Isoform-dependent effects of tau phosphorylation in Alzheimer's disease

阿尔茨海默病中 tau 磷酸化的异构体依赖性效应

• 批准号: 10536450
• 负责人: Nima Nick Naseri
• 金额: $ 6.76万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10536450
• 关键词: Isoform; dependent; effects; tau; phosphorylation

Neuropathology of the microtubule-associated protein tau is central to numerous devastating neurological disorders termed tauopathies, including Alzheimer's disease (AD). Tau promotes microtubule (MT) polymerization and stability via its interaction with tubulin, regulated by phosphorylation to tau in multiple domains. Six major tau isoforms are alternatively spliced in an age-dependent manner. While FTDP-17-causing mutations in tau show isoform-dependent effects on aggregation and microtubule stabilization, there are no tau mutations linked to AD. Meanwhile, isoform-dependent effects of phosphorylation to tau remain unexplored. Interestingly, I discovered that numerous AD-relevant phospho-tau (pTau) sites are also phosphorylated during normal embryonic development, and then decrease over age. Why are specific pTau sites, which are correlated with disease, also expressed during normal development? The parallel between the developmental and pathological states suggests that understanding the function of pTau during development would elucidate the mechanism(s) underlying tau pathology in AD. My preliminary data indicate that i) numerous AD-relevant pTau sites are expressed in normal embryonic mouse brains, ii) pTau is soluble and likely functional in fetal tissue, iii) phosphorylation at sites T231, S235 and S262 impair MT polymerization by the fetal tau isoform but not an adult isoform, and iv) these same pTau sites accelerate aggregation of the adult isoform over the fetal isoform. Recent evidence demonstrates that tau initiates nucleation of MT polymerization. Based on my preliminary data, ! propose that tau phosphorylation has isoform-dependent effects, in which specific pTau sites are detrimental to adult isoforms in an aging brain but functional in the fetal isoform in developing brains. I will explore two aims which will provide critical insight to understanding the effects of tau phosphorylation in each isoform. In Aim 1, I will determine the relationship between the interaction between pTau with both soluble tubulin and stabilized MTs in each isoform. In Aim 2, I will test for the relationship between pTau sites which cause tau to dissociate from MTs with those that accelerate its aggregation and/or seeding activity. Importantly, our lab has uniquely synthesized full-length tau with genuine chemical phosphorylation at sites T231, S235 and S262 for these studies. These experiments will be tested using multiple platforms including these chemically-synthesized proteins, high-resolution single-molecule techniques (single-molecule FRET, fluorescence correlative spectroscopy, and time-lapse TIRF microscopy), and injection of semi-synthesized proteins into mouse brains. The investigator has 6+ years of expertise in working with the biological assays and mouse models described here, and the sponsor and collaborators will guide training in the described chemical and biophysical techniques. I anticipate that my findings will elucidate the function of tau in developing neurons, and will provide insight into mechanisms of pathological tau hyperphosphorylation across numerous tauopathies including AD.

微管相关蛋白tau的神经病理学是许多破坏性神经系统疾病的核心。 称为tau蛋白病的疾病，包括阿尔茨海默病（AD）。Tau促进微管（MT） 通过其与微管蛋白的相互作用的聚合和稳定性，在多个细胞中通过磷酸化tau调节。 域.六种主要的tau亚型以年龄依赖性方式选择性剪接。虽然FTDP-17导致 tau蛋白的突变显示出对聚集和微管稳定的亚型依赖性作用， 与AD相关的突变与此同时，tau蛋白磷酸化的亚型依赖性作用仍未被探索。 有趣的是，我发现许多AD相关的磷酸化tau（pTau）位点在治疗过程中也被磷酸化。 正常的胚胎发育，然后随着年龄的增长而减少。为什么特定的pTau位点， 在正常发育过程中也有表达吗？发展与 病理状态表明，了解pTau在发育过程中的功能将阐明 AD中tau病理学的潜在机制。我的初步数据表明i）许多AD相关的pTau ii）pTau是可溶性的并且可能在胎儿组织中起作用，iii） T231、S235和S262位点的磷酸化通过胎儿tau亚型而不是成人损害MT聚合 iv）这些相同的pTau位点相对于胎儿同种型加速了成体同种型的聚集。最近 证据表明tau引发MT聚合的成核。根据我的初步数据，！ 提出tau磷酸化具有亚型依赖性效应，其中特定的pTau位点对 在老化的大脑中，成人同种型具有功能，但在发育中的大脑中，胎儿同种型具有功能。我将探讨两个目标 这将为理解每种同种型中tau磷酸化的作用提供关键的见解。在目标1中， 将确定pTau与可溶性微管蛋白和稳定的微管蛋白之间的相互作用之间的关系。 每种亚型的MT。在目标2中，我将测试导致tau解离的pTau位点之间的关系 从MT与那些加速其聚集和/或播种活动。重要的是，我们的实验室拥有独特的 合成的全长tau蛋白在位点T231、S235和S262处具有真正的化学磷酸化， 问题研究这些实验将使用多个平台进行测试，包括这些化学合成的 蛋白质，高分辨率单分子技术（单分子FRET，荧光相关 光谱和延时TIRF显微镜），以及将半合成蛋白质注射到小鼠脑中。 研究者在使用所述生物测定和小鼠模型方面拥有6年以上的专业知识 在这里，赞助者和合作者将指导所述化学和生物物理技术的培训。 我预计我的发现将阐明tau蛋白在发育神经元中的功能，并将为以下方面提供见解： 包括AD在内的许多tau蛋白病的病理性tau蛋白过度磷酸化机制。