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: 8335977
• 负责人: Huaibin Cai
• 金额: $ 67.24万
• 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/8335977
• 关键词: Acute; Address; Affect; Alleles; Autophagocytosis; Behavioral; Biological Markers; Biota; Bradykinesia; Brain; Brain region; Cell Line; Cells; Cellular biology; Colitis; Complex; Crohn' s disease; Cytoplasm; Data; Databases; Defect; Deposition; Development; Down-Regulation; Environmental Risk Factor; Equilibrium; Event; Future; Gastrointestinal tract structure; Gene Expression; Gene Mutation; Genetic; Genome; Glycine; Human; Image; Immune; Immune response; Immune system; Immunology; Inflammatory; Inflammatory Bowel Diseases; Inherited; Knockout Mice; Knowledge; Legal patent; Life; Link; Lipopolysaccharides; Location; Lymphoid Tissue; Manuscripts; Mediator of activation protein; MicroRNAs; Midbrain structure; Modeling; Molecular; Movement Disorders; Mus; Mutant Strains Mice; Mutation; Myeloid Cells; NFAT Pathway; Nature; Nerve Degeneration; Neurodegenerative Disorders; Neurons; PARK8 gene; Parkinson Disease; Participant; Pathogenesis; Patients; Phosphorylation; Phosphotransferases; Positioning Attribute; Predisposition; Process; Property; Proteins; Proteomics; RNA Interference; Regulatory T-Lymphocyte; Research; Rest Tremor; Risk; Role; Serine; Severe Combined Immunodeficiency; Severities; Signal Transduction; Sodium Dextran Sulfate; Susceptibility Gene; Transgenic Mice; Transgenic Organisms; Ulcerative Colitis; Up-Regulation; Vesicle; aged; dopaminergic neuron; genome wide association study; insight; leucine-rich repeat kinase 2; mutant; neurite growth; new therapeutic target; novel; pathogenic bacteria; protein expression; protein function; research study; response; therapeutic target; trafficking

1. LRRK2 is a novel regulator of NFAT that modulates the severity of inflammatory bowel disease. Inflammatory bowel disease (IBD), which is generally thought to develop from a dysregulated immune response to the gut luminal biota, includes two major forms: Crohns disease (CD) and ulcerative colitis. Both genetic and environmental factors contribute to the development of CD1. Genome-wide association studies (GWAS) for Crohns disease have identified over 40 susceptibility loci2-4. The large number of the susceptibility genes likely reflects the complexity of the inflammatory process taking place along the gastrointestinal tract in IBD. The lymphoid tissue within the gastrointestinal tract constantly encounters normal commensal microbiota as well as potentially pathogenic bacteria; hence it is critical in this location to maintain a delicate balance between immune responsiveness and tolerance. Research on the susceptibility genes identified by GWAS has exemplified this concept, and yielded important insights into the pathogenesis of IBD. For example, the regulatory function of Treg cells, the involvement of autophagy, and IL23/IL23R signaling events have been implicated in IBD5. Despite this progress, the majority of susceptibility genes have no known mechanism to explain their involvement with CD. We investigated the role of LRRK2 in IBD and the underlying molecular mechanism. Various experimental murine models have been established to study the roles of innate and adaptive immune systems in the