Investigating the Role of LRRK2 Hyperactivity in Autophagic and Synaptic Deficits

研究 LRRK2 过度活跃在自噬和突触缺陷中的作用

• 批准号: 10549286
• 负责人: Dan Dou
• 金额: $ 3.5万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10549286
• 关键词: Affect; Age; Aging; Autophagocytosis; Axon; Biogenesis; Clinical; Complement; Complex; Data; Dementia; Dependence; Development; Disease; Disease model; Environment; Foundations; Functional disorder; Future; Genetic; Goals; Homeostasis; Human; Hyperactivity; Idiopathic Parkinson Disease; Image; Impaired cognition; Impairment; Induced pluripotent stem cell derived neurons; LRRK2 gene; Label; Link; Lysosomes; Mentors; Mitochondria; Motor; Mutation; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Organelles; PHluorin; Parkinson Disease; Pathogenesis; Pathogenicity; Pathologic; Pathway interactions; Patients; Peptide Hydrolases; Phospho-Specific Antibodies; Phosphorylation; Phosphotransferases; Physicians; Physiological; Play; Population; Prevalence; Process; Proteins; Regulation; Repression; Research; Rodent; Role; Scientist; Severities; Synapses; Synaptic Transmission; Synaptic Vesicles; Synaptophysin; Testing; Therapeutic; Training; Transmembrane Transport; Travel; Vesicle; Work; Writing; age related; burden of illness; causal variant; cognitive impairment in Parkinson' s; dominant genetic mutation; experimental study; fluorescence imaging; healthy aging; human disease; improved; in vivo; induced pluripotent stem cell; inhibitor; neuronal cell body; non-motor symptom; presynaptic; rab GTP-Binding Proteins; recruit; retrograde transport; therapeutically effective; vesicle transport; vesicular release

PROJECT SUMMARY Parkinson’s disease (PD) is the second-most prevalent neurodegenerative disease and the fastest- growing. PD increases in both prevalence and severity with age, and the disease burden is projected to dramatically increase in coming decades as the population ages. Current therapies do not address the underlying neurodegeneration, and development of improved therapeutics is hindered by poor understanding of the pathogenesis. Changes in homeostatic regulation of both autophagy and synaptic transmission have been implicated in not only PD but also physiological aging. Leucine-rich repeat kinase 2 (LRRK2) is a leading candidate for a nexus bridging autophagy and PD pathogenesis. Mutations in LRRK2 are the most common genetic cause of PD, and increased LRRK2 kinase activity has also been linked to idiopathic PD. There is accumulating evidence that multiple PD-causative pathways may converge on disrupting autophagy through mechanisms dependent on LRRK2 activity. Recently, LRRK2 was shown to phosphorylate a subset of Rab GTPases, providing an important opportunity to make advances in the understanding of mechanisms downstream of LRRK2. The goal of this proposal is to elucidate the role that LRRK2 may play in autophagy and synaptic homeostasis, in the context of pathogenic mutations associated with PD. Our preliminary data suggests that the most common pathogenic mutation in LRRK2, p.G2019S, disrupts autophagic vesicle (AV) transport in the axons of rodent and iPSC-derived human neurons. We hypothesize that multiple pathogenic PD mutations disrupt autophagy and synaptic vesicle precursor (SVP) transport through mechanisms dependent on increased LRRK2 kinase activity. To test this hypothesis, we will exploit iPSC-derived neurons as a human disease model. Experiments proposed in Aim 1a will explore whether LRRK2-p.G2019S causes deficits in AV cargo degradation, as a potential mechanism of neurodegeneration. In Aim 1b, we will determine whether the VPS35-p.D620N mutation, a different pathogenic PD mutation that increases LRRK2 activity, causes similar autophagy deficits as LRRK2-p.G2019S. Finally, in Aim 2, we will explore whether SVP transport from the soma to the axon is impaired by mutations causing LRRK2 hyperactivity, with potential ramifications for synaptic homeostasis. Collectively, we expect that these Aims will help elucidate mechanisms by which multiple causes of PD can converge on a LRRK2-dependent pathway to disrupt neuronal autophagy and synaptic homeostasis. Additionally, this work may provide a foundation for future work to shed light on pathways by which physiological aging alters these processes in the absence of diseases of aging. To complement this research plan, a comprehensive training plan will help the trainee meet specific Research Goals, Professional Goals (including writing, mentoring, and presentation), and Clinical Goals (tying together an integrated physician-scientist training pathway), all in a supportive, collaborative scientific environment.

项目概要 帕金森病 (PD) 是第二常见的神经退行性疾病，也是发病最快的疾病 生长。 PD 的患病率和严重程度随着年龄的增长而增加，预计疾病负担将增加 随着人口老龄化，未来几十年将急剧增加。目前的治疗方法并不能解决 潜在的神经退行性变，并且由于了解不足而阻碍了改进疗法的发展 的发病机制。自噬和突触传递的稳态调节的变化 不仅与PD有关，而且与生理衰老有关。富含亮氨酸重复激酶 2 (LRRK2) 是一种领先的 桥接自噬和 PD 发病机制的候选者。 LRRK2 突变是最常见的 PD 的遗传原因，LRRK2 激酶活性增加也与特发性 PD 相关。有 越来越多的证据表明，多种 PD 致病途径可能会通过以下方式集中破坏自噬： 机制依赖于 LRRK2 活性。最近，LRRK2 被证明可以磷酸化 Rab 的一个子集 GTPases，为增进对机制的理解提供了重要机会 LRRK2 的下游。该提案的目的是阐明 LRRK2 在自噬中可能发挥的作用 和突触稳态，在与 PD 相关的致病突变的背景下。我们的初步数据 表明 LRRK2 最常见的致病性突变 p.G2019S 会破坏自噬囊泡 (AV) 啮齿动物和 iPSC 衍生的人类神经元轴突中的运输。我们假设多种致病性 PD 突变通过机制破坏自噬和突触小泡前体 (SVP) 运输 依赖于 LRRK2 激酶活性的增加。为了验证这一假设，我们将利用 iPSC 衍生的神经元 作为人类疾病模型。目标 1a 中提出的实验将探讨 LRRK2-p.G2019S 是否会导致 AV 货物降解的缺陷，作为神经退行性变的潜在机制。在目标 1b 中，我们将确定 VPS35-p.D620N 突变是否是一种增加 LRRK2 活性的不同致病性 PD 突变， 引起与 LRRK2-p.G2019S 类似的自噬缺陷。最后，在目标 2 中，我们将探讨 SVP 是否 从体细胞到轴突的运输受到导致 LRRK2 过度活跃的突变的损害，有可能 突触稳态的影响。总的来说，我们期望这些目标将有助于阐明机制 PD 的多种原因可以汇聚到 LRRK2 依赖性途径上，从而破坏神经元自噬 和突触稳态。此外，这项工作可能为未来的工作奠定基础，以阐明 在没有衰老疾病的情况下，生理衰老改变这些过程的途径。到 作为该研究计划的补充，全面的培训计划将帮助受训者满足特定的研究 目标、专业目标（包括写作、指导和演示）和临床目标（将这些目标结合在一起） 综合的医师-科学家培训途径），所有这些都在支持性、协作性的科学环境中进行。