Regulation of Leucine Rich Repeat Kinase 2 (LRRK2)

富含亮氨酸重复激酶 2 (LRRK2) 的调节

• 批准号: 8191191
• 负责人: Matthew J Lavoie
• 金额: $ 26.76万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8191191
• 关键词: Accounting; Biochemical; Biological Assay; Biology; Cell Line; Cells; Complex; Cytoplasm; Data; Dimerization; Disease; Engineering; Environment; Event; Foundations; Functional disorder; Future; Genes; Genetic; Genetic Determinism; Goals; Guanosine Triphosphate Phosphohydrolases; In Situ; Investigation; Life; Link; Luciferases; Membrane; Missense Mutation; Molecular Conformation; Mutation; Nature; Neurodegenerative Disorders; Parkinson Disease; Pathogenesis; Pathway interactions; Patients; Penetrance; Phosphotransferases; Physiological; Play; Population; Process; Property; Protein Conformation; Protein Fragment; Protein-Serine-Threonine Kinases; Proteins; Regulation; Reporter; Reporting; Resources; Rest; Role; Screening procedure; Signal Transduction; Small Interfering RNA; Stress; System; Time; Validation; Viral; Work; base; dimer; high throughput screening; improved; in vitro Assay; inhibitor/antagonist; innovation; insight; interest; leucine-rich repeat kinase 2; mutant; novel; small molecule; stressor; tool

DESCRIPTION (provided by applicant): Missense mutations within the multi-domain kinase, LRRK2, are the most common cause of familial Parkinson's disease (PD), accounting for up to 40% of cases in some populations. LRRK2 mutations also appear in an unprecedented number of sporadic PD patients, implicating this protein in all forms of PD. While the genetic links between LRRK2 dysfunction and PD pathogenesis are clear, less is known about the physiological activity of LRRK2 or how it is regulated. The weak penetrance of LRRK2 disease mutations, however, does suggest that other genes may potently influence LRRK2 function and play a role in its regulation. Recent data indicate that like other kinases, LRRK2 can form a functional dimer within the cell. We've shown that the LRRK2 dimer is several-fold more active than its monomeric counterpart. In addition, the majority (~75%) of total LRRK2 is found within the cytoplasm at rest, while the membrane fraction is selectively enriched in the LRRK2 dimer. This is highly reminiscent of other proteins involved in intracellular signaling, where various kinases and GTPases shuttle to and from the membrane, and can be activated through dimerization. Therefore, we hypothesize that these newly discovered biochemical properties of LRRK2 are consistent with canonical mechanisms used throughout nature to regulate kinase activity and intracellular signaling. However, the precise importance of these processes to LRRK2 are not known. Our long-term goal is to uncover the biochemical nature of PD-linked LRRK2 mutants. However, this has remained elusive, partly due to our incomplete understanding of LRRK2 biology. At this time, a broader understanding of the physiological function and regulation of wild-type LRRK2 is critically needed. The goal of this application is to optimize a newly developed, quantitative assay of LRRK2 dimerization, and use this tool to uncover critical aspects of LRRK2 regulation. Specifically, we will 1) develop a protein-fragment complementation assay of LRRK2 dimerization, and 2) identify cellular and genetic determinants of LRRK2 dimerization in the intact cell. This work will not only inform us as to the pathways involved in intracellular LRRK2 signaling, but also provide novel tools to examine the underlying mechanisms of several pathogenic LRRK2 mutations in future studies. PUBLIC HEALTH RELEVANCE: Mutations in the LRRK2 kinase are the most common genetic cause of Parkinson's disease. The goal of this application is to optimize a novel, high-throughput assay of LRRK2 biology to uncover how cells regulate LRRK2 function. Through this new resource, and an improved understanding of the pathways controlling LRRK2 activity, we hope to gain valuable insights into the physiological and pathological functions of LRRK2.

描述（由申请人提供）：多域激酶 LRRK2 内的错义突变是家族性帕金森病 (PD) 的最常见原因，在某些人群中占病例的 40%。 LRRK2 突变也出现在数量空前的散发性 PD 患者中，表明该蛋白与所有形式的 PD 有关。虽然 LRRK2 功能障碍与 PD 发病机制之间的遗传联系很清楚，但人们对 LRRK2 的生理活性或其调节方式知之甚少。然而，LRRK2 疾病突变的弱外显率确实表明其他基因可能会有效影响 LRRK2 功能并在其调节中发挥作用。最近的数据表明，与其他激酶一样，LRRK2 可以在细胞内形成功能性二聚体。我们已经证明 LRRK2 二聚体的活性是其单体对应物的数倍。此外，总 LRRK2 的大部分 (~75%) 存在于静止的细胞质中，而膜部分则选择性富集 LRRK2 二聚体。这很容易让人想起参与细胞内信号传导的其他蛋白质，其中各种激酶和 GTP 酶在细胞膜之间穿梭，并且可以通过二聚化来激活。因此，我们假设 LRRK2 这些新发现的生化特性与自然界中用于调节激酶活性和细胞内信号传导的规范机制一致。然而，这些过程对 LRRK2 的确切重要性尚不清楚。我们的长期目标是揭示 PD 连锁 LRRK2 突变体的生化性质。然而，这一点仍然难以捉摸，部分原因是我们对 LRRK2 生物学的了解不完全。此时，迫切需要对野生型 LRRK2 的生理功能和调控有更广泛的了解。该应用的目标是优化新开发的 LRRK2 二聚化定量测定，并使用该工具揭示 LRRK2 调控的关键方面。具体来说，我们将 1) 开发 LRRK2 二聚化的蛋白质片段互补测定，2) 鉴定完整细胞中 LRRK2 二聚化的细胞和遗传决定因素。这项工作不仅让我们了解细胞内 LRRK2 信号传导的途径，而且还为未来研究中检查几种致病性 LRRK2 突变的潜在机制提供了新的工具。 公共卫生相关性：LRRK2 激酶突变是帕金森病最常见的遗传原因。此应用的目标是优化 LRRK2 生物学的新型高通量测定，以揭示细胞如何调节 LRRK2 功能。通过这一新资源，以及对控制 LRRK2 活性的途径的深入了解，我们希望获得对 LRRK2 生理和病理功能的宝贵见解。