AUTOPHAGY FUNCTION & DYSFUNCTION IN ALZHEIMER'S DISEASE

自噬是细胞降解受损细胞器和细胞质以及保护自身免受蛋白酶体不能降解的有毒蛋白质聚集的主要机制。我们将讨论揭示阿尔茨海默病(AD)中自噬的诱导和损害的新线索，确定其在Abeta生成中的作用，并展示自噬功能中对早老素(PS1)的需求。自噬在AD中受损的假设将在人脑和FAD成纤维细胞、细胞系和AD病理的小鼠模型中进行研究(目标1和2)。标记自噬途径和其他细胞器特定区段的新型抗体将用于双重免疫荧光和超微结构研究(包括免疫金)，以及其他最先进的技术，以确定 AD的自噬功能障碍。将确定PS1、PS2和g-分泌酶活性在调节自噬中的作用，并将调查观察到的FAD相关PS1突变导致的自噬介导的蛋白质降解选择性损害的基础(目标2)。我们还将调查21三体(唐氏综合征)中独特的自噬异常的基础，并测试FAD相关APP突变(目标3)也会损害自噬的假设。自噬作为一种神经保护机制将在正常小鼠、AD小鼠模型和一种新型转基因小鼠中进行研究，该转基因小鼠表达GFP标记的LC3，一种与自噬诱导相关的蛋白质，将使实时 自噬的可视化，并将有助于在体内研究自噬的调节(目标4)。作为调节自噬作为神经退行性疾病治疗干预的前奏，我们将研究在活体中操纵大脑自噬的策略(目标4)。这些正在进行的研究是第一次系统地研究AD的自噬，将全面评估自噬和自噬功能障碍的影响，并确定治疗包括AD在内的蛋白质病的新方法。

Autophagy is the cell's primary mechanism to degrade damaged organelles and cytoplasm and to protect itself from toxic protein aggregates not degradable by the proteasome. We will address new leads revealing induction and impairment of autophagy in Alzheimer's Disease (AD), identifying its role in Abeta generation, and showing for a requirement for presenilin (PS1) in autophagy function. The hypothesis that autophagy is impaired in AD will be investigated in human brain and FAD fibroblasts, cell lines, and mouse models of AD pathology (Aims 1 and 2). Novel antibodies marking specific compartments of the autophagic pathway and other organelles will be used in double-immunofluorescence and ultrastructural studies (including immunogold), along with other state-of-the-art techniques, to define the nature and significance of autophagy dysfunction in AD. The roles of PS1, PS2, and g-secretase activity in modulating autophagy will be determined and the basis for the observed selective impairment in autophagy-mediated protein degradation caused by FAD-related PS1 mutations will be investigated (Aim 2). We will also investigate the basis for distinctive autophagy abnormalities in Trisomy 21 (Down syndrome) and test the hypothesis that FAD-related APP mutations (Aim 3) also impair autophagy. Autophagy as a neuroprotective mechanism will be studied in primary neurons from normal mice, AD mouse models, and a novel transgenic mouse expressing GFP-tagged LC3, a protein associated with autophagy induction, which will enable real-time visualization of autophagy and will facilitate in vivo studies of autophagy modulation (Aim 4). As a prelude to modulating autophagy as a therapeutic intervention in neurodegenerative disease, we will investigate strategies to manipulate autophagy in the brain in vivo (Aim 4). These ongoing studies, the first to investigate autophagy systematically in AD, will provide a comprehensive assessment of the impact of autophagy and autophagy dysfunction and identify novel approaches to treat proteinopathies, including AD.