Deciphering the tau phosphorylation code

破译 tau 磷酸化密码

• 批准号: 10115965
• 负责人: Daniel T Pak
• 金额: $ 23.09万
• 依托单位: GEORGETOWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-01-15 至 2022-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10115965
• 关键词: Age; Alzheimer' s Disease; Alzheimer' s disease model; Amyloid beta-Protein; Atlases; Chemicals; Chronic; Code; Cognitive deficits; Complication; Databases; Dementia; Development; Disease; Disease Progression; Disease model; Etiology; Event; Female; Filament; Frontotemporal Dementia; High Pressure Liquid Chromatography; Hippocampus (Brain); Knock-in; Knock-in Mouse; Lead; MAPT gene; Mass Spectrum Analysis; Memory impairment; Methods; Microtubules; Modeling; Modification; Mus; Mutation; Nerve Degeneration; Neurodegenerative Disorders; Neurofibrillary Tangles; Neuronal Injury; Neurons; Pathogenesis; Pathogenicity; Pathologic; Pathology; Pathway interactions; Pattern; Phosphorylation; Phosphorylation Site; Phosphotransferases; Physiological; Physiology; Play; Protocols documentation; Regulation; Resources; Role; Senile Plaques; Severity of illness; Sex Differences; Signal Pathway; Site; Synaptic plasticity; Tauopathies; Time; Work; age related; base; density; experimental study; familial Alzheimer disease; hyperphosphorylated tau; improved; in vivo; interest; kinase inhibitor; male; mouse model; nano; neurotoxicity; next generation; novel; novel therapeutic intervention; paired helical filament; prion-like; response; slow potential; tandem mass spectrometry; targeted treatment; tau Proteins; tau aggregation; tau function; tau mutation; tau phosphorylation; tau-1; tau-protein kinase; therapeutic target

Alzheimer’s disease (AD) is a neurodegenerative disorder characterized by plaques comprised of Aβ, and neurofibrillary tangles (NFTs) containing the microtubule associated protein tau. Tau pathology closely correlates with neuronal degeneration and cognitive deficits. As the loss of tau protects against Aβ-induced neurotoxicity, tau is thought to act as a pathogenic downstream effector of Aβ to induce neuronal injury. Furthermore, aggregated tau can propagate in a prion-like manner to initiate a self-perpetuating toxic cascade. For these reasons, tau-directed approaches and novel mechanisms of treatment are essential. Hyperphosphorylation of tau is a consistent feature of all tauopathies, suggesting it may be obligatory step in pathogenesis and hence a rational target of modulation. Phosphorylation dissociates tau from microtubules and promotes tau aggregation into paired helical filaments that further accumulate to form NFTs. Tau is natively unfolded, and under normal conditions has little tendency to aggregate; therefore hyperphosphorylated tau in NFTs is a prominent sign of neurodegeneration, and understanding the etiology and pattern of phosphorylated tau is essential. A complication in this analysis is that there are over 80 potential tau phosphorylation sites. Due to the complexity of this problem, tau “hyperphosphorylation” is not precisely defined. Additionally, tau plays important roles during both associative and homeostatic forms of synaptic plasticity, but little is known regarding the phosphorylation events involved in these pathways. Homeostatic responses to hyperexcitation are of particular interest as this type of aberrant overactivity is observed in early stage AD and may be relevant for the initiation of pathogenesis. Here, we will use unbiased mass spectrometry (MS) to perform comprehensive mapping of tau phosphorylation patterns during specific physiological conditions as well as in disease models. Compiling phosphomaps into an “atlas” of tau modifications will begin to decipher the phosphorylation code that governs tau function in physiology and pathology. In Aim 1, we will use qualitative and quantitative MS approaches to define tau phosphorylation patterns during different forms of synaptic plasticity in cultured hippocampal neurons, examining a time course after each paradigm to observe time-dependent changes. We will also use specific kinase inhibitors with the above stimulation protocols to facilitate identification of endogenous tau kinases and signaling pathways. In Aim 2, we will use MS to identify tau phosphorylation patterns in a humanized double knock-in model of familial AD. As a primary tauopathy model of frontotemporal dementia, we will use P301L-tau in the background of humanized tau knock-in mice. We will examine different ages to understand the earliest tau modifications and profile of disease progression, as well as male vs. female mice to elucidate sex differences in disease severity. Defining the physiological regulation and phosphorylation code of tau is highly significant for understanding basic mechanisms involved in synaptic plasticity. Furthermore, this work will help guide new therapeutic approaches for targeting pathogenic tau in AD and other tauopathies.

阿尔茨海默氏病（AD）是一种神经退行性疾病，其特征在于由Aβ组成的斑块， 神经元缠结（NFT），其含有微管相关蛋白tau。Tau病理学与 神经元退化和认知缺陷由于tau蛋白的缺失可以防止Aβ诱导的神经毒性， tau被认为是Aβ的致病性下游效应物以诱导神经元损伤。此外，委员会认为， 聚集的tau可以朊病毒样方式传播以启动自我延续的毒性级联。为这些 因此，针对tau的方法和新的治疗机制至关重要。过度磷酸化 tau蛋白是所有tau蛋白病的一致特征，这表明它可能是发病机制中的必要步骤， 合理的调制目标磷酸化使tau蛋白从微管上解离并促进tau蛋白聚集 形成成对的螺旋状细丝，其进一步积累形成NFT。Tau是天然展开的，在正常情况下， 条件下几乎没有聚集的倾向;因此，NFT中的过度磷酸化tau是NFT的显著标志。 因此，了解磷酸化tau蛋白的病因和模式是至关重要的。该分析中的复杂性在于存在超过80个潜在的tau磷酸化位点。由于这个问题的复杂性，tau蛋白的“过度磷酸化”并没有被精确定义。此外，tau蛋白在突触可塑性的联合和稳态形式中都起着重要作用，但关于其磷酸化的研究却知之甚少。 参与这些途径的事件。对过度兴奋的稳态反应特别令人感兴趣，因为这 在早期AD中观察到一种异常的过度活动，并且可能与发病机制的启动有关。 在这里，我们将使用无偏质谱法（MS）对tau蛋白磷酸化进行全面的定位 在特定的生理条件下以及在疾病模型中的模式。将磷酸图谱编译成 tau修饰的“图谱”将开始破译在生理学中支配tau功能的磷酸化密码 和病理在目标1中，我们将使用定性和定量MS方法来定义tau蛋白磷酸化 在培养的海马神经元中不同形式的突触可塑性的模式，检查时间过程 每个范例后观察随时间变化。我们还将使用特定的激酶抑制剂， 上述刺激方案以促进内源性tau激酶和信号传导途径的鉴定。在Aim中 2，我们将使用MS来鉴定家族性AD的人源化双敲入模型中的tau磷酸化模式。 作为额颞叶痴呆的原发性tau蛋白病模型，我们将在以下背景下使用P301 L-tau蛋白： 人源化tau基因敲入小鼠。我们将研究不同的年龄，以了解最早的tau修饰， 疾病进展概况以及雄性与雌性小鼠，以阐明疾病严重程度的性别差异。 明确tau蛋白的生理调节和磷酸化密码对于理解tau蛋白的基础功能具有重要意义。 参与突触可塑性的机制。此外，这项工作将有助于指导新的治疗方法 用于靶向AD和其他tau蛋白病中的致病性tau蛋白。