Determining the Interacting Effects of GBA, SNCA, and APOE on a-Synuclein Pathology Severity in Dementia with Lewy Bodies and Parkinson's Disease

项目摘要 包括路易体痴呆症(DLB)和帕金森氏病(PD)在内的联核病正在日益增多 美国有100多万人受到影响，全球估计有1000万人受到影响。 突触核病是由于蛋白质α-突触核蛋白(α-SYN)积聚到大脑中非自然的纤维中而引起的 并可表现为运动症状、认知症状或两者的组合。目前有 没有有效的治疗方法来治愈或预防联核症，很大程度上是因为没有明确的原因 疾病。有许多已确定的遗传和环境风险因素与联核病有关， 但这些因素导致疾病的背景仍然未知。除了直接突变到SNCA之外， 编码α-SYN、gba和apoE突变的基因是公认的危险因素，并已被 表现为增加α-SYN聚集。然而，这些风险因素都不是疾病的重要预测因素。 提示未知的遗传和环境相互作用可能影响疾病的发生、严重程度和 α-SYN病理的临床转归。在这项建议中，我将重点放在细胞类型和基因如何与 相互调节α-SYN的病理严重程度。我假设α-SYN病理是由上位性因素促成的 破坏不同细胞类型动态平衡的遗传因素与叶片多巴胺能之间的相互作用 神经元容易受到有害的α-SYN病理和神经变性的影响。我将使用以下工具研究这一假设 两种独立的、互补的方法来剖析细胞和分子调节机制 α-syn表型。首先，我将GBA基因敲除与SNCA结合--三倍增殖型、野生型和基因敲除细胞系。 在gba突变和α-syn聚集之间有明显的联系，但这些蛋白是如何连接的仍是个未知数。 未知。我将使用高含量成像和基因编码的荧光标记来量化溶酶体和 活细胞中的线粒体动力学。这将揭示gba如何响应不同的α-syn剂量，以及如何 影响细胞的动态平衡。其次，我将利用含有APOE等位基因变异和GBA的等基因系 击倒对手。GBA和APOE参与脂类代谢，提示脂类之间可能存在联系 代谢紊乱和α-SYN病理，尽管这两种蛋白之间没有公开发表的联系。 我将使用下一代基因组测序和蛋白质组学方法来确定相互作用的影响 SNCA、Gba和ApoE等位基因变异对α-SYN病理严重程度的影响。完成这一目标将揭开 无论GBA和APOE是通过汇聚的还是独立的途径发挥作用，扩大了我们对 对疾病病理学有贡献的网络。通过将多个遗传风险因素结合成多细胞类型 模型，我将开创新的技术和方法来剖析影响α-SYN的上位机制 病理学。这些见解将指导我们开发新的治疗方法，使更早的检测成为可能， 治疗和预防联体核病。

Project Summary Synucleinopathies, including dementia with Lewy bodies (DLB) and Parkinson’s disease (PD), are a growing health crisis, affecting over 1 million people in the United States and an estimated 10 million people globally. Synucleinopathies are caused by accumulation of the protein α-synuclein (α-syn) into unnatural fibrils in the brain and can manifest with motor symptoms, cognitive symptoms, or a combination of the two. There are currently no effective treatments to cure or prevent synucleinopathies, in large part because there is no clear cause for disease. There are many identified genetic and environmental risk factors associated with synucleinopathies, but the context in which these factors lead to disease is still unknown. In addition to direct mutations to SNCA, the gene that encodes α-syn, mutations to GBA and APOE are well established risk factors and have been shown to increase α-syn aggregation. However, none of these risk factors are great predictors of disease on their own, suggesting unknown genetic and environmental interactions likely influence the initiation, severity, and clinical outcomes of α-syn pathology. In this proposal, I focus on how both cell type and genotype interact with each other to modulate α-syn pathology severity. I hypothesize that α-syn pathology is precipitated by epistatic interactions between genetic factors that disrupt homeostasis in different cell types and leaves dopaminergic neurons vulnerable to deleterious α-syn pathology and neurodegeneration. I will investigate this hypothesis using two independent, complementary approaches to dissect the cellular and molecular mechanisms that modulate α-syn phenotypes. First, I will couple GBA knockdown with SNCA-triplication, wild-type, and knockout cell lines. There is a clear link between GBA mutations and α-syn aggregation, but how these proteins connect is still unknown. I will use high-content imaging with genetically encoded fluorescent markers to quantify lysosomal and mitochondrial dynamics in live cells. This will reveal how GBA responds to different α-syn dosages and how that affects cellular homeostasis. Second, I will leverage isogenic lines containing APOE allelic variants and GBA knockdown. GBA and APOE are involved in lipid metabolism, suggesting a putative link between lipid metabolism disruption and α-syn pathology, although there is no published connection between the two proteins. I will use next generation genomic sequencing and proteomic approaches to determine the interacting effects between SNCA, GBA, and APOE allelic variants on α-syn pathology severity. Completion of this aim will uncover whether GBA and APOE are acting through converging or independent pathways, expanding our knowledge of the network that contributes to disease pathology. By coupling multiple genetic risk factors into a multi-cell type model, I will pioneer new technology and approaches to dissect epistatic mechanisms that influence α-syn pathology. These insights will guide us in developing new therapeutic approaches, enabling earlier detection, treatment, and prevention of synucleinopathies.