Neuroprotective mechanism of DJ-1 in Parkinson's disease

描述（由申请人提供）：最近在帕金森病（PD）的常染色体隐性形式中发现了DJ-1的功能丧失突变。DJ-1在正常和病理状态下的作用尚不清楚，其假定功能的证据一直存在争议。DJ-1在保护神经元免受氧化应激中的作用在理解帕金森病的这种遗传形式和帕金森病的散发形式中的细胞死亡的潜在共同机制方面特别有用，帕金森病的散发形式被认为是由线粒体功能障碍产生的氧化应激引起的。虽然DJ-1缺陷细胞对氧化应激更敏感，但DJ-1似乎不具有活性氧（ROS）的直接清除作用。相反，DJ-1可能作用于ROS的下游途径。具体而言，DJ-1可能是重要的泛素化和DJ-1-空小鼠脑衰减增加泛素化反应氧化应激。DJ-1也被证明具有伴侣功能。DJ-1在氧化应激下转变为酸性形式。为了研究DJ-1缺陷引起的帕金森病的发病机制，我们成功地产生了DJ-1基因敲除小鼠。我们的初步研究表明，DJ-1基因敲除小鼠出现与多巴胺神经传递改变相关的年龄依赖性运动缺陷，这与多巴胺转运蛋白（DAT）功能亢进一致。基于这些观察，我们假设DJ-1促进靶蛋白如DAT的泛素化和随后的降解，并且这种功能被氧化应激激活。我们提出以下实验来验证这些假设，并确定DJ-1在维持最佳多巴胺传递和细胞存活中发挥作用的靶分子和途径。目标1.研究DJ-1基因敲除小鼠的多巴胺神经元是否更容易受到衰老和百草枯攻击的影响。我们将研究衰老和百草枯治疗对DJ-1基因敲除小鼠生化、行为和解剖学特性的影响。目标2.研究DJ-1是否影响氧化应激下的泛素蛋白酶体途径。我们将检查总的泛素化蛋白水平，并比较DJ-1-null和WT脑中多巴胺能神经元死亡所涉及的特异性底物蛋白的相对水平。此外，我们将研究DJ-1对泛素-蛋白酶体系统的影响如何改变DAT功能，通过测量其表面表达和泛素化。目标3。研究DJ-1的分子伴侣功能及其与泛素途径的氧化依赖性协同作用。了解DJ-1的神经保护功能可能会导致帕金森病的合理治疗，以防止其表现或进展。由于帕金森病导致相当一部分老年人严重残疾，这些发现将对公共卫生产生重大影响。

DESCRIPTION (provided by applicant): Loss-of-function mutations in DJ-1 were recently identified in an autosomal recessive form of Parkinson's disease (PD). The role of DJ-1 in normal and pathological states is still unclear, and evidences for its putative functions have been quite controversial. The role of DJ-1 in protecting neurons from oxidative stress is particularly informative in understanding a potential common mechanism for cell death in this genetic form of Parkinson's disease and in sporadic forms of Parkinson's disease which is thought to result from oxidative stress generated by mitochondrial dysfunction. Although DJ-1 deficient cells are more susceptible to oxidative stress, DJ-1 does not seem to have direct scavenging effect of reactive oxygen species (ROS). Instead, DJ-1 may work on a downstream pathway from ROS. Specifically, DJ-1 may be important in ubiquitination and DJ-1-null mouse brain attenuated increase in ubiquitination in response to oxidative stress. DJ-1 has been also shown to have chaperone function. DJ-1 changes into an acidic form under oxidative stress. To investigate the pathogenesis of Parkinson's disease caused by DJ-1 deficiency, we have successfully generated DJ-1-null mice. Our preliminary studies indicate that DJ-1-null mice develop age-dependent motor deficits associated with alteration of dopamine neurotransmission, consistent with hyperactive dopamine transporter (DAT) function. Based on these observations, we hypothesize that DJ-1 facilitates ubiquitination and subsequent degradation of target proteins such as DAT and this function is activated by oxidative stress. We propose the following experiments to test these hypotheses and identify the target molecules and pathways in which DJ-1 plays a role to maintain optimal dopamine transmission and cell survival. Aim 1. To investigate whether dopamine neurons in DJ-1-null mice are more vulnerable to aging and paraquat challenge. We will examine the effect of aging and paraquat treatment in DJ-1-null mouse for its biochemical, behavioral and anatomical properties. Aim 2. To investigate whether DJ-1 influences the ubiquitin proteasomeal pathway under oxidative stress. We will examine the total ubiquitinated proteins levels and compare the relative levels of specific substrate proteins implicated in dopaminergic neuronal death in DJ-1-null and WT brains. In addition, we will examine how the effect of DJ-1 on ubiquitin-proteasome system alters DAT function by measuring its surface expression and ubiquitination. Aim 3. To investigate the chaperone function of DJ-1 and its oxidation dependent cooperation with ubiquitin pathway. Relevance Understanding the neuroprotective function of DJ-1 may lead to rational therapy for Parkinson's disease to prevent the manifestation or progression. Since Parkinson's disease leads to significant disability in a significant portion of the elderly population, such findings will have a significant impact in public health.