ATP-BINDING CASSETTEE TRANSPORTER-2 (ABCA2) REGULATION OF BETA APP PROCESSING

这个子项目是许多利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 该项目的长期目标集中于确定负责淀粉样前体蛋白（APP）加工的机制。 APP的淀粉样蛋白生成加工导致A β肽片段的产生，所述A β肽片段参与阿尔茨海默病（AD）的病因学。调节APP加工的机制尚未完全阐明。 我们进行了一个屏幕的ATP结合盒转运蛋白表达在大脑中的APP代谢的影响。 我们目前的研究结果表明，ATP结合盒转运蛋白（ABCA 2）是APP加工和Abeta生产的调节剂。通过单核苷酸多态性（SNP）作图的遗传证据确定了ABCA 2中与早发性AD显著相关的单个同义突变。 我们确定ABCA 2在携带APP瑞典突变（APPsw）的HEK 293细胞中的过表达增加了细胞APP全蛋白水平和Abeta分泌。 使用基因表达谱的微阵列分析稳定转染ABCA 2的HEK 293细胞，我们检测到一些与AD相关的基因水平升高。 最近生产的ABCA 2基因敲除小鼠将允许明确确定ABCA 2转运蛋白在体内调节APP加工和Abeta产生的功能。 该授权申请的中心假设是ABCA 2在神经元细胞中起调节APP加工和Abeta产生的作用，从而在维持APP代谢的淀粉样蛋白生成和非淀粉样蛋白生成途径之间的稳态中起关键作用。深入了解ABCA 2转运蛋白在体内调节APP加工的作用机制将为开发治疗策略以减少AD预防和治疗中的Abeta负荷提供重要基础。 这一假设将通过追求以下具体目标进行调查： 具体目标1。确定ABCA 2在调节神经元细胞中APP加工和Abeta产生中的作用。由于我们先前已经证明ABCA 2调节非神经元HEK 293细胞中的细胞APP全蛋白水平和Abeta分泌，因此我们建议调查ABCA 2可能调节神经元细胞中的APP加工和Abeta产生的特定假设。 具体目标二：确定ABCA 2在神经元细胞中通过Apoe/低密度脂蛋白受体相关蛋白（LRP）进行APP加工和Abeta产生中的作用。ApoE 4等位基因是AD的一个强遗传风险因子，是低密度脂蛋白受体相关蛋白的配体，介导HDL衍生胆固醇的酯化。 LRP还介导APP加工和Abeta生成。 由于我们报道ABCA 2可能通过胆固醇运输调节N2 a神经母细胞瘤细胞中LRP的表达，我们提出了ABCA 2可能通过ApoE/LRP调节神经元细胞中APP加工和Abeta产生的具体假设。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The long-term goals of this project focus on defining mechanisms responsible for processing of the amyloid precursor protein (APP). Amyloidogenic processing of APP results in the generation of Abeta peptide fragments that are involved in the etiology of Alzheimer's disease (AD). The complete elucidation of the mechanisms that regulate APP processing has not been realized. We undertook a screen of ATP-binding cassette transporters expressed in the brain for their effects on APP metabolism. Our current results suggest that the ATP-binding cassette transporter (ABCA2) is a regulator of APP processing and Abeta production. Genetic evidence by single nucleotide polymorphism (SNP) mapping identified a single synonymous mutation in ABCA2 that is significantly associated with early-onset AD. We determined that ABCA2 overexpression in HEK293 cells bearing the APP Swedish mutation (APPsw) increased cellular APP holoprotein levels and Abeta secretion. Using microarray analysis of gene expression profiles in HEK293 cells stably transfected with ABCA2, we detected elevated levels of a number of genes associated with AD. The recent production of an ABCA2 knockout mouse will permit the unequivocal determination of the function of the ABCA2 transporter in regulating APP processing and Abeta production in vivo. A central hypothesis of this grant application is that ABCA2 functions in neuronal cells to regulate APP processing and Abeta production and thereby serves a critical role in maintaining homeostasis between amyloidogenic and non-amyloidogenic pathways of APP metabolism. In depth knowledge of the mechanisms of action of the ABCA2 transporter in vivo on regulation of APP processing will provide a critical basis for the development of therapeutic strategies to reduce Abeta burden in prevention and treatment of AD. This hypothesis will be investigating by pursuing the following the following specific aims: Specific Aim 1. Determine the role of ABCA2 in regulating APP processing and Abeta production in neuronal cells. Since we have previously demonstrated that ABCA2 regulates cellular APP holoprotein levels and Abetasecretion in non-neuronal HEK293 cells, we propose to investigate the specific hypothesis that ABCA2 may regulate APP processing and Abeta production in neuronal cells. Specific Aim 2: Determine the role of ABCA2 in APP processing and Abeta production through ApoE/low-density lipoprotein receptor-related protein (LRP) in neuronal cells. The ApoE4 allele is a strong genetic risk factor for AD and is a ligand for the low-density lipoprotein receptor-related protein, where it mediates esterification of HDL-derived cholesterol. LRP also mediates APP processing and Abeta generation. Since we report that ABCA2 regulates LRP expression in N2a neuroblastoma cells, perhaps by cholesterol trafficking, we propose the specific hypothesis that ABCA2 may regulate APP processing and Abeta generation through ApoE/LRP in neuronal cells.