Understanding the Role GPRC6a During Tau Metabolism

了解 GPRC6a 在 Tau 代谢过程中的作用

• 批准号: 10338842
• 负责人: Daniel Carl Lee
• 金额: --
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-07-01 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10338842
• 关键词: Understanding; Role; GPRC6a; During; Tau

Tauopathies including Alzheimer's disease (AD) consist of age-associated neurodegenerative diseases for which no disease-modifying treatments exist. Our group has uncovered a unique interaction between the arginine metabolism and tauopathies. Arginine metabolism affects multiple biological processes that show considerable influence upon tau biology. We hypothesize that L-arginine acts as a sensor for amino acid abundance and that the depletion of arginine initiates autophagy to replenish amino acid levels. Arginase 1 (Arg1) remains one of several different enzymes capable of metabolizing the semi-essential amino acid L- arginine. In fact numerous isoforms exist for Arg and nitric oxide synthases (NOS). We demonstrate in cells and animal models of tauopathy the benefits of increasing Arg1 in reducing many aspects of the tau phenotype (hallmarks comprised of tau effects). Several seminal findings show lysosomal and cytoplasmic arginine sensors that modulate mechanistic target of rapamycin complex 1 (mTORC1). GPRC6a is a G-protein coupled receptor that binds L-α amino acids including L-arginine and may serve as an extracellular arginine sensor for mTORC1. Our preliminary data indicates increased activation of the arginine-sensing mTORC1 pathway in AD brains and animal models of tauopathies. We show that arginine producing and metabolizing enzymes, arginine sensors, and mTORC1 complexes increase in the hippocampus of AD patients compared to control aged matched brains. We also find that tau increased total arginine levels in the brain by 35%, increased basal levels of extracellular arginine and arginine release following neuronal stimulation in the brain. Furthermore, novel allosteric antagonist to GPRC6a clears various forms of tau in cell lines, primary neurons, and mice. Genetic reduction of GPRC6a also reduced tau expression and seemingly regulates several intracellular arginine sensors. We posit that tauopathies (including AD) cause impaired arginine metabolism and uncoupling of arginine-sensing mTORC1 signaling, which lead to hyper-mTORC1 activation creating a positive feed- forward loop to augment the tau phenotype. We will test the hypothesis that decreased GPRC6a signaling from allosteric antagonists or shRNA to GPRC6a modulate arginine-sensing mTORC1 signaling, induces autophagy and increases tau clearance in vitro and in vivo. We will use tetracycline inducible tau cell lines with a photo- switchable fluorescent tracer to measure tau metabolism, split tau-GFP constructs that measure tau oligomerization, and primary neurons to determine the role of GPRC6a and tau fate. We will determine if genetic knockdown and pharmacological inhibition of GPRC6a impacts tau biology in vitro and in vivo. Success in this application would provide a new receptor target that may impact arginine-sensing mTORC1 signaling. Furthermore, we aim to identify an entirely new class of drugs that targets GPCR6a to mitigate the tau phenotype provide new strategies for tauopathies and AD.

包括阿尔茨海默氏病 (AD) 在内的 Tau蛋白病是一种与年龄相关的神经退行性疾病。 我们的研究小组发现了一种独特的相互作用。 精氨酸代谢和 tau蛋白病会影响多种生物过程。 我们勇敢地承认 L-精氨酸可以作为氨基酸的传感器。 丰度并且精氨酸的消耗会启动自噬以补充精氨酸酶 1 的水平。 (Arg1) 仍然是能够代谢半必需氨基酸 L- 的几种不同酶之一 事实上，我们在细胞中证明了精氨酸和一氧化氮合酶 (NOS) 存在多种同工型。 和 tau 蛋白病动物模型 增加 Arg1 在减少 tau 表型的许多方面的益处 （由 tau 效应组成的标志）。几个开创性的发现显示了溶酶体和细胞质精氨酸。 调节雷帕霉素复合物 1 (mTORC1) 机械靶点的传感器是一种 G 蛋白偶联物。 结合 L-α 氨基酸（包括 L-精氨酸）的受体，可作为细胞外精氨酸传感器 mTORC1。我们的初步数据表明 AD 中精氨酸感应 mTORC1 通路的激活增加。 我们证明精氨酸产生和代谢酶， 与对照组相比，AD 患者海马中的精氨酸传感器和 mTORC1 复合物增加 我们还发现，tau 蛋白使大脑中的总精氨酸水平增加了 35%，增加了基础值。 大脑神经元刺激后细胞外精氨酸和精氨酸释放的水平。 GPRC6a 的新型变构拮抗剂可清除细胞系、原代神经元和小鼠中各种形式的 tau 蛋白。 GPRC6a 的基因减少也减少了 tau 表达，并且似乎调节了多种细胞内 我们认为 tau蛋白病（包括 AD）会导致精氨酸代谢受损和解偶联。 精氨酸感应 mTORC1 信号传导，导致 mTORC1 过度激活，产生正向反馈 我们将测试减少 GPRC6a 信号传导的假设。 GPRC6a 的变构拮抗剂或 shRNA 调节精氨酸感应 mTORC1 信号传导，诱导自噬 并增加体内和体外的 tau 清除率。我们将使用具有光诱导作用的四环素诱导 tau 细胞系。 用于测量 tau 代谢的可切换荧光示踪剂，用于测量 tau 的分裂 tau-GFP 结构 寡聚化和初级神经元的作用，以确定 GPRC6a 和 tau 的命运。 GPRC6a 的基因敲除和药理学抑制影响 tau 体外和体内生物学的成功。 该应用将提供一个可能影响精氨酸感应 mTORC1 信号传导的新受体靶点。 此外，我们的目标是找到一类全新的药物，以 GPCR6a 为靶点，以减轻 tau 蛋白的影响。 表型为 tau蛋白病和 AD 提供新策略。