Beclin Deficiency and Microglial Impairment in Alzheimer's Disease

描述（由申请人提供）： 阿尔茨海默氏病（AD）产生大脑的进行性退化，缓慢地破坏受害者的认知能力，并对家庭和整个社会造成巨大的社会和经济负担。据估计，在500万患有AD的美国人中，多达20%是退伍军人，由于目前没有有效的治疗方法，阿尔茨海默氏症协会预计，在未来20年内，患者人数将增加两倍。在大多数情况下，AD的病因尚不清楚，但越来越多的证据表明，该疾病的细胞清除和蛋白质降解途径可能受损，为治疗干预提供了潜在的新靶点。 最近，包括我们实验室在内的几项研究表明，自噬蛋白beclin 1与A？周转和神经退行性变有关。这些研究表明，全长beclin 1及其结合伴侣vps34在AD大脑中强烈减少，并且在AD模型小鼠中减少beclin 1导致更多的A？积累和神经变性。这些影响的确切机制仍然不清楚。Beclin 1被经典地归因于启动自噬，这是参与长寿命蛋白质和细胞器降解的主要途径。然而，在酵母中的研究表明，beclin 1也可能在其他细胞过程中发挥关键作用。最值得注意的是，beclin 1直系同源物Atg 6调节酵母中的液泡蛋白分选，这是一种基本的溶酶体分选途径，可以促进细胞组分的降解或再利用。此外，在哺乳动物细胞中，自噬蛋白，包括beclin 1，定位于吞噬杯，自噬可以增强吞噬作用。我们的初步数据表明，减少小胶质细胞中的beclin 1会损害它们从原位脑切片中清除淀粉样蛋白斑块以及吞噬培养中的乳胶珠的能力。我们发现，敲低beclin 1导致逆转录蛋白vps 35水平降低，vps 35是一种哺乳动物蛋白，参与将细胞组分分选到溶酶体或回到限定的区室（例如，细胞表面）和受损的吞噬受体CD36的再循环。有趣的是，beclin 1可以被半胱天冬酶切割，产生的片段可能会干扰正常的beclin 1功能。 综上所述，我们提出了这样的假设，即减少beclin 1或beclin 1片段通过破坏vps35介导的吞噬受体再循环来损害小胶质细胞的吞噬作用，导致AD期间A？和细胞碎片的积累。为了验证这一假设及其与AD进展的相关性，我们建议确定beclin 1减少后吞噬功能受损是否由vps35水平降低和吞噬受体再循环受损介导，以及全长beclin 1或其半胱天冬酶切割片段是否调节吞噬作用和受体再循环。此外，我们建议确定减少小胶质细胞beclin 1对小鼠吞噬作用和阿尔茨海默病样疾病的后果。在拟议的研究结束时，我们希望能更好地了解beclin 1在小胶质细胞中的功能，以及在AD患者中观察到的beclin 1缺陷如何损害小胶质细胞对A？和大脑中其他蛋白质或颗粒的清除。这些知识将有望帮助我们确定治疗这种毁灭性疾病的新靶点。

DESCRIPTION (provided by applicant): Alzheimer's disease (AD) produces a progressive degeneration of the brain that slowly destroys a victim's cognitive abilities and inflicts tremendous social and economic burden on families and society in general. As many as a 20% of the estimated five million Americans with AD are Veterans, and with no effective treatments available currently, the Alzheimer's Association expects the number of patients will triple in the next 20 years. The cause(s) of AD is unknown in most cases but growing evidence suggests that cellular clearance and protein degradation pathways may be impaired in the disease providing potential new targets for therapeutic intervention. Recently, several studies, including from our lab, have implicated the autophagy protein, beclin 1, in A¿ turnover and neurodegeneration. These studies show that full length beclin 1 together with its binding partner vps34 is strongly reduced in AD brains and that reducing beclin 1 in AD model mice results in more A¿ accumulation and neurodegeneration. Still the precise mechanisms of these effects remain unclear. Beclin 1 is classically attributed with initiating autophagy, which is the major pathway involved in the degradation of long- lived proteins and organelles. However, studies in yeast show that beclin 1 may also play a critical role in other cellular processes. Most notably, the beclin 1 ortholog Atg6 regulates vacuolar protein sorting in yeast, a rudimentary lysosomal sorting pathway that can promote the degradation or re-use of cellular components. Moreover, in mammalian cells autophagy proteins, including beclin 1, localize to phagocytic cups and autophagy can enhance phagocytosis. Our preliminary data demonstrate that reducing beclin 1 in microglia impairs their capacity to clear amyloid plaques from brain sections in situ as well as to phagocytose latex beads in culture. We find that knocking down beclin 1 leads to reduced levels of the retromer protein vps35, a mammalian protein involved in sorting cellular components to the lysosome or back to defined compartments (e.g., the cell surface) and impaired recycling of the phagocytic receptor CD36. Interestingly, beclin 1 can be cleaved by caspases and the resulting fragments may interfere with normal beclin 1 function. Taken together we propose the hypothesis that reduced beclin 1 or beclin 1 fragments impair microglial phagocytosis by disrupting vps35-mediated recycling of phagocytic receptors, resulting in the accumulation of A¿ and cellular debris during AD. To test this hypothesis and its relevance in contributing to the progression of AD we propose to determine whether impaired phagocytosis following beclin 1 reduction is mediated by diminished vps35 levels and impaired phagocytic receptor recycling, and whether full-length beclin 1 or its caspase-cleaved fragments modulate phagocytosis and receptor recycling. In addition, we propose to determine the consequences of reduced microglial beclin 1 on phagocytosis and Alzheimer's-like disease in mice. At the end of the proposed studies we expect to have a better understanding of the function of beclin 1 in microglia and how beclin 1 deficiency observed in AD patients may impair microglia clearance of A¿ and other proteins or particles in the brain. This knowledge will hopefully help us to identify novel targets for the treatment of this devastating disease.