Function and Pathogenic Mechanism of LRRK2 in Parkinson's Disease

LRRK2在帕金森病中的功能及发病机制

基本信息

批准号：
8552520
负责人：
Huaibin Cai
金额：
$ 82.7万
依托单位：
NATIONAL INSTITUTE ON AGING
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/8552520
关键词：
A kinase anchoring protein Actins Affect Behavioral Binding Biological Markers Bradykinesia Brain Brain region Catalytic Domain Cell Line Cellular biology Characteristics Cyclic AMP Cyclic AMP-Dependent Protein Kinases Cytoskeleton Data Defect Dendritic Spines Development Down-Regulation Family Filopodia Functional disorder Gene Expression Gene Mutation Genome Glutamate Receptor Glycine Head Holoenzymes Human Image Inherited Knowledge LIM Domain Kinase 1 Legal patent Life Link Maintenance Manuscripts Mediating MicroRNAs Microfilaments Midbrain structure Molecular Molecular Genetics Morphogenesis Morphology Movement Disorders Mus Mutant Strains Mice Mutation Neck Nerve Degeneration Neurites Neurodegenerative Disorders Neurons PARK8 gene Parkinson Disease Participant Pathway interactions Patients Phosphorylation Phosphotransferases Positioning Attribute Process Property Proteins Proteomics Rest Rest Tremor Role Serine Signal Pathway Signal Transduction Specificity Synaptic Transmission Transgenic Mice Transgenic Organisms Up-Regulation Vertebral column Vesicle aged base cofilin critical period density dopaminergic neuron flexibility leucine-rich repeat kinase 2 mutant neurite growth neuron development new therapeutic target postnatal protein expression protein function protein phosphatase inhibitor-2 research study synaptogenesis therapeutic target trafficking

项目摘要

1. Upregulation of Leucine-rich Repeat Kinase 2 Expression in Sporadic Parkinson Disease involves Specific MicroRNA. LRRK2 has been implicated in the progression of sporadic Parkinson Disease (PD). However, the mechanisms regulating LRRK2 protein expression and function in the brains of patients with sporadic PD remain to be determined. Here we show that the expression of LRRK2 protein is significantly increased in the brains of patients with sporadic PD. Moreover, we found a significant inverse-correlation between the expression of LRRK2 and microRNA-205 (miR-205) in the PD brains. The expression of LRRK2 and miR-205 were also dynamically regulated and inversely correlated in multiple regions of the brain as mice aged, suggesting a potential post-transcriptional regulatory role of miR-205 in modulating LRRK2 expression. Indeed over-expression of miR-205 suppressed the expression of LRRK2 in both cell lines and primary neuronal cultures, as well as rescued the neurite growth defects induced by over-expressing the PD-related LRRK2 R1441G mutation. In summary, we demonstrate that LRRK2 protein is up-regulated in the brains of patients with sporadic PD possibly due to down-regulation of miR-205. Our findings also suggest that over-expression of miR-205 may help to suppress the pathogenic elevation of LRRK2 in the brains of patients with PD. * The utility of miR-205 as a biomarker and therapeutic target has been submitted for patent application. * The manuscript of miR-205 data is under revision by Human Molecular Genetics. 2. LRRK2 as a Modulator of PKA Pathway during Synaptogenesis LRRK2 is actively involved in cytoskeletal dynamics for review (Parisiadou and Cai, 2010). In accordance, a critical connection between LRRK2 and actin dynamics is indicated in the neurite outgrowth during neuron development (Parisiadou et al., 2009). Dendritic spine formation that underlies the basis for neuron connectivity and plasticity in the brain is also critically determined by the actin cytoskeleton (Ethell and Pasquale, 2005; Tada and Sheng, 2006; Schubert and Dotti, 2007). Spine morphogenesis include the transition from initial long, thin, highly flexible filopodia to more stable spines that are considered mature when show characteristic bulbous enlargements at their tips (spine head) and distinct neck (Oray et al., 2006; Yoshihara et al., 2009). The importance of the maintenance of spine morphology and density is reflected by the correlation between abnormal spine properties and brain dysfunction (Kasai et al., 2003). Cofilin, a modulator of actin filament turnover, critically regulates the actin-based dynamics of spine formation (Meng et al., 2002). Cofilin activities are inhibited by LIM kinase-mediated phosphorylation and activated by Slingshot-induced de-phosphorylation on a highly conserved Serine at residue three (S3) (Bernstein and Bamburg, 2010). However, alternative regulatory mechanisms have been described, including a protein kinase A (PKA)-dependent pathway (Paavilainen et al., 2004). PKA-mediated signaling pathways are critical for neuron development and function (Frey et al., 1993; Greenberg et al., 1987; Choi et al., 2002). PKA is a holoenzyme that consists of two regulatory and two catalytic subunits. At resting state, each regulatory subunit binds to a catalytic unit and keeps it inactive. Upon binding with cAMP, the regulatory subunit dissociates with catalytic unit, which then acts to phosphorylate its substrates (Scott, 1991; Francis and Corbin, 1994). The mammalian PKA family could be subdivided into types I and II based on their regulatory subunits (Brandon et al., 1997). The II-beta; regulatory (RII-beta) subunit is highly expressed in neurons (Ventra et al., 1996) (Brandon et al., 1998). PKA spatial intracellular distribution possesses a critical role in PKA signaling since it might contribute to signaling specificity and efficacy (Lu et al., 2007; Zaccolo et al., 2002). The subcellular localization of type II PKA is controlled by the A kinase anchoring proteins (AKAPs) (Wong and Scott, 2004; Zhong et al., 2009). Here we show that LRRK2 is a critical regulator of PKARII-beta's subcellular distribution in neurons, and assign a new role on LRRK2 as a modulator of PKA pathway particularly around postnatal two to three weeks, a critical period for the synapse formation in the mouse brain. LRRK2-absence resulted to increased PKA activity as evidenced by increased phosphorylation of cofilin and AMPA type glutamate receptor subunit GR1A1. The alterations of cofilin and GR1A1 phosphorylation impaired the spine formation and synaptic transmission. Overall, our data reveal a new regulatory role of LRRK2 during synaptogenesis, in which LRRK2 might function as part of the developmental switch imposed onPKA-related pathways.

