DJ-1 and neurodegeneration: its roles in mitochondria and in protein misfolding

帕金森病（PD）是仅次于阿尔茨海默病的第二种最常见的神经退行性疾病，其特征在于多巴胺能神经元的损失和路易体形式的错误折叠的α-突触核蛋白的积累。临床症状包括肌肉僵硬、静止性震颤、运动迟缓和姿势不稳定，这是由于黑质延髓部多巴胺能神经元的丢失。在后期阶段，虽然异常的α-突触核蛋白聚集和氧化应激可能导致特定神经元群体的死亡，但散发性PD中神经元丢失的病因尚不清楚。虽然对PD的发病机制知之甚少，但对与家族性PD相关的几个基因的详细研究正在增强对与疾病相关的分子机制的理解。其中一个基因突变-PARK 7-占早发性隐性PD散发病例的约1-2%。PARK 7编码DJ-1，DJ-1是一种小的保守蛋白，广泛表达并主要定位于细胞质，但也存在于细胞核中并与线粒体相关。尽管这种蛋白质的重要性，DJ-1在PD形成中的作用仍不清楚。DJ-1有几种功能，包括氧化应激传感器，处理细胞中错误折叠蛋白质的伴侣蛋白，以及线粒体功能的调节剂。在这个项目中，我们试图澄清DJ-1如何有助于线粒体功能和蛋白质错误折叠，并最终如何在这些细胞过程中的缺陷，由于DJ-1突变有助于PD的发病机制。对于这些研究，我们计划实施永生化细胞系，小鼠的原代细胞，以及果蝇。在初步实验中，我们观察到DJ-1的突变改变了其在氧化应激条件下定位于哺乳动物细胞线粒体的能力。因此，我们现在寻求进一步表征这种影响与不同的氧化剂，也在α-突触核蛋白过表达的背景下。此外，我们计划研究这些突变体对小鼠细胞线粒体形态和功能的影响，以及对线粒体基因表达的影响。最后，我们计划通过研究DJ-1突变体对果蝇线粒体结构和功能的影响来扩展这种方法。接下来，我们试图通过研究DJ-1和Tau蛋白之间的关系，进一步探索DJ-1在蛋白质错误折叠中的作用，Tau蛋白与几种神经退行性疾病有关。Tau蛋白参与阿尔茨海默病的发病机制，并且是这种疾病中存在的神经元缠结的主要组分。编码Tau的基因最近被遗传地与PD相关联。此外，DJ-1与阿尔茨海默病和其他脑部疾病中的Tau包涵体共定位。然而，尚未研究DJ-1和Tau之间任何可能的物理/功能相互作用，并且尚未探索它们在PD背景下的关系。因此，我们建议在活的哺乳动物细胞中研究DJ-1和Tau之间的相互作用，以及在多大程度上，如果有的话，这会改变Tau的毒性或纤维的形成。此外，我们计划将这项工作扩展到Tauopathies的果蝇模型，以确定DJ-1是否调节这些动物的疾病相关“症状”。在初步工作中，我们已经观察到DJ-1和Tau在哺乳动物细胞中表达时直接相互作用。因此，我们的工作将更好地确定DJ-1在线粒体功能和蛋白质错误折叠中的作用，这将有助于阐明其在PD中的致病作用。这增加了对DJ-1生物学的了解，最终可能有助于为这种和其他神经退行性疾病的治疗策略提供信息。

Parkinson's disease (PD), the second most common neurodegenerative disorder after Alzheimer's disease, is characterized by loss of dopaminergic neurons and the accumulation of misfolded alpha-synuclein in the form of Lewy bodies. The clinical symptoms include muscle rigidity, resting tremor, bradykinesia and postural instability as a result of the loss of dopaminergic neurons in the substantia nigra pars compacta. At later stages, the etiology of neuronal loss in sporadic PD is unknown, although aberrant alpha-synuclein aggregation and oxidative stress may contribute to the death of specific neuronal populations. Although little is known about the pathogenesis of PD, the detailed study of several genes associated with familial PD is enhancing the understanding of the molecular mechanisms associated with the disease. Mutations in one of these genes - PARK7 - accounts for ~1-2% of the sporadic cases of early onset recessive PD. PARK7 encodes for DJ-1, a small conserved protein which is broadly expressed and primarily localized to the cytoplasm, but also found in the nucleus and associated with mitochondria. Despite the importance of this protein, the role of DJ-1 in in formation of PD remains unclear. Several functions have been suggested for DJ-1, including an oxidative stress sensor, a chaperone which deals with misfolded protein in the cells, and a regulator of mitochondrial function. In this project we seek to clarify how DJ-1 contributes to mitochondrial function and protein misfolding, and ultimately how defects in these cellular processes due to DJ-1 mutations contribute to pathogenesis in PD. For these studies we plan to implement immortalized cell lines, primary cells from mice, as well as fruit flies. In preliminary experiments we have observed that mutations in DJ-1 alter its ability to localize to mitochondria in mammalian cells under oxidative stress conditions. We thus now seek to further characterize this effect with varied oxidants, and also in the context of alpha-synuclein overexpression. In addition, we plan to investigate the impact of these mutants on both morphology and function of mitochondria in mouse cells, as well as the effects on mitochondrial gene expression. Finally we plan to extend this approach by studying the effect of DJ-1 mutants on the structure and function of mitochondria in fruit flies. Next we seek to further explore the role of DJ-1 in protein misfolding by investigating the relationship between DJ-1 and the Tau protein, which has been implicated in several neurodegenerative disorders. The Tau protein is involved in the pathogenesis of Alzheimer's disease and is a major component of neurofibrillary tangles present in this disorder. The gene encoding Tau has recently been genetically linked to PD. In addition, DJ-1 co-localizes with Tau inclusions in Alzheimer's disease and other brain disorders. However any possible physical/functional interactions between DJ-1 and Tau have not been studied and their relationship in the context of PD has not been explored. Thus we propose to study the interaction between DJ-1 and Tau in living mammalian cells, and to what extent, if any, this alters Tau toxicity or formation of fibers. Furthermore, we plan to extend this work to a fruit fly model of Tauopathies, to ascertain whether DJ-1 modulates disease-relevant "symptoms" in these animals. In preliminary work we have observed that DJ-1 and Tau directly interaction when expressed in mammalian cells. Thus, our work will better define the role of DJ-1 in mitochondrial function and in protein misfolding which will help clarify its pathogenic role in PD. This added insight into DJ-1 biology may ultimately help inform therapeutic strategies for this, and other, neurodegenerative disorders.