Alpha4's regulation of PP2A activity: Role in tau hyperphosphorylation

Alpha4 对 PP2A 活性的调节：在 tau 过度磷酸化中的作用

• 批准号: 8203280
• 负责人: Michele Laura LeNoue-Newton
• 金额: $ 2.64万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-12-01 至 2014-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8203280
• 关键词: Address; Affect; Affinity; Age; Alzheimer' s Disease; American; Antibodies; Binding; Brain; Caspase; Catalytic Domain; Cell Line; Cells; Complement; Complex; Data; Elderly; Electrons; Elements; Engineering; Event; Exhibits; Face; Goals; Impaired cognition; Lead; Measures; Memory Loss; Microtubule-Associated Proteins; Microtubules; Modeling; Molecular; Molecular Structure; Mono-S; Nerve Degeneration; Neurodegenerative Disorders; Neurofibrillary Tangles; Neurons; Patients; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphotransferases; Play; Polyubiquitination; Post-Translational Protein Processing; Protein Dephosphorylation; Protein Phosphatase 2A Regulatory Subunit PR53; Protein phosphatase; Proteins; Regulation; Research; Role; Senile Plaques; Spectrum Analysis; Structure; Symptoms; Ubiquitin; Ubiquitination; Work; base; flexibility; genetic regulatory protein; hyperphosphorylated tau; insight; interest; multicatalytic endopeptidase complex; mutant; new therapeutic target; prevent; protein protein interaction; scaffold; tau Proteins; tau phosphorylation; ubiquitin-protein ligase

DESCRIPTION (provided by applicant): Alzheimer's disease is a progressive neurodegenerative disease causing cognitive decline and memory loss that effects 1 in 8 Americans over the age of 65 (1). It is estimated that by the year 2050 between 11 million and 16 million Americans age 65 and older will be affected (2). Currently, there are strategies to temporarily reduce the symptoms of Alzheimer's, but there are no therapies to stop the progression of neurodegeneration or cure AD (3). There are two main pathophysiological features of AD, beta amyloid plaques and neurofibrillary tangles (4). Neurofibrillary tangles are composed of hyperphosphorylated tau protein (5, 6). Normal tau is a microtubule-associated protein that stabilizes microtubules (7). Hyperphosphorylation of tau decreases its affinity for microtubules and hyperphosphorylated tau sequesters normal tau preventing it from binding to microtubules (8-10). Tau is phosphorylated by numerous kinases, but PP2A has been implicated as being the predominant phosphatase involved in tau dephosphorylation (11-13). Alpha4 has been shown to regulate PP2A stability and expression (14, 15) and localizes to microtubules due to its association with Mid1 (16). Specifically, alpha4 regulates ubiquitination of PP2Ac and its degradation by the proteasome by suppressing polyubiquitination of PP2Ac (15). Based on previous studies and preliminary structures, the working hypothesis is that the UIM of alpha4 binds to monoubiquitinated PP2Ac capping the ubiquitin chain and preventing polyubiquitination and that this protection is regulated through the ubiquitination of alpha4. In order to understand the mechanism by which alpha4 regulates PP2Ac, double electron-electron resonance (DEER) studies will be conducted to look at the role of flexibility of alpha4 in interacting with both PP2Ac and ubiquitin. To investigate the mechanism of alpha4 regulation of PP2A, a neuronal cell line will be transfected with alpha4 constructs that disrupt protein-protein interactions and PP2Ac ubiquitination and stability will be assessed. Activity of PP2A towards tau will be assessed using by measuring levels of phosho-tau using available phospho-tau antibodies. Our data indicate that alpha4 may be regulated via ubiqutination. To investigate this, 293FT cells will be transfected with alpha4 constructs that disrupt protein-protein interactions and alpha4 ubiquitination will be measured. The goal of this project is to elucidate the mechanism of alpha4 regulation of PP2A and investigate the role alpha4 plays in regulating PP2A activity towards tau. PUBLIC HEALTH RELEVANCE: Tau hyperphosphorylation, a hallmark of Alzheimer's disease (AD), is regulated by protein phosphatase 2A (PP2A). This research addresses the mechanism of alpha4's regulation of PP2A and attendant effects on tau phosphorylation. The results of this research will further our understanding of the molecular basis of tau hyperphosphorylation and lead to possible new therapeutic targets.

描述（由申请人提供）：阿尔茨海默病是一种进行性神经退行性疾病，导致认知能力下降和记忆丧失，影响八分之一的65岁以上美国人(1)。据估计，到2050年，将有1100万至1600万65岁及以上的美国人受到影响(2)。目前，有一些策略可以暂时减轻阿尔茨海默病的症状，但没有治疗方法可以阻止神经退行性疾病的进展或治愈AD(3)。AD有两个主要的病理生理特征，β -淀粉样斑块和神经原纤维缠结(4)。神经原纤维缠结由过度磷酸化的tau蛋白组成（5,6）。正常的tau蛋白是一种稳定微管的微管相关蛋白(7)。tau蛋白的过度磷酸化降低了其对微管的亲和力，过度磷酸化的tau蛋白会隔离正常的tau蛋白，阻止其与微管结合（8-10）。Tau被许多激酶磷酸化，但PP2A被认为是参与Tau去磷酸化的主要磷酸酶（11-13）。Alpha4已被证明调节PP2A的稳定性和表达（14,15），并且由于其与Mid1的关联而定位于微管（16）。具体来说，alpha4通过抑制PP2Ac的泛素化来调节PP2Ac的泛素化及其被蛋白酶体降解（15）。基于以往的研究和初步结构，我们的工作假设是，alpha4的UIM结合到单泛素化的PP2Ac上，覆盖泛素链，阻止多泛素化，这种保护是通过alpha4的泛素化来调节的。为了了解alpha4调控PP2Ac的机制，将进行双电子-电子共振（double electron-electron resonance， DEER）研究，以观察alpha4的柔韧性在与PP2Ac和泛素相互作用中的作用。为了研究alpha4调控PP2A的机制，我们将破坏蛋白-蛋白相互作用的alpha4构建体转染一个神经元细胞系，并评估PP2Ac泛素化和稳定性。PP2A对tau蛋白的活性将通过使用可用的磷酸化tau抗体测量磷酸化tau蛋白的水平来评估。我们的数据表明，alpha4可能通过泛素化调控。为了研究这一点，293FT细胞将被转染破坏蛋白-蛋白相互作用的alpha4构建体，并测量alpha4泛素化。本课题的目的是阐明alpha4调控PP2A的机制，探讨alpha4在调控PP2A对tau的活性中所起的作用。