Gene Environment Interactions in Parkinson's Disease

帕金森病的基因环境相互作用

基本信息

批准号：
7894952
负责人：
KIM TIEU
金额：
$ 19.13万
依托单位：
UNIVERSITY OF ROCHESTER
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-07-16 至 2012-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7894952
关键词：
1-Methyl-4-phenylpyridinium Attenuated Biological Assay Cell Death Cell Survival Cells Cessation of life Chemicals Chimeric Proteins Data Dominant-Negative Mutation Dopamine Dose Electron Transport Experimental Models Exposure to Gene Mutation Genetic Goals Herbicides High Pressure Liquid Chromatography Human Immunoblotting Induced Mutation Intervention Lead Measures Mediating Mitochondria Molecular Mutation Nerve Degeneration Neurodegenerative Disorders Neurotoxins PTEN-induced putative kinase Paraquat Parkinson Disease Pathogenesis Pathway interactions Patients Phosphotransferases Play Polarography Predisposition Process Production Proteins Reporting Research Role Shapes Small Interfering RNA System Testing Time Toxic Environmental Substances Toxic effect Translating base design functional outcomes gene environment interaction genetic manipulation inhibitor/antagonist insight mitochondrial dysfunction mutant neuroprotection neurotoxicity novel overexpression prevent public health relevance research study response small molecule stable cell line toxicant vector vector control

项目摘要

DESCRIPTION (provided by applicant): Our long term goal is to study the mechanism of neurodegeneration induced by environmental neurotoxicants. This proposal is submitted to investigate gene-environment interactions in the pathogenesis of Parkinson's disease mediated through the mitochondrial pathway. Mitochondrial dysfunction has been proposed as a major mechanism of neurodegeneration in PD for years, but direct evidence in humans was inadequate until the recent discoveries of mutations in PTEN-induced putative kinase 1 (PINK1), which encodes a mitochondrial kinase. Although PINK1 mutations are associated with autosomal recessive PD, interestingly, an increasing number of PD patients carrying single heterozygous mutations have been reported. These observations suggest that a heterozygous PINK1 mutation may act as a susceptibility factor that interplays with environmental insults. To determine the interactions between PINK1 mutations and environmental toxicants, we have created stable cell lines with inducible over-expression of various PINK1 mutants. Based on our preliminary results, we hypothesize that PINK1 mutations increase cell susceptibility to environmental toxicants such as paraquat (PQ) through a novel mechanism: mitochondrial fragmentation via the mitochondrial fission / fusion machinery. In the first specific aim, we will characterize neurotoxicity induced by PQ and MPP+ (two toxicants representing different mechanisms of toxicity) through the mitochondrial fission and fusion pathway in N27 cells with a PINK1 mutation (L347P) and empty vector control, as well as in N27 cells with PINK1 knockdown mediated by siRNA. We will assess cell viability, functional outcomes (ATP production, dopamine release and electron transport chain activity), as well as mitochondrial fragmentation (size/shape) and alterations in mitochondrial fission and fusion proteins. In specific aim 2, we will perform neuroprotective experiments against PQ and MPP+ toxicity in mutant PINK1 cells by targeting the mitochondrial fission and fusion pathway, through genetic manipulations and a small molecule. We will transfect cells with relevant constructs and use a chemical inhibitor to attenuate mitochondrial fragmentation. The neuroprotective effects of these two strategies against MPP+ and PQ toxicity will be determined using cell viability and functional assays as described in specific aim 1. PUBLIC HEALTH RELEVANCE This research may provide insights into the complexity of gene-environment interactions in the cause of cell death as seen in Parkinson's disease by unraveling a still unrecognized molecular pathway. Furthermore, this novel mechanism may offer an additional avenue to develop neuroprotective therapy for patients with Parkinson's disease.

描述（由申请人提供）：我们的长期目标是研究由环境神经毒性引起的神经变性机制。该提议提交了研究通过线粒体途径介导的帕金森氏病的发病机理中的基因环境相互作用。多年来，已经提出了线粒体功能障碍为PD神经退行性的主要机制，但是人类的直接证据不足，直到最近发现PTEN诱导的假定激酶1（PINK1）突变的发现，该突变编码了线粒体激酶。尽管PINK1突变与常染色体隐性PD有关，但有趣的是，携带单杂合突变的PD患者数量越来越多。这些观察结果表明，杂合的PINK1突变可能是与环境损害相互作用的敏感因素。为了确定Pink1突变与环境有毒物质之间的相互作用，我们创建了稳定的细胞系，具有诱导的各种Pink1突变体的过表达。基于我们的初步结果，我们假设PINK1突变通过一种新的机制来增加细胞对环境有毒物质（例如Paraquat（PQ））的敏感性：通过线粒体裂变 /融合机械的线粒体碎片化。在第一个具体目的中，我们将通过线粒体裂变和粉红色1突变（L347p）和空位矢量控制的N27细胞中的N27细胞中的线粒体裂变和融合途径来表征PQ和MPP+（两种代表毒性不同机制）引起的神经毒性。我们将评估细胞活力，功能结果（ATP产生，多巴胺释放和电子传输链活性）以及线粒体碎片（尺寸/形状）以及线粒体裂变和融合蛋白的改变。在特定的目标2中，我们将通过基因操纵和小分子靶向突变体PINK1细胞中PQ和MPP+毒性的神经保护实验。我们将使用相关构建体转染细胞，并使用化学抑制剂减轻线粒体碎片。这两种针对MPP+和PQ毒性的神经保护作用将使用特定目的1中所述的细胞活力和功能测定确定。公共卫生相关性，这项研究可能会在帕金森氏病中发现的细胞死亡的复杂性，从而对基因 - 环境相互作用的复杂性进行见解，从而通过未识别的是未识别的一条未识别的分子途径。此外，这种新型机制可能会为帕金森氏病患者开发神经保护疗法提供额外的途径。