pathogenesis of IBD. Given the predominant expression of LRRK2 in innate immune cells but not in T cells21, dextran sulfate sodium (DSS) induced colitis was used in our study. Acute colitis induced by DSS depends on the innate immune system, because it occurs in severe combined immunodeficiency (SCID) mice which lack B and T lymphocytes22. Here we show that LRRK2 deficiency exacerbated colitis in mice treated with DSS. We searched for potential mediators of the effect using data from a RNAi screen in D. Melanogaster and found that LRRK2 was an interactor with NFAT. Examining this connection biochemically, we found that LRRK2 restrained NFAT1 in the cytoplasm within the NRON complex and inhibited NFAT1 activation. We also determined that LRRK2 did not alter NFAT1 phosphorylation but instead potently modulated cytoplasmic retention and the interaction between NFAT1 and the NRON complex in response to selected inducers such as lipopolysaccharide (LPS). Finally, a LRRK2 risk allele associated with CD in humans resulted in subtly reduced LRRK2 protein levels and delicately enhanced NFAT activity consistent with its role as part of a multifactorial susceptibility to human IBD. Collectively, our data demonstrate that LRRK2 negatively regulates NFAT-driven innate immune responses in myeloid cells and that lower levels of LRRK2 in humans provides a molecular insight into how this protein can contribute to an increased risk of CD. Future studies will address the involvement of LRRK2/NFAT pathway in the potential microgliosis and neurodegeneration in the brain. * The manuscript of the above data has been accepted by Nature Immunology. 2. 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 the final revision by Brain. 3. Deposition of LRRK2 WT and G2019S transgenic and LRRK2 knockout mice in the JAX for public access. JAX Mice Database - 012441 C57BL/6J-Tg(tetO-LRRK2*G2019S)E3Cai/J JAX Mice Database - 012449 TC57BL/6J-Tg(teto-LRRK2)C7874Cai/J JAX Mice Database - 012453 B6.129X1(FVB)-Lrrk2tm1.1Cai/J

1。LRRK2是NFAT的新型调节剂，可调节炎症性肠病的严重程度。 炎症性肠病（IBD）通常被认为是由于对肠道腔生物植物的免疫反应而发展的，包括两种主要形式：克罗恩斯病（CD）和溃疡性结肠炎。遗传和环境因素都有助于CD1的发展。针对克罗恩斯疾病的全基因组关联研究（GWAS）已鉴定出40多个敏感性基因座。大量的敏感性基因可能反映了沿着IBD胃肠道发生的炎症过程的复杂性。胃肠道内的淋巴组织不断遇到正常的共生微生物群以及潜在的致病细菌。因此，在这个位置至关重要的是要在免疫反应和耐受性之间保持微妙的平衡。 GWAS鉴定的易感基因的研究已经举例说明了这一概念，并对IBD的发病机理产生了重要的见解。例如，Treg细胞的调节功能，自噬的参与和IL23/IL23R信号事件已与IBD5有关。尽管取得了这种进步，但大多数易感基因都没有已知的机制来解释其参与CD。我们研究了LRRK2在IBD和潜在的分子机制中的作用。已经建立了各种实验性鼠模型，以研究先天和适应性免疫系统在IBD发病机理中的作用。鉴于LRRK2在先天免疫细胞中的主要表达，而在T细胞中不具有lrRK2，因此在我们的研究中使用了硫酸葡萄糖钠（DSS）诱导的结肠炎。 DSS诱导的急性结肠炎取决于先天免疫系统，因为它发生在缺乏B和T淋巴细胞22的严重组合免疫缺陷（SCID）小鼠中。在这里，我们表明LRRK2缺乏加剧了用DSS治疗的小鼠的结肠炎。我们使用D. melanogaster中的RNAi屏幕的数据搜索了效果的潜在介体，发现LRRK2是与NFAT的相互作用。通过生化研究这种联系，我们发现LRRK2在NRON复合物中的细胞质中限制了NFAT1并抑制了NFAT1激活。我们还确定LRRK2没有改变NFAT1磷酸化，而是有效调节细胞质保留率以及NFAT1与NRON复合物之间的相互作用，以响应选定 诱导剂，例如脂多糖（LPS）。最后，与人类CD相关的LRRK2风险等位基因导致LRRK2蛋白水平略有降低，并精致增强了NFAT活性，与其作为对人IBD多因素敏感性的一部分一致的作用一致。总体而言，我们的数据表明，LRRK2负调节髓样细胞中NFAT驱动的先天免疫反应，并且人类中LRRK2的较低水平为该蛋白如何促进CD的风险增加的分子见解。未来的研究将涉及LRRK2/NFAT途径在大脑中潜在的小胶质细胞和神经变性中的参与。 *上述数据的手稿已被自然免疫学接受。 2。富含亮氨酸的重复激酶2在零星帕金森氏病中的表达涉及特定的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数据的手稿在大脑的最终修订下。 3。在JAX中，LRRK2 WT和G2019S转基因和LRRK2敲除小鼠的沉积。 JAX小鼠数据库-012441 C57BL/6J-TG（TETO-LRK2*G2019S）E3CAI/J JAX小鼠数据库-012449 TC57BL/6J-TG（TETO-LRK2）C7874CAI/J JAX小鼠数据库-012453 B6.129x1（FVB）-LRRK2TM1.1CAI/J