Intersection between familial Alzheimer's disease and Notch signal transduction pathway

已知早老素-1或早老素-2基因的染色体突变可导致早发性家族性阿尔茨海默病（AD）。从秀丽隐杆线虫的遗传分析和早老素-1敲除小鼠的表型分析来看，早老素还参与Notch信号转导通路，对细胞分化的调控具有重要意义。为了第一步阐明早老素的分子功能，并进一步评估Notch通路参与AD病因学的可能性，我们详细研究了早老素-1的膜拓扑结构。我们还利用酵母双杂交系统研究了早老素-1和Notch分子之间可能的物理相互作用。早老素的膜拓扑分析：通过在体外转录-翻译系统中表达c端缺失的早老素-1与报告基因之间的一系列融合蛋白，并检测微粒体膜存在下的n-糖基化和对蛋白酶K的保护作用，研究了早老素-1的膜拓扑结构。结果表明，其c端结构与其他研究小组提出的结构不同。基于本研究结果，我们提出了早老素膜拓扑结构的“七跨一嵌”模型。早老素-1与Notch-1的相互作用分析：采用酵母双杂交分析方法评估早老素-1的n端、亲水环或c端区域与Notch-1细胞质区域之间可能存在的直接相互作用。我们的结果排除了早老素-1和至少Notch-1细胞质区之间物理相互作用的可能性。

Chromosomal mutations in the gene for presenilin-1 or presenilin-2 are known to cause early-onset familial Alzheimer's disease (AD). From genetic analysis of C.elegans and phenotypical analysis of presenilin-1 knock-out mice, presenilins are also implicated to be involved in the Notch signal transduction pathway, which is important to the regulation of cell differentiation. For the first step to elucidate the molecular function of presenilins, which are situated in the intersection between AD and Notch pathway, and further to assess the possibility of Notch pathway involvement in the etiology of AD,we studied the membrane topology of presenilin-1 in detail. We also examined the possible physical interaction between presenilin-1 and Notch molecule using yeast two-hybrid system.1.Membrane topological analysis of presenilin : Membrane topology of presenilin-1 was studied by expressing a series of fusion proteins between C-terminal deleted presenilin-1 and a reporter in in vitro transcription-translation system and examining N-glycosylation and protection against proteinase K in the presence of microsomal membrane. Our results indicated that the structure of its C-terminal portion is different from those suggested by other groups. Based on the results of this study, we propose the "seven-spanning and one-embedded" model for presenilin membrane topology.2.Analysis of the interaction between presenilin-1 and Notch-1 : Yeast two-hybrid analysis was employed to assess possible direct interaction between the n-terminal, hydrophilic loop or C-terminal region of presenilin-1 and the cytosolic region of Notch-1. Our result excluded the possibility of physical interaction between presenilin-1 and, at least, the cytosolic region of Notch-1.