The role of Rab1 GTPase and its activators in selective autophagy and neurodegenerative disease

摘要 影响数百万人生活的与年龄相关的神经退行性疾病(ND)是 由神经功能的逐渐丧失引起的。这种损失与累积有关。 聚集的蛋白，例如帕金森病的α-突触核蛋白和阿尔茨海默病的APP 疾病。这些疾病背后的致病机制，目前还没有 治疗方法，还没有完全被理解。自噬，一种细胞循环途径，穿梭于过剩和 将受损的细胞成分转移到溶酶体进行降解。我们认为这是一种构成 选择性自噬途径清除ND相关蛋白聚集体。在哺乳动物细胞中， 在压力条件下，自噬已被广泛研究，但几乎一无所知。 关于构成选择性自噬过程或其在人类细胞中的调节。在这里，我们 建议扩展我们最近在酵母中关于两种成分选择的研究 人细胞自噬途径及其由Rab1酵母同系物Ypt1的调节。 Rab1 GTP酶与ND有关，其激活被认为可以抑制 ND相关聚集体。然而，Rab1同时调节分泌和自噬，而不是 明确它的激活通过哪个过程影响ND。此外，Rab1在分泌物中被激活 与肿瘤的发生有关。因此，为了阐明Rab1通过哪个过程影响ND，它是 对分离其在分泌和自噬中的功能至关重要。在酵母方面，我们成功地 用两种方法分离Ypt1在分泌和自噬中的功能：生成 自噬特异性Ypt1突变和自噬特异性Ypt1激活剂的特征。 这些激活剂在两条结构性选择性自噬途径中刺激Ypt1：清除 过量的膜蛋白和细胞质蛋白复合体的清除。因为所有的 玩家从酵母到人类细胞都是保守的，我们建议确定我们的发现是否 与人类细胞有关。在目标1中，我们将尝试产生自噬特异的Rab1突变 在AIMS 2-3中，我们将确定自噬选择性激活剂是否刺激Rab1 两条截然不同的结构性选择性自噬途径。 如果成功，这些工具将被用于未来的实验，以确定自噬是否- 特异的Rab1调节影响ND相关蛋白聚集体的清除。实现 这项提案的目标和未来的实验将为以下方面提供一种新的监管范式 清除与ND相关的蛋白质聚集体。此外，识别Rab1调节子的特异性 结构性选择性自噬将提供新的新城疫治疗靶点，符合 国家神经疾病和中风研究所的任务。

Abstract Age-related neurodegenerative diseases (ND), which affect the lives of millions of people, are caused by a progressive loss of neuronal function. This loss is associated with the accumulation of aggregated proteins, such as α-synuclein in Parkinson's disease and APP in Alzheimer's disease. The pathogenic mechanisms underling these diseases, for which currently there are no cures, are not fully understood. Autophagy, a cellular recycling pathway, shuttles excess and damaged cellular components to the lysosome for degradation. We propose that constitutive selective autophagy pathways clear ND-related protein aggregates. In mammalian cells, autophagy has been extensively studied under stress conditions, but almost nothing is known about constitutive selective autophagy processes or their regulation in human cells. Here, we propose to extend our recent studies in yeast about two types of constitutive selective autophagy pathways and their regulation by the Rab1 yeast homolog, Ypt1, to human cells. Rab1 GTPase was implicated in ND and its activation was proposed to inhibit accumulation of ND-related aggregates. However, Rab1 regulates both secretion and autophagy and it is not clear through which process its activation affects ND. Moreover, Rab1 activation in secretion was implicated in oncogenesis. Thus, to elucidate through which process Rab1 affects ND, it is crucial to separate its functions in secretion and autophagy. In yeast, we succeeded in separating the functions of Ypt1 in secretion and autophagy using two approaches: generating autophagy-specific Ypt1 mutations and characterizing autophagy-specific Ypt1 activators. These activators stimulate Ypt1 in two constitutive selective autophagy pathways: clearance of excess membrane proteins and clearance of cytoplasmic protein complexes. Because all the players are conserved from yeast to human cells, we propose to determine whether our findings pertain to human cells. In Aim 1, we will attempt to generate autophagy-specific Rab1 mutations and in Aims 2-3, we will determine whether autophagy-selective activators stimulate Rab1 in two distinct constitutive selective autophagy pathways. If successful, these tools will be used in future experiments to determine whether autophagy- specific Rab1 modulation affects clearance of ND-related protein aggregates. Achieving the goals of this proposal and the future experiments would provide a novel regulation paradigm for clearance of ND-related protein aggregates. Moreover, identification of Rab1 regulators specific for constitutive selective autophagy would provide novel ND therapeutic targets, in line with the missions of the National Institute of Neurological Disorders and Stroke.