LRRK2 Biology in Parkinson's Disease

项目摘要-项目4：帕金森氏病的LRRK2生物学 帕金森病(PD)是一种复杂的神经退行性疾病，既有散发性的，也有家族性的。 富含亮氨酸的重复蛋白2(LRRK2)的突变最近被证明导致4%的常染色体 世界范围内主要的家族性病例和1%的散发病例。多发性骨髓瘤的临床和病理表型 LRRK2PD患者与经典的晚发性帕金森病相似，LRRK2基因敲除动物对α- 突触核蛋白多巴胺能(DA)神经退行性变，进一步强调了该基因的潜在重要性。 这个项目将解决异常的激酶活性如何导致疾病的机制。 我们的假设是，导致LRRK2 GS的疾病通过致病引起翻译去调控 S15的磷酸化，导致蛋白表达改变和神经元功能障碍和死亡。 全面了解mRNA翻译、转录组和细胞周期蛋白的分子变化 LRRK2GS诱导的蛋白质组是理解DA脆弱性所必需的。还有一个重要的 与α-突触核蛋白的相互作用影响DA脆弱性，但其机制尚不清楚。 目的1解决关于增加的激酶活性的重要性的持续争议 建立LRRK2GS突变对DA神经元活性的影响，建立LRRK2GS/DA小鼠模型。 这些小鼠也提供了一个模型来探索LRRK2的非激酶作用，而LRRK2是从 菲尔德。随着时间的推移，将监测行为、神经解剖学和神经化学的变化。功能界别 LRRK2和α之间的相互作用-突触核蛋白毒性和传递将在LRRK2KO和 确定LRRK2功能缺失的转基因株系是保护性的，获得的LRRK2功能是有毒的。 目标2将解决新的观察结果，即磷酸化的s15是LRRK2GS的致病靶点， S15和磷酸化s15的表达将在转基因小鼠和人类死后组织中进行监测。它 将确定拟磷酸盐s15是否足以引起神经变性，以及是否磷缺乏s15 可以在全球和空间受限的模型中提供对LRRK2 GS神经毒性的保护。这些研究 将在体内确认s15是否是LRRK2GS的致病底物。 目标3将定义在mRNA翻译、转录组和蛋白质组使用的具体变化 基因工程小鼠和人类多巴胺能培养中的先进技术。两者的使用 老鼠和人类模型允许在完整的大脑中进行评估，同时研究导致 到相关人类系统中的慢性神经变性。无偏的综合数据集将是 对所有警局调查人员来说都是宝贵的资源。 该项目的目标是确定神经退行性变信号级联中的结点，这些结点可以 为今后帕金森病治疗策略的发展提供新的靶点。

PROJECT SUMMARY - PROJECT 4: LRRK2 BIOLOGY IN PARKINSON'S DISEASE Parkinson's disease (PD) is a complex neurodegenerative disorder that is both sporadic and familial. Mutations in the leucine-rich repeat kinase 2 (LRRK2) have recently been shown to result in 4% of autosomal dominant familial cases and 1% of sporadic cases worldwide. The clinical and pathological phenotypes of LRRK2 PD patients are similar to classic late-onset PD and LRRK2 knock out animals are resistant to α- synuclein dopaminergic (DA) neurodegeneration, further emphasizing the potential importance of this gene. This project will address mechanisms of how aberrant kinase activity leads to disease. Our hypothesis is that disease causing LRRK2 GS elicits translational deregulation through pathogenic phosphorylation of s15, leading altered protein expression and neuronal dysfunction and death. Comprehensive understanding of the molecular changes in mRNA translation, the transcriptome and the proteome elicited by LRRK2 GS is required to understand DA vulnerability. There is also an important interaction with α-synuclein that impacts DA vulnerability but the mechanism is not yet known. Aim 1 to address the ongoing controversy regarding the importance of the increased kinase activity of LRRK2 GS mutation on DA neuronal viability, LRRK2 GS kinase-dead (LRRK2 GS/DA) mice were made. These mice also provide a model to explore the non-kinase actions of LRRK2 that has been lacking from the field. Behavioral, neuroanatomical and neurochemical changes will be monitored over time. The functional interaction between LRRK2 and α-synuclein toxicity and transmission will be monitored in LRRK2 KO and transgenic lines to determine loss of LRRK2 function is protective and gain of LRRK2 function is toxic. Aim 2 will address the new observation that phosphorylated s15 is a pathogenic target of LRRK2 GS, expression of s15 and phospho-s15 will be monitored in transgenic mice and human postmortem tissue. It will be determined if phosphomimetic s15 is sufficient to elicit neurodegeneration and if phosphodeficient s15 can provide protection from LRRK2 GS neurotoxicity in global and spatially restricted models. These studies will confirm in vivo whether s15 is a pathogenic substrate of LRRK2 GS. Aim 3 will define the specific changes in mRNA translation, the transcriptome and proteome using advanced technologies in genetically engineered mice and human dopaminergic cultures. The use of both mouse and human models allows assessments in the intact brain while investigating acute changes that lead to chronic neurodegeneration in a relevant human system. The unbiased comprehensive datasets will be a valuable resource to all PD investigators. The goal of this project is to identify nodal points in the signal cascade of neurodegeneration that can provide new targets for future development of therapeutic strategies for the treatment of PD.