Glutaredoxin, a Critical Regulator of Parkinson Disease Pathogenesis

我们研究的总体目标是确定和验证帕金森氏症治疗的新靶点 疾病(PD)。帕金森病的特征是多巴胺能神经元的丧失，这些神经元特别容易患上帕金森病 氧化应激。谷氧还蛋白(GRX)是一种抗氧化防御酶，能中和氧化应激和 通过催化蛋白质-谷胱甘肽混合二硫化物的可逆形成维持硫醇动态平衡 半胱氨酸残基(蛋白质-SSG)。这种氧化修饰与许多功能的改变有关 参与细胞生存的蛋白质，包括先前已被鉴定为家族形式的蛋白质 警察。以线虫为模式生物，我们发现Grx1的线虫同源物中存在缺陷， GRX在哺乳动物中的主要亚型，导致由 突变的LRRK2(G2019S或R1441C)、突触核蛋白或酪氨酸羟基酶的过度表达。这些发现 提示Grx1缺乏在动物模型中易患帕金森病相关表型。 Grx1缺乏参与了帕金森病的发病。支持这些遗传模型的相关性，我们有 获得了帕金森病患者死后脑内Grx1含量降低的初步证据。 因此，我们建议表征和验证Grx1在哺乳动物动物模型中的神经保护作用。 并探讨家族性帕金森病关键蛋白谷胱甘肽基化状态的改变是否也与家族性帕金森病有关。 在疾病的发病机制中起着重要作用。检查Grx1缺乏是否会使人易受 哺乳动物的多巴胺能变性，我们在目标1中提出了一种新的小鼠模型 使现有的转基因人类LRRK2突变动物与Grx1基因敲除动物杂交。转基因生物 带有Grx1基因敲除的LRRK2突变小鼠代表了一种基因工程哺乳动物动物模型 氧化应激增加和硫醇稳态的特异性缺陷。我们将确定这部小说是否 小鼠模型表现为帕金森病特有的多巴胺能神经元的丢失。这项研究的结果将是 验证Grx1是否是帕金森病发病的关键调节因子。在目标2中，我们将鉴定氧化半胱氨酸 参与帕金森病发病机制的蛋白质的修饰和功能变化，包括突触核蛋白， Parkin、UCH-L1、DJ-1、PINK1和LRRK2。半胱氨酸修饰对PD样表型的影响 将通过使用哺乳动物细胞和线虫中的非氧化性突变形式的蛋白质进行检测 帕金森病的模型。神经变性易感性与蛋白质氧化修饰的对应关系 家族性帕金森病的发生将为理解其致病机制提供重要的进展 散发性帕金森病。

The overall objective of our research is to identify and validate novel targets for the treatment of Parkinson's disease (PD). PD is characterized by loss of dopaminergic neurons, which are especially vulnerable to oxidative stress. Glutaredoxin (Grx) is an antioxidant defense enzyme that counteracts oxidative stress and maintains thiol homeostasis by catalyzing the reversible formation of protein-glutathione mixed disulfides on cysteine residues (protein-SSG). This oxidative modification is implicated in change of function for many proteins involved in cell survival, including proteins that have been previously identified with familial forms of PD. Using C. elegans as a model organism, we have found that deficiency in the nematode homolog of Grx1, the major isoform of Grx in mammals, leads to exacerbation of dopaminergic degeneration elicited by overexpression of mutant LRRK2 (G2019S or R1441C), ¿-synuclein, or tyrosine hydroxylase. These findings indicate that Grx1 deficiency can predispose to PD-relevant phenotype in an animal model, suggesting that Grx1 deficiency contributes to PD pathogenesis. Supporting the relevance of these genetic models, we have obtained preliminary evidence that Grx1 content is decreased in postmortem brains of PD patients. Accordingly, we propose to characterize and validate the neuroprotective role of Grx1 in mammalian models of PD, and to investigate whether changes in glutathionylation status of key proteins implicated in familial PD also contribute importantly to the disease pathogenesis. To examine if Grx1 deficiency confers vulnerability to dopaminergic degeneration in mammals, we propose in Aim 1 to characterize a novel mouse model generated from crossing existing transgenic human LRRK2 mutant animals with Grx1-knockout animals. The transgenic LRRK2 mutant mice with Grx1-knockout represent a genetically engineered mammalian animal model of elevated oxidative stress and specific deficiency in thiol homeostasis. We will determine whether this novel mouse model manifests loss of dopaminergic neurons characteristic of PD. The outcome of this study would validate if Grx1 serves as a critical regulator of PD pathogenesis. In Aim 2, we will identify oxidative cysteine modifications and changes in function of the proteins implicated in PD pathogenesis including ¿-synuclein, parkin, UCH-L1, DJ-1, PINK1, and LRRK2. The contribution of cysteine modifications to PD-like phenotype will be examined by the use of non-oxidizable mutant forms of the proteins in mammalian cell and C. elegans models of PD. Correspondence between susceptibility to neurodegeneration and oxidative modification of proteins implicated in familial PD would provide an important advance in understanding pathogenic mechanisms underlying sporadic PD.