BIOSYNTHESIS AND PROCESSING OF AB IN NT2N CELLS

老年斑--阿尔茨海默病的主要神经病理特征 (AD)，主要由衍生的淀粉样肽(Abeta)组成 通过加工淀粉样前体蛋白(APP)。两种主要形式 淀粉样蛋白的长度分别为40和42个氨基酸。而Abeta(1-40)则更多 丰富的Abeta(1-40)更容易聚集，并构成大多数 SPS中淀粉样蛋白的含量。APP中与家族性相关的突变 通过增加Abeta的总量来改变Abeta的生产 Abeta产生的，或通过导致 产生Abeta(1-40)。因此，应用程序处理和Abeta中的扰动 生产可能在AD的发展中发挥重要作用。因此， 项目1的目标是继续描述应用程序处理 人类神经元模型系统(NT2N细胞)中的通路。我们最近发现 APP在内质网/中间体中的滞留 内质网(ER/IC)阻断Abeta的分泌，但细胞内Abeta被抑制 还在生产。定量ELISA法显示ER/IC相关的Abeta是 完全由Abeta(1-42)组成。这种神经元内的Abeta(1-42) 在NT2N中积累数周，同时变得不那么溶解。 这些观察结果对于理解AD的发病机制具有重要意义。 包括到早老素(即PS1和PS2)的可能链接，这些链接是 膜蛋白主要定位于导致AD的ER/IC，当它们 在流行的家系中发生了突变。有趣的是，与时尚相关的形式 突变的早老素增加了Abeta(1-42)的水平。长期的 聚集在神经元内的Abeta(1-42)可能具有神经毒性 并最终导致神经元死亡后SPS的形成。我们 建议扩大我们对这一新途径的研究，以研究其他 产生Abeta的亚细胞隔间，并研究其影响 早老素通过以下特定的方式影响Abeta的生产 目标：1)继续我们最近对ER/IC通路的研究 产生细胞内Abeta(1-42)；2)研究潜在的相互作用 在早老素和APP之间，并检测野生型如何表达 或突变体PS1和PS影响细胞内Abeta的产生；3)检查 内吞途径在细胞内和细胞内分泌的作用 分泌Abeta(1-40)和Abeta(1-42)；以及4)研究野生型和 突变形式的APP751和770由NT2N神经元处理。

Senile plaques, a major neuropathologic feature of Alzheimer's disease (AD), are composed largely of the amyloid peptide (Abeta) that is derived by processing of the amyloid precursor protein (APP). The two major forms of amyloid are 40 and 42 amino acid long. While Abeta(1-40) is more abundant, Abeta(1-40) aggregates more readily and comprises the majority of the amyloid in SPs. Mutations in APP that are associated with familial AD (FAD) alter the Abeta production by increasing the total amount of Abeta generated, or by causing a relative increase in the amount of Abeta(1-40) produced. Thus, perturbations in APP processing and Abeta production may play an important role in the development of AD. Therefore, the goals of Project 1 are to continue delineation of APP processing pathways in human neuronal model system (NT2N cells). We recently found that retention of APP in the endoplasmic reticulum/intermediate compartment (ER/IC) blocked Abeta secretion, but intracellular Abeta was still produced. Quantitative ELISA showed that ER/IC-associated Abeta is composed exclusively of Abeta(1-42). This intra-neuronal Abeta(1-42) accumulates over a period of weeks in NT2N while becoming less soluble. These observations have implications for understanding AD pathogenesis, including possible links to the presenilins (i.e., PS1 and PS2) which are membrane proteins largely localized to the ER/IC that cause AD when they are mutated in FAD pedigrees. Interestingly, FAD-associated forms of mutant the presenilins increased the levels of Abeta(1-42). Long-term accumulation of aggregated intra-neuronal Abeta(1-42) might be neurotoxic and ultimately lead to the formation of SPs following neuronal death. We propose to extend our studies on this novel pathway, to examine other subcellular compartments where Abeta is produced, and to study the effects of the presenilins on Abeta production through the following Specific Aims: 1) to continue our recent studies on the ER/IC pathway by which intracellular Abeta(1-42) is produced; 2) to study potential interactions between the presenilins and APP, and examine how expression of wild type or mutant PS1 and PS affects intracellular Abeta production; 3) to examine the role of the endocytic pathway in the production of intracellular and secreted Abeta(1-40) and Abeta(1-42); and 4) to study how wild-type and mutant forms of APP751 AND 770 are processed by NT2N neurons.