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

已知早老素-1或早老素-2基因的染色体突变会导致早发性家族性阿尔茨海默病(AD)。从线虫的遗传分析和早老素-1基因敲除小鼠的表型分析中，早老素也被认为参与了Notch信号转导途径，这对细胞分化的调节是重要的。为了阐明位于AD和Notch通路交叉点的早老素的分子功能，并进一步评估Notch通路参与AD发病的可能性，我们对早老素-1的膜拓扑结构进行了详细研究。我们还利用酵母双杂交系统研究了早老素-1与Notch分子之间可能的物理相互作用。1.早老素的膜拓扑分析：通过在体外转录-翻译系统中表达一系列C端缺失的早老素-1与报告分子之间的一系列融合蛋白，并在微体膜存在下检测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。我们的结果表明，它的C-末端部分的结构与其他组的不同。2.早老素-1与Notch-1相互作用的分析：酵母双杂交分析早老素-1的N-末端、亲水环或C-末端区域与Notch-1的细胞质区域之间可能的直接相互作用。我们的结果排除了早老素-1和至少Notch-1的胞浆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.