了如何最 普遍存在的神经元丝氨酸/苏氨酸蛋白磷酸酶 (PPP) 钙调神经磷酸酶 (CN) 招募其底物。即，CN 通过 CN 特异性募集基序（PxIxIT 和 LxVP）结合调节剂和底物。此外，我们还 发现 CN 使用活性位点识别序列 (TxxP) 来靶向底物磷酸位点， 反过来，驱动重要的生物功能。这是第一个定义的 PPP 活性位点识别序列， 改变我们识别新型 CN 特异性磷酸位点的能力。在这里，我们将利用我们新成立的 实现三个目标的工具和发现。首先，我们将建立 CN 相互作用组和底物组 不同的神经元群体。这将使我们能够证明神经元中 CN 功能的多样性，定义 如果它们对刺激的反应不同，并确定这些反应所需的分子底物 发生的变化。在上一个资助期成功的基础上，我们还将进一步推进我们的 通过研究两个关键的 CN 底物信号传导平台对 CN 底物募集进行分子理解： CN-AKAP5 和 CN-TAK1。具体来说，我们将展示这些信号平台利用多个、 竞争性 PxIxIT/LxVP 基序通过不同的蛋白质招募 CN，并展示这些不同的机制如何 定义 CN 底物去磷酸化功效。至关重要的是，我们的初步数据表明翻译后 AKAP5 的修饰可调节 CN 对 PKA 活性的控制，并最终调节受体。最后，我们 最近还证实了我们的预测，即转化生长因子-β 激活激酶 1 (TAK1) 直接与 CN 结合。然而，出乎意料的是，TAK1 调节剂 TAB2 也通过不同的方式直接与 CN 结合。 LxVP 和 PxIxIT 基序。我们将研究这种相互作用对 CN 的机制和后果 募集和 TAK1 功能。 CN 在 AKAP5 和 TAK1 信号传导中的作用和调节模式 平台的出现是出乎意料的，并突显了用于调节 CN 活动的多种机制。采取 总之，拟议的研究利用了一种强大的综合方法，结合了原子分辨率 生物化学和细胞生物学实验技术，以获得对分子的新见解 用于指导 CN 活动的机制。因为 CN 在一般人类疾病中起着关键作用，并且在 特别是大脑，并且因为 CN 是唯一成功的治疗靶向 PPP（CN 是 重磅免疫抑制剂环孢菌素 A 和 FK-506），我们提出的工作将提供一个关键的 需要对 CN 活动和神经元功能调节进行分子和细胞了解。