1. 散发性帕金森病中富含亮氨酸的重复激酶 2 表达的上调涉及特定的 MicroRNA。 LRRK2 与散发性帕金森病 (PD) 的进展有关。然而，散发性帕金森病患者大脑中调节 LRRK2 蛋白表达和功能的机制仍有待确定。在这里，我们发现散发性帕金森病患者大脑中 LRRK2 蛋白的表达显着增加。此外，我们发现 PD 大脑中 LRRK2 和 microRNA-205 (miR-205) 的表达之间存在显着的负相关。随着小鼠年龄的增长，LRRK2 和 miR-205 的表达也在大脑的多个区域中受到动态调节并呈负相关，这表明 miR-205 在调节 LRRK2 表达中具有潜在的转录后调节作用。事实上，miR-205 的过度表达抑制了细胞系和原代神经元培养物中 LRRK2 的表达，并挽救了因过度表达 PD 相关 LRRK2 R1441G 突变而引起的神经突生长缺陷。总之，我们证明散发性 PD 患者大脑中 LRRK2 蛋白上调，可能是由于 miR-205 下调所致。我们的研究结果还表明，miR-205 的过度表达可能有助于抑制 PD 患者大脑中 LRRK2 的致病性升高。 * miR-205 作为生物标志物和治疗靶点的用途已提交专利申请。 * miR-205 数据的手稿正在由人类分子遗传学修订。 2. LRRK2 作为突触发生过程中 PKA 通路的调节剂 LRRK2 积极参与细胞骨架动力学审查（Parisiadou 和 Cai，2010）。因此，神经元发育过程中神经突的生长表明了 LRRK2 和肌动蛋白动力学之间的关键联系（Parisiadou 等人，2009）。作为大脑神经元连接和可塑性基础的树突棘形成也主要由肌动蛋白细胞骨架决定（Ethell 和 Pasquale，2005；Tada 和 Shen，2006；Schubert 和 Dotti，2007）。脊柱形态发生包括从最初的长、薄、高度灵活的丝状伪足到更稳定的脊柱的转变，当在其尖端（脊柱头）和明显的颈部显示出特征性的球状增大时，脊柱被认为是成熟的（Oray等人，2006年；Yoshihara等人， 2009）。维持脊柱形态和密度的重要性反映在异常脊柱特性与脑功能障碍之间的相关性（Kasai et al., 2003）。 Cofilin 是肌动蛋白丝周转的调节剂，它严格调节基于肌动蛋白的脊柱形成动力学（Meng 等，2002）。 Cofilin 活性被 LIM 激酶介导的磷酸化抑制，并被 Slingshot 诱导的残基 3 (S3) 高度保守的丝氨酸去磷酸化激活 (Bernstein 和 Bamburg, 2010)。然而，已经描述了替代的调节机制，包括蛋白激酶 A (PKA) 依赖性途径 (Paavilainen 等人，2004)。 PKA 介导的信号通路对于神经元发育和功能至关重要（Frey 等，1993；Greenberg 等，1987；Choi 等，2002）。 PKA 是一种全酶，由两个调节亚基和两个催化亚基组成。在静止状态下，每个调节亚基与催化单元结合并使其保持非活性。与 cAMP 结合后，调节亚基与催化单元解离，然后催化单元磷酸化其底物（Scott，1991；Francis 和 Corbin，1994）。哺乳动物 PKA 家族根据其调节亚基可分为 I 型和 II 型（Brandon 等，1997）。 II-β；调节 (RII-β) 亚基在神经元中高度表达 (Ventra 等人, 1996) (Brandon 等人, 1998)。 PKA 细胞内空间分布在 PKA 信号传导中起着至关重要的作用，因为它可能有助于信号传导的特异性和功效（Lu 等人，2007；Zaccolo 等人，2002）。 II 型 PKA 的亚细胞定位由 A 激酶锚定蛋白 (AKAP) 控制（Wong 和 Scott，2004；Zhong 等，2009）。在这里，我们证明 LRRK2 是 PKARII-β 在神经元中亚细胞分布的关键调节因子，并赋予 LRRK2 作为 PKA 通路调节剂的新作用，特别是在出生后两到三周，这是小鼠大脑中突触形成的关键时期。 LRRK2 缺失导致 PKA 活性增加，这一点可以通过 cofilin 和 AMPA 型谷氨酸受体亚基 GR1A1 磷酸化的增加来证明。 cofilin 和 GR1A1 磷酸化的改变损害了脊柱的形成和突触传递。总的来说，我们的数据揭示了 LRRK2 在突触发生过程中的新调节作用，其中 LRRK2 可能作为 PKA 相关途径发育开关的一部分发挥作用。