Molecular Dissection of Synucleinopathies Using Yeast, Rodent and Human iPS Cells

描述（由申请人提供）：突触核蛋白病，包括帕金森病和路易体痴呆，是与α-突触核蛋白的神经元聚集相关的常见衰老依赖性神经退行性疾病。体内大鼠突触核蛋白病模型描绘了明显神经元变性之前的早期关键病理变化。这些可能代表可能导致疾病进展的关键病理生理学变化，而不会受到细胞死亡相关事件的混淆。我假设这些在前退行性阶段受到干扰的通路是突触核蛋白病发病机制的重要贡献者，并将产生疾病修饰疗法的新靶点。 将使用两种互补的体外模型来确定这些退行性前变化与α-突触核蛋白毒性之间的关系。在酵母中过表达人a-syn重现了在人突触核蛋白病中观察到的细胞缺陷，产生了对由a-syn错误折叠引起的路径生物学的见解，并创建了适合于高通量分析的模型。利用无与伦比的遗传工具，我将使用酵母模型来建立这些变化和α-syn依赖性细胞毒性之间的因果关系，并研究建立这种联系的机制基础。 其次，我将表征来自家族性α-突触核蛋白病患者的诱导多能干细胞（iPS），包括A53 T和α-syn的倍增（重复，三倍）突变。最近的突破性发现使人类体细胞重新编程为多能iPS细胞，为人类疾病的研究提供了前所未有的方法。这些细胞可以稳健地分化成神经元，提供了高度相关的背景，其中验证和扩展来自大鼠AAV和酵母突触核蛋白病模型的发现，并使用无偏基因表达谱产生新的退行性前变化。使用酵母、大鼠和人iPS细胞的多模型方法将促进重要致病途径的验证，为药物开发和其他治疗策略提供新的治疗靶点。

DESCRIPTION (provided by applicant): Synucleinopathies, including Parkinson's disease and dementia with Lewy bodies, are common aging-dependent neurodegenerative diseases associated with neuronal aggregation of a-synuclein. An in vivo rat synucleinopathy model delineated early key pathological changes preceding overt neuronal degeneration. These may represent critical pathophysiological changes potentially causal to disease progression, without being confounded by cell death-related events. I hypothesized that these pathways perturbed in the pre-degenerative stages are important contributors to the pathogenesis of synucleinopathies, and will yield novel targets for disease-modifying therapies. Two complementary in vitro models will be used to determine the relationships between these pre-degenerative changes and a-synuclein toxicity. Over expressing human a-syn in yeast recapitulates cellular defects seen in the human synucleinopathies, yielding insights into the path biology caused by a-syn misfolding and creating a model amenable to high throughput analyses. Taking advantage of unparalleled genetic tools available, I will use the yeast model to establish causal relationships between these changes and a-syn-dependent cytotoxicity, and to investigate the mechanistic underpinning of such connections when established. Second, I will characterize induced pluripotent stem (iPS) cells derived from familial a-synucleinopathy patients including the A53T and multiplication (duplication, triplication) mutations of a-syn. The recent groundbreaking discovery enabling reprogramming of human somatic cells into pluripotent iPS cells offers an unprecedented approach to the study of human diseases. These cells can be robustly differentiated into neurons, providing a highly relevant context in which to validate and extend findings from the rat AAV and yeast synucleinopathy models, and to generate novel pre-degenerative changes using unbiased gene expression profiling. The multi- model approach using yeast, rat and human iPS cells will facilitate the validation of important pathogenetic pathways, offering novel therapeutic targets for drug development and other therapeutic strategies.