Beclin Deficiency and Microglial Impairment in Alzheimer's Disease

描述(由申请人提供)： 阿尔茨海默病(AD)会导致大脑的进行性退化，慢慢破坏患者的认知能力，给家庭和整个社会造成巨大的社会和经济负担。据估计，美国500万阿尔茨海默病患者中有20%是退伍军人，由于目前还没有有效的治疗方法，阿尔茨海默氏症协会预计，未来20年，患者数量将增加两倍。AD的病因(S)在大多数病例中尚不清楚，但越来越多的证据表明，细胞清除和蛋白质降解途径可能在该病中受损，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及其结合伙伴Vps34在AD脑中强烈减少，而在AD模型小鼠中，减少beclin 1会导致更多的A？积累和神经退化。尽管如此，这些影响的确切机制仍不清楚。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同源基因Atg6调节酵母中的空泡蛋白分类，这是一种基本的溶酶体分类途径，可以促进细胞成分的降解或重新利用。此外，在哺乳动物细胞中，包括Beclin 1在内的自噬蛋白定位于吞噬杯中，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会削弱它们从原位脑切片上清除淀粉样斑块的能力，以及在培养中吞噬乳胶珠的能力。我们发现，敲除beclin 1会导致逆转录蛋白VPS35水平降低，VPS35是一种哺乳动物蛋白质，参与将细胞成分分选到溶酶体或返回到特定的隔室(例如细胞表面)，并损害吞噬细胞受体CD36的循环。有趣的是，Beclin 1可以被caspase切割，由此产生的片段可能会干扰正常的beclin 1功能。综上所述，我们提出假设，减少的Beclin 1或Beclin 1片段通过扰乱VPS35介导的吞噬受体的循环而损害小胶质细胞的吞噬功能，导致AD期间A和细胞碎片的积累。为了验证这一假说及其在AD进展中的作用，我们建议确定Beclin 1减少后的吞噬功能受损是否由VPS35水平降低和吞噬受体循环受损所介导，以及全长Beclin 1或其caspase裂解片段是否调节吞噬和受体循环。此外，我们建议确定小胶质细胞Beclin 1减少对小鼠吞噬功能和阿尔茨海默病样疾病的影响。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和其他蛋白质或颗粒的清除。这一知识有望帮助我们确定治疗这种毁灭性疾病的新目标。

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.