Regulation of PTEN-induced kinase 1 (PINK 1)

描述（由申请人提供）：帕金森病是一种以多巴胺能中脑神经元死亡为特征的衰弱性运动障碍。最近在家族性帕金森综合征和一些早发性散发性帕金森病人群中发现了PTEN诱导的推定激酶1（PINK 1）突变。为了评估突变的潜在致病作用，有必要首先了解野生型PINK 1的调控和功能。一个尚未解决的关键问题是PINK 1是否是一种功能性激酶。该提案旨在研究一个新的研究方向的可行性，该方向专注于帕金森神经毒素引起的应激条件下PINK 1磷酸化，定位和功能的调节。我们以前的研究表明，神经靶向激酶通路在毒素诱导的多巴胺能细胞死亡中起着核心作用。因此，我们假设PINK 1是一种功能性激酶，与定位于线粒体的一组细胞死亡调节激酶相互作用。为了解决这一假设，我们将使用无细胞和培养系统来确定PINK 1是否显示出受已知的神经靶向信号通路调节并参与其中的激酶活性。我们将确定PINK 1的亚细胞定位是否在多巴胺能细胞死亡的神经毒素模型中受到调节，并比较PINK 1和激酶死亡的PINK 1突变体对多巴胺能细胞系和原代中脑培养物中线粒体细胞死亡的影响。这些研究的完成将有助于更好地了解PINK 1在多巴胺能神经元细胞损伤中的作用，以及它如何参与影响这些细胞存活或死亡的信号网络。在这个探索/发展阶段之后，更广泛的研究将旨在确定PINK 1激活的下游靶点以及疾病相关突变对PINK 1调节的影响，因为这些可能导致新的治疗方法。此外，一组免疫化学和分子试剂将被创建，可以促进其他研究人员研究帕金森氏症和相关疾病机制的工作。

DESCRIPTION (provided by applicant): Parkinson's disease is a debilitating movement disorder characterized by death of dopaminergic midbrain neurons. Mutations in PTEN-induced putative kinase 1 (PINK1) have been recently identified in familial parkinsonism, and some populations of early onset sporadic Parkinson's disease. In order to assess potential causative effects of mutations, it is necessary to first understand the regulation and function of wild type PINK1. One key unresolved issue is whether PINK1 is a functional kinase. This proposal is designed to investigate the feasibility of a novel research direction focused upon the regulation of PINK1 phosphorylation, localization, and function under conditions of stress elicited by parkinsonian neurotoxins. Our previous studies suggest that mitochondrially targeted kinase pathways play a central role in toxin-induced dopaminergic cell death. Thus, we hypothesize that PINK1 is a functional kinase that interacts with a set of cell death- regulatory kinases that localize to mitochondria. To address this hypothesis, we will determine if PINK1 shows kinase activity that is regulated by and participates in known mitochondrially targeted signaling pathways, using both cell free and culture systems. We will determine whether subcellular localization of PINK1 is regulated in neurotoxin models of dopaminergic cell death, and compare the effects of PINK1 and a kinase dead PINK1 mutant on mitochondrial cell death in dopaminergic cell lines and primary midbrain cultures. Completion of these studies will lead to a better understanding of the role of PINK1 during dopaminergic neuronal cell injury, and how it participates in signaling networks that influence whether these cells live or die. Following this exploratory/developmental phase, more extensive studies will be aimed at identifying downstream targets of PINK1 activation and the effects of disease-associated mutations on PINK1 regulation, as these may lead to novel therapeutic approaches. In addition, a group of immunochemical and molecular reagents will have been created that can facilitate the work of other investigators studying mechanisms of Parkinson's and related diseases.