Role of PP2A in Tauopathies

描述（由申请人提供）：阿尔茨海默病（AD）是痴呆症的主要原因，每年影响超过450万美国人，总成本至少为1000亿美元。蛋白、tau蛋白和淀粉样前体蛋白（APR）加工过程的改变是AD发病的关键事件。临床研究表明，膳食叶酸缺乏与血浆同型半胱氨酸水平升高有关，同型半胱氨酸是AD和与年龄相关的认知能力下降的危险因素。越来越多的证据表明，蛋白磷酸酶2A （PP2A）是一个主要的脑酶家族，参与了阿尔茨海默病的发病。PP2A催化亚基可以通过特异性甲基转移酶（PPMT）甲基化，也可以通过专用甲基化酯酶（PME-1）去甲基化，其性质及其在神经元中的调节尚未被探索。甲基化调节PP2A的功能特异性。值得注意的是，PPMT活性在体外被Hey前体s -腺苷- l-同型半胱氨酸（SAH）抑制，而SAH水平在AD脑中升高。值得注意的是，我们的初步数据表明：1)神经元PPMT和PP2A甲基化在AD中下调；2)低叶酸饮食小鼠大脑中去甲基化的PP2A积累；3) PP2A甲基化降低与tau和APP的改变相关。我们的研究计划的目标是进一步验证PPMT是叶酸依赖的Hey代谢途径（AD、PP2A和tau和APP的调节的危险因素）之间的关键中介的假设。因此，叶酸缺乏可能诱导PP2A甲基化下调，并有助于AD的发病机制。在Aim 1中，将利用培养细胞中野生型和突变型蛋白的沉默策略和表达来研究PPMT和PME-1的失调对PP2A、tau、APP和微管调控的影响。在Aim 2中，将在野生型和转基因AD小鼠模型中进一步验证慢性膳食叶酸缺乏抑制PP2A甲基化并促进tau磷酸化和淀粉样蛋白形成的假设。在Aim 3中，将进行Western blot和免疫组织化学分析，比较正常衰老过程中PPMT、PME-1和去甲基化PP2A在人脑区域的分布，并评估其是否在AD和其他tau病变中发生改变。如果我们的模型得到验证，膳食叶酸补充和PPMT可能是新的治疗靶点，旨在减缓阿尔茨海默病的疾病进程，以及可能的其他神经退行性痴呆。

DESCRIPTION (provided by applicant): Alzheimer's disease (AD) is a leading cause of dementia and affects over 4.5 millions Americans at a total cost of at least $100 billion per year. Deregulation of the protein, tau, and altered processing of the amyloid precursor protein (APR) are key events in AD pathogenesis. Clinical studies suggest that dietary folate deficiency is associated with elevated plasma levels of homocysteine (Hey), a risk factor for AD and age- related cognitive decline. There is increasing evidence that protein phosphatase 2A (PP2A), a family of major brain enzymes, is involved in AD pathogenesis. PP2A catalytic subunit can be methylated by a specific methyltransferase (PPMT), and demethylated by a dedicated methylesterase (PME-1), whose properties and regulation in neurons have not yet been explored. Methylation modulates PP2A functional specificity. Notably, PPMT activity is inhibited in vitro by the Hey precursor, S-adenosyl-L-homocysteine (SAH), and SAH levels are elevated in AD brain. Significantly, our preliminary data indicate that: 1) Neuronal PPMT and PP2A methylation are downregulated in AD; 2) demethylated PP2A accumulates in the brain of mice fed a low-folate diet; and 3) reduced PP2A methylation correlates with alterations in tau and APP. The goal of our research proposal is to further test the hypothesis that PPMT is a critical intermediate between folate-dependent Hey metabolic pathways that are a risk factor for AD, PP2A, and the regulation of tau and APP. As such, folate deficiency may induce downregulation of PP2A methylation and contribute to AD pathogenesis. In Aim 1, silencing strategies and expression of wild-type and mutant proteins in cultured cells will be utilized to investigate the effects of deregulating PPMT and PME-1 on the regulation of PP2A, tau, APP and microtubules. In Aim 2, the hypothesis that chronic dietary folate deficiency inhibits PP2A methylation and promotes tau phosphorylation and amyloidogenesis will be further tested in wild-type and transgenic AD mouse models. In Aim 3, Western blot and immunohistochemical analyses will be performed to compare the distribution of PPMT, PME-1 and demethylated PP2A in human brain regions during normal aging, and assess whether it becomes altered in AD and other tauopathies. If our model is validated, dietary folate supplementation and PPMT may be novel therapeutic targets for strategies aimed at slowing down the disease process in AD, and possibly other neurodegenerative dementias.