Characterization of the role of cyclin G-associated kinase in Parkinson disease

描述(由申请人提供)：我们对家族性帕金森病(PD)的全基因组关联研究(GWAS)首次发现细胞周期蛋白G相关激酶(GAK)基因与帕金森病风险相关。Meta-PD Gwas分析现已证明GAK基因中rs1564282 SNP在增加帕金森病风险方面具有明确的全基因组效应(至少P=4.8x10-15)。值得注意的是，这种影响似乎在家族性帕金森病中尤其强烈。最近，我们发表了GAK与1-突触核蛋白毒性相关的研究。对帕金森病患者和对照组大脑死后额叶皮质的微阵列表达分析发现，rs1564282与1-突触核蛋白表达增加显著相关，这可能与疾病的发病机制有关。此外，在帕金森病神经细胞模型中，GAK基因敲除显著增加了1-突触核蛋白的毒性。Gak通过与组织蛋白D(CTSD)的相互作用在内吞作用中发挥关键作用，CTSD是参与1-突触核蛋白降解的主要溶酶体酶。综上所述，这些研究暗示了GAK在帕金森病发病机制中的新作用。由于激酶，如GAK，是治疗干预的有吸引力的靶点，解决GAK基因区域负责的功能变异及其对GAK表达和功能的影响有望为帕金森病的研究和治疗带来重要的进展。这项应用建议识别负责的突变，并进一步评估它们在涉及的内吞途径和帕金森病发病机制中的作用。我们提出了一系列权威性的DNA测序、RNA测序、选择性剪接、基因表达和临床风险描述研究，这些研究具有重要的翻译意义。具体地说，AIM 1将通过对480例家族性帕金森病患者和96名对照的GenePD家族性帕金森病队列中的225kb重新测序来识别GAK区域的功能序列变异，在这些序列中发现了基因，从而极大地提高了成功发现功能变异的可能性。目的2建议对34个PD和29个对照大脑进行RNA测序，这些序列已经有了广泛的SNP基因分型和微阵列数据。RNA-Seq数据将用于研究GAK和相关基因在内吞途径中的编码序列和表达。RNA测序还将使我们能够检查所有与Gwas有关的PD基因(例如SNCA、MAPT、BST1等)。并对病例和对照进行转录组范围的比较。最后，目标3探讨了携带目标1和目标2中发现的变异和突变的帕金森病病例的特征，并在基因-基因和基因-环境相互作用和遗传风险概况研究中利用已识别的序列变异的基因分型。遗传风险分析研究对帕金森病的风险预测和诊断以及对帕金森病临床试验中的治疗反应的解释具有重要的翻译意义。这些帕金森病研究人员对GAK基因和区域有着独特的经验，并且已经掌握了进行这些决定性研究所需的所有样本。 公共卫生相关性：这项研究建议检测帕金森病患者和健康人的DNA序列(遗传)和基因表达(基因组)谱，以确定细胞周期蛋白G相关激酶基因在帕金森病风险中的作用。这一发现将为开发新的治疗方法以及预测和诊断这种毁灭性的神经退行性疾病提供重要的见解。

DESCRIPTION (provided by applicant): Our genome wide association study (GWAS) for familial Parkinson's disease (PD) was the first to identify the cyclin G-associated kinase (GAK) gene as associated with PD risk. Meta-PD GWAS analyses have now demonstrated an unequivocal genome-wide effect (at least P = 4.8 x 10-15) for the rs1564282 SNP in the GAK gene on increased PD risk. Notably, the effect appears to be particularly strong in familial PD. Recently, we published that GAK is associated with 1-synuclein toxicity. Microarray expression analysis of post-mortem frontal cortex from PD and control brains found a significant association of rs1564282 with increased 1- synuclein expression, which has implications for disease pathogenesis. Further, knockdown of GAK significantly increases 1-synuclein toxicity in neuronal cell models of PD. GAK plays a critical role in endocytosis through its interaction with cathepsin-D (CTSD), which is the main lysosomal enzyme involved in 1-synuclein degradation. Taken together, these studies implicate a novel role for GAK in PD pathogenesis. Since kinases, such as GAK, are attractive targets for therapeutic intervention, resolving the responsible functional variants in the GAK gene region and their effects on GAK's expression and function promise important advancements for PD research and therapeutics. This application proposes to identify the responsible mutations, and to further evaluate their role in the implicated endocytic pathway and in the pathogenesis of PD. We propose a definitive series of DNA sequencing, RNA sequencing, alternative splicing, gene expression, and clinical risk profiling studies which have important translational implications. Specifically, Aim 1 will identify functional sequence variants in the GAK region by resequencing 225 Kb in 480 familial PD cases and 96 controls from our GenePD familial PD cohort, in which the gene was identified, thus greatly enhancing the likelihood for successful discovery of functional variants. Aim 2 proposes to perform RNA-sequencing in a large well characterized series of 34 PD and 29 control brains, for which extensive SNP genotyping and microarray data are already available. The RNA-Seq data will be used to study coding sequence and expression of GAK and related genes in the endocytic pathway. RNA sequencing will also allow us to examine all the GWAS implicated PD genes (e.g. SNCA, MAPT, BST1 etc.) and to perform transcriptome-wide comparison of cases and controls. Finally, Aim 3 explores the characteristics of PD cases carrying the variants and mutations found in Aims 1 and 2 and utilizes the genotyping of identified sequence variants in gene-gene and gene- environment interaction and genetic risk profiling studies. Genetic risk profiling studies have important translational implications in risk prediction and diagnostics for PD as well as for interpretation of therapeutic response in clinical trials for PD. These PD investigators are uniquely experienced with the GAK gene and region and have already in hand all the samples needed to carry out these definitive studies. PUBLIC HEALTH RELEVANCE: This research proposes to examine the DNA sequence (genetic) and gene expression (genomic) profiles of Parkinson disease cases compared with healthy individuals to determine the role of the cyclin G-associated kinase gene in Parkinson's disease risk. The findings will provide important insight for the development of new treatments as well as in predicting and diagnosing this devastating neurodegenerative disease.