Glutaredoxin, a Critical Regulator of Parkinson Disease Pathogenesis

我们研究的总体目标是确定和验证治疗帕金森病的新靶点 疾病（PD）。PD的特征在于多巴胺能神经元的损失，多巴胺能神经元特别容易受到 氧化应激谷氧还蛋白（Grx）是一种抗氧化防御酶，可以抵消氧化应激， 通过催化蛋白质-谷胱甘肽混合二硫化物在 半胱氨酸残基（蛋白质-SSG）。这种氧化修饰与许多人的功能改变有关。 参与细胞存活的蛋白质，包括先前已鉴定为家族性形式的 警局利用C.线虫作为模式生物，我们已经发现Grx 1的线虫同源物的缺陷， 哺乳动物中Grx的主要同种型，导致由 突变体LRRK 2（G2019 S或R1441 C）、γ-突触核蛋白或酪氨酸羟化酶的过表达。这些发现 表明Grx 1缺乏可在动物模型中导致PD相关表型，这表明 Grx 1缺陷有助于PD发病机制。支持这些遗传模型的相关性，我们有 初步证明PD患者死后脑组织中Grx 1含量降低。 因此，我们建议在哺乳动物模型中表征和验证Grx 1的神经保护作用。 并研究家族性PD中涉及的关键蛋白谷胱甘肽化状态的变化是否也 对疾病的发病机制有重要作用。为了检查Grx 1缺陷是否会导致 在哺乳动物中的多巴胺能变性，我们在目的1中提出了一种新的小鼠模型， 将现有的转基因人类LRRK 2突变动物与Grx 1敲除动物杂交。转基因 Grx 1基因敲除的LRRK 2突变小鼠代表了遗传工程改造的哺乳动物模型， 升高的氧化应激和巯基稳态的特异性缺乏。我们将决定这部小说 小鼠模型表现出PD特征性的多巴胺能神经元的损失。这项研究的结果将 验证Grx 1是否是PD发病机制的关键调节因子。在目标2中，我们将确定氧化半胱氨酸 PD发病机制中涉及的蛋白质（包括突触核蛋白）的功能修饰和变化， parkin、UCH-L1、DJ-1、PINK 1和LRRK 2。半胱氨酸修饰对PD样表型的贡献 将通过使用哺乳动物细胞和C. elegans PD模型神经变性易感性与蛋白质氧化修饰的对应关系 与家族性PD有关的研究将为理解潜在的致病机制提供重要的进展 偶发性PD。

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.