Mechanisms of LRRK2 Mediated Neurotoxicity

LRRK2 介导的神经毒性机制

基本信息

批准号：
8071508
负责人：
Andrew B West
金额：
$ 31.41万
依托单位：
UNIVERSITY OF ALABAMA AT BIRMINGHAM
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-09-01 至 2015-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8071508
关键词：
Alanine Animal Model Animals Aspartic Acid Biochemical Biological Assay Biological Models Cell Death Cells Complex Corpus striatum structure Data Development Dimerization Disease Disease Progression Dissociation Dopamine Dopaminergic Cell Drug Design Electron Transport Employee Strikes Ethnic Origin Etiology Fiber GTP Binding Genes Guanosine Triphosphate Phosphohydrolases Human Imagery In Vitro Life Link Mass Spectrum Analysis Mechanics Mediating Missense Mutation Molecular Conformation Morbidity - disease rate Mus Mutation Nerve Degeneration Neurons Open Reading Frames Parkinson Disease Pathway interactions Phosphorylation Phosphotransferases Play Population Protein Kinase Protein Kinase Inhibitors Proteins Proteome Regulation Resolution Role Route Signal Transduction Pathway Site Structure Technology Testing Toxic effect Transgenic Mice Ubiquitin United States Viral Vector alpha synuclein cancer type density design dimer disease phenotype disease-causing mutation dopaminergic neuron enzyme activity in vivo in vivo Model insight kinase inhibitor late disease onset leucine-rich repeat kinase 2 mimetics monoamine mortality mutant neuroprotection neurotoxicity novel protein kinase inhibitor public health relevance receptor small molecule therapeutic development therapeutic target vector

项目摘要

DESCRIPTION (provided by applicant): Parkinson's disease (PD) is a major cause of morbidity and mortality in the United States. The etiology is largely unknown and therapies that slow or halt the relentless progression of disease do not yet exist. Dominant missense mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are the most common known specific cause of PD, and LRRK2 mutations associate with disease phenotypes that mimic typical late-onset disease. LRRK2 encodes an unusually large protein kinase, and the most common mutations that cause PD may up-regulate LRRK2 kinase activity. Since small molecule protein kinase inhibitors have made an impact on the treatment of multiple types of cancer, LRRK2 represents a potential therapeutic target for the treatment of PD should kinase activity correlate with neurodegeneration in model systems. Through the design of novel high capacity viral vectors encoding the LRRK2 open-reading frame modified with either PD-causing mutations that up-regulate kinase activity or mutations that inactivate kinase activity, the importance of LRRK2 kinase activity in directing dopaminergic neurodegeneration in vivo will be determined. The specific effects of pathogenic mutations on LRRK2 function and activity are not fully understood, but may allow for a straightforward route to decipher disease-related LRRK2 functions. LRRK2 encodes an active kinase and GTPase protein, a near unique arrangement in the human proteome, and pathogenic mutations can occur in both enzymatic domains. Oligomerization and dimerization are intrinsic mechanisms of regulation for many protein kinases and GTPase proteins. We will biochemically characterize oligomeric and dimeric LRRK2 protein and determine the effect of pathogenic LRRK2 mutations on the composition of LRRK2 conformations and associated activities. Further, we develop technology that may allow direct visualization of LRRK2 enzyme activity in living cells and explore both inhibition and activation of kinase activity. Through the resolution of LRRK2 autophosphorylation sites, we have uncovered an unexpected complexity in the probable reciprocal regulation of kinase and GTPase activities, where kinase activity may play a dual role in mediating GTPase-dependent LRRK2 dimerization and kinase-mediated phosphorylation of substrates. We will characterize the role of LRRK2 autophosphorylation on enzymatic regulation and how PD-associated mutations perturb normal activities. PUBLIC HEALTH RELEVANCE: The biochemical mechanisms underlying disease-causing mutations in the LRRK2 gene, currently the most common known cause of Parkinson's disease, are explored in order to determine potential targets for therapeutics development. Identification of cellular pathways linked with LRRK2-mediated neurodegeneration will further enhance our understanding of Parkinson's disease and spur the development of rationally designed drugs that slow or halt the disease.

描述（由申请人提供）：帕金森氏病（PD）是美国发病率和死亡率的主要原因。病因在很大程度上是未知的，并且尚不存在缓慢或停止疾病进展的疗法。富含亮氨酸的重复激酶2（LRRK2）基因中的显性错义突变是最常见的PD特异性原因，而LRRK2突变与模仿典型的晚期发作疾病的疾病表型相关。 LRRK2编码一种异常大的蛋白激酶，导致PD的最常见突变可能会上调LRRK2激酶活性。由于小分子蛋白激酶抑制剂对多种类型的癌症的治疗产生了影响，因此LRRK2代表了治疗PD的潜在治疗靶标，如果激酶活性与模型系统中的神经变性相关。通过设计编码LRRK2开放式读取框架的新型高容量病毒载体，该框架通过PD引起的突变进行了修改，该突变会引起激酶活性或灭活激酶活性的突变，LRK2激酶活性在指导多巴胺神经化学因素在VIVO中的重要性都可以确定。致病突变对LRRK2功能和活性的具体影响尚不完全了解，但可能可以直接途径来破译与疾病相关的LRRK2功能。 LRRK2编码活性激酶和GTPase蛋白，这是人类蛋白质组中几乎独特的排列，并且在两个酶结构域中都可能发生致病突变。寡聚化和二聚化是许多蛋白激酶和GTPase蛋白调节的内在机制。我们将在生物化学上表征寡聚和二聚体LRRK2蛋白，并确定致病性LRRK2突变对LRRK2构象和相关活性组成的影响。此外，我们开发了可以直接可视化活细胞中LRRK2酶活性的技术，并探索激酶活性的抑制和激活。通过LRRK2自磷酸化位点的分辨率，我们发现了激酶和GTPase活性的可能相互调节的意外复杂性，在这种情况下，激酶活性可能在介导GTPase依赖性的LRRK2二维和激酶介导的副磷酸化的底物中介导GTPase依赖性LRRRK2二维。我们将表征LRRK2自磷酸化对酶调节的作用以及与PD相关的突变如何扰动正常活动。公共卫生相关性：LRRK2基因中引起疾病的突变的生化机制，目前是目前最常见的帕金森氏病原原因，以确定疗法开发的潜在靶标。与LRRK2介导的神经变性相关的细胞途径的鉴定将进一步增强我们对帕金森氏病的理解，并刺激速度缓慢或停止疾病的合理设计的药物的发展。