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，影响超过450万美国人，每年的总成本至少为1000亿美元。tau蛋白的失调和淀粉样前体蛋白（APR）的加工改变是AD发病机制中的关键事件。临床研究表明，膳食叶酸缺乏与血浆同型半胱氨酸（Hey）水平升高有关，这是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（PP 2A），一个主要的脑酶家族，参与AD的发病机制。PP 2A催化亚基可以被一种特异性的甲基转移酶（PPMT）甲基化，也可以被一种特异性的甲基酯酶（PME-1）去甲基化，但其在神经元中的性质和调控尚不清楚。甲基化调节PP 2A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-高半胱氨酸（SAH）抑制，并且在AD脑中SAH水平升高。值得注意的是，我们的初步数据表明：1）神经元PPMT和PP 2A甲基化在AD中下调; 2）去甲基化的PP 2A在低叶酸饮食的小鼠脑中积累;以及3）PP 2A甲基化的降低与tau和APP的改变相关。我们的研究建议的目标是进一步检验PPMT是叶酸和ATP之间的关键中间体的假设。因此，叶酸缺乏可诱导PP 2A甲基化的下调并有助于AD发病机制。在目标1中，将利用沉默策略和野生型和突变蛋白在培养细胞中的表达来研究去调节PPMT和PME-1对PP 2A、tau、APP和微管的调节的影响。在目标2中，将在野生型和转基因AD小鼠模型中进一步测试慢性膳食叶酸缺乏抑制PP 2A甲基化并促进tau磷酸化和淀粉样蛋白生成的假设。在目标3中，将进行蛋白质印迹和免疫组织化学分析，以比较正常衰老期间人脑区域中PPMT、PME-1和去甲基化PP 2A的分布，并评估其是否在AD和其他tau蛋白病中发生改变。如果我们的模型得到验证，膳食叶酸补充剂和PPMT可能是新的治疗目标，旨在减缓AD和其他神经退行性痴呆的疾病进程。

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.