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

• 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
• 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

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。在零星帕金森氏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与零星帕金森氏病（PD）的进展有关。但是，调节LRRK2蛋白表达和功能的机制在零星PD患者的大脑中仍有待确定。在这里，我们表明，偶发性PD患者的大脑中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逆相关。随着小鼠的老化，LRRK2和miR-205的表达在大脑的多个区域也受到动态调节，并成反比，这表明miR-205在调节LRRK2表达中的潜在转录后调节作用。实际上，miR-205的过度表达抑制了在细胞系和原发性神经元培养物中LRRK2的表达，并通过过表达与PD相关的LRRK2 R1441g突变诱导的神经突生长缺陷。总而言之，我们证明了LRRK2蛋白在零星PD患者的大脑中被上调，这可能是由于miR-205的下调。我们的发现还表明，miR-205的过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标的效用已提交用于专利申请。 * miR-205数据的手稿通过人分子遗传学的修订。 2。LRRK2作为突触发生过程中PKA途径的调节剂 LRRK2积极参与综述的细胞骨架动力学（Parisiadou and Cai，2010年）。根据依从性，在神经元发育过程中，神经突生长中指出了LRRK2与肌动蛋白动力学之间的关键联系（Parisiadou等，2009）。肌动蛋白细胞骨架的脑中神经元连通性和可塑性基础的基础的树突状脊柱形成（Ethell and Pasquale，2005; Tada and Sheng，2006; Schubert and Dotti，2007年）。脊柱的形态发生包括从初始长，薄，高度柔韧的丝状卵形到更稳定的棘突的过渡，这些刺是在其尖端（脊柱头）和独特的颈部显示出特征性的球茎肿胀时被认为成熟的（Oray等，2006; Yoshihara等，2009）。脊柱形态和密度维持的重要性反映了异常的脊柱特性与脑功能障碍之间的相关性（Kasai等，2003）。 Cofilin是肌动蛋白丝传递的调节剂，严格调节基于肌动蛋白的脊柱形成动力学（Meng等，2002）。 Cofilin活性受到LIM激酶介导的磷酸化的抑制，并在残基三（S3）的高度保守丝氨酸上被弹弓诱导的去磷酸化激活（Bernstein和Bamburg，2010年）。然而，已经描述了替代性调节机制，包括蛋白激酶A（PKA）依赖性途径（Paavilainen等，2004）。 PKA介导的信号通路对于神经元的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保持不活性。与cAMP结合后，调节亚基与催化单元解离，然后起作用以磷酸化其底物（Scott，1991； Francis和Corbin，1994）。哺乳动物的PKA家族可以根据其监管亚基细分为I和II（Brandon等，1997）。 ii-beta;调节性（RII-BETA）亚基在神经元中高度表达（Ventra等，1996）（Brandon等，1998）。 PKA空间内分布在PKA信号传导中具有关键作用，因为它可能有助于信号特异性和功效（Lu等，2007； Zaccolo等，2002）。 II型PKA的亚细胞定位由A锚定蛋白（Akaps）控制（Wong and Scott，2004； Zhong等，2009）。 在这里，我们表明LRRK2是PKARII-BETA在神经元中的亚细胞分布的关键调节剂，并在LRRK2上分配了新作用，作为PKA途径的调节剂，特别是在产后两到三周，特别是在小鼠大脑突触形成的关键时期。 LRRK2-少量导致PKA活性的增加，这是Cofilin和AMPA型谷氨酸受体亚基GR1A1的磷酸化增加所证明的。 Cofilin和Gr1a1磷酸化的改变损害了脊柱形成和突触传播。总体而言，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可能是施加与OnPKA相关途径的发育开关的一部分。