Reversible and Temporally Inducible LRRK2 Knockout Mice

可逆且暂时诱导的 LRRK2 敲除小鼠

• 批准号: 7229920
• 负责人: Ted M. Dawson
• 金额: $ 21.47万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-02-01 至 2008-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7229920
• 关键词: Aging; Animals; Biochemical; Brain; Cell Death; Cells; Cessation of life; Class; Deposition; Development; Dissection; Dopaminergic Cell; Evaluation; Exhibits; Experimental Models; Family; Gene Targeting; Generations; Genes; Guanosine Triphosphate Phosphohydrolases; Human; Inherited; Injection of therapeutic agent; Interruption; Knock-out; Knockout Mice; LRRK2 gene; Lead; MAP Kinase Kinase Kinase; Maintenance; Methods; Missense Mutation; Modeling; Molecular; Monkeys; Mus; Mutation; Nerve Degeneration; Neurons; Neurotoxins; Numbers; Parkinson Disease; Pathogenesis; Pathology; Pathway interactions; Pharmaceutical Preparations; Phosphotransferases; Principal Investigator; Process; Protein Family; Protein Kinase; Proteins; Role; Signal Transduction; System; Time; Tissues; Toxic Environmental Substances; Toxic effect; Transgenic Mice; WD Repeat; alpha synuclein; dopamine system; dopaminergic neuron; drinking water; inhibitor/antagonist; innovation; insight; leucine-rich repeat kinase 2; novel; novel therapeutics; null mutation; postnatal; prevent; programs; research study; synuclein; tau Proteins; therapeutic target

DESCRIPTION (provided by applicant): Mutations in the LRRK2/Dardarin gene are a common cause of autosomal dominant Parkinson's disease (PD). Several dominantly inherited missense mutations have been identified in a number of families that exhibit a broad spectrum of neuropathological features, including deposition of alpha-synuclein and tau proteins. LRRK2 (leucine-rich repeat kinase 2) encodes a large, multifunctional protein. It belongs to the ROCO protein family and includes a protein kinase domain of the MAPKKK class and several other major functional domains including Ras/GTPase and WD40 domains. It is difficult at this juncture to fully appreciate how mutations in the LRRK2 gene cause PD, as its function is largely unknown. We propose to generate and characterize LRRK2 knockout mice to gain a better understanding of LRRK2 function and to determine whether the absence of LRRK2 function is the cause of PD due to LRRK2 mutations. We plan to utilize the recent technological advances in the generation of inhibitor sensitizing mutations to introduce a mutation into the LRRK2 kinase domain that will allow the specific inhibition of the LRRK2 signaling cascade. Introduction of the drug in drinking water or by injection will permit the specific and reversible inhibition of LRRK2 kinase activity at any time during development or aging. Such an innovative approach provides an extremely valuable method to dissect the role of LRRK2 signaling in PD. Moreover, it will enable a molecular dissection of the role of LRRK2 kinase activity in maintaining a functional nigrostriatal dopamine system during development, early postnatal development and within the mature and diseased brain. Our targeting strategy will also enable us to create a null mutation in LRRK2 as well as tissue specific deletion of LRRK2 using the Cre-Lox system. Accordingly experiments are proposed to further characterize the role of LRRK2 in the pathogenesis of PD. In Specific Aim #1 we will develop and characterize a reversible and temporally inducible LRRK2 knockout mouse using a novel method of reversible and temporally specific interruption of kinase activity in cells and animals. In Specific Aim #2 we will we will evaluate the sensitivity of LRRK2 gene targeted mice to environmental toxins including MPTP-induced dopaminergic cell death. In Specific Aim #3 we will determine whether LRRK.2, a-synuclein and/or tau participate in a common pathogenic pathway by crossing LRRK2 gene-targeted mice with a-synuclein and tau transgenic mice. Development and characterization of a reversible and temporally inducible LRRK2 knockout mouse, will increase our understanding of the relationship of LRRK2 kinase activity in the pathogenesis of PD. Moreover, it may provide insight into the molecular mechanisms by which mutations in this gene induce neuronal damage and may provide novel therapeutics targets to prevent the toxic effects of this familial associated gene in the degenerative process of PD.

描述（由申请人提供）：LRRK2/Dardarin 基因突变是常染色体显性帕金森病 (PD) 的常见原因。在许多家族中已经发现了几种显性遗传的错义突变，这些突变表现出广泛的神经病理学特征，包括 α-突触核蛋白和 tau 蛋白的沉积。 LRRK2（富含亮氨酸重复激酶 2）编码一种大型多功能蛋白质。它属于 ROCO 蛋白家族，包括 MAPKKK 类蛋白激酶结构域和几个其他主要功能结构域，包括 Ras/GTPase 和 WD40 结构域。目前还很难完全理解 LRRK2 基因突变如何导致帕金森病，因为其功能很大程度上未知。我们建议生成并表征 LRRK2 敲除小鼠，以更好地了解 LRRK2 功能，并确定 LRRK2 功能的缺失是否是 LRRK2 突变引起的 PD 的原因。我们计划利用抑制剂致敏突变生成方面的最新技术进展，将突变引入 LRRK2 激酶结构域，从而特异性抑制 LRRK2 信号级联。在饮用水或注射中引入药物将允许在发育或衰老过程中的任何时间特异性且可逆地抑制 LRRK2 激酶活性。这种创新方法提供了一种极其有价值的方法来剖析 LRRK2 信号传导在 PD 中的作用。此外，它将能够对 LRRK2 激酶活性在发育、产后早期发育以及成熟和患病大脑内维持功能性黑质纹状体多巴胺系统中的作用进行分子剖析。我们的靶向策略还将使我们能够使用 Cre-Lox 系统在 LRRK2 中创建无效突变以及 LRRK2 的组织特异性删除。因此，建议进行实验以进一步表征 LRRK2 在 PD 发病机制中的作用。在特定目标#1中，我们将使用一种可逆且暂时特异性中断细胞和动物激酶活性的新方法来开发并表征可逆且可暂时诱导的 LRRK2 敲除小鼠。在具体目标#2 中，我们将评估 LRRK2 基因靶向小鼠对环境毒素（包括 MPTP 诱导的多巴胺能细胞死亡）的敏感性。在具体目标 #3 中，我们将通过将 LRRK2 基因靶向小鼠与 a-突触核蛋白和 tau 转基因小鼠杂交来确定 LRRK.2、a-突触核蛋白和/或 tau 是否参与共同的致病途径。可逆且可暂时诱导的 LRRK2 敲除小鼠的开发和表征将增加我们对 LRRK2 激酶活性在 PD 发病机制中关系的理解。此外，它可以深入了解该基因突变引起神经元损伤的分子机制，并可能提供新的治疗靶点，以防止该家族相关基因在帕金森病退行性过程中产生毒性作用。