PRESENILIN GENE STUDY IN DROSOPHILA MELANOGASTER AS MODEL

对早发性家族性阿尔茨海默病(AD)家系的遗传连锁研究已经确定，至少三个基因座的突变将导致该病：淀粉样β蛋白前体基因、早老素-1基因(PS-1)和早老素-2基因(PS-2)。虽然已有报道表明，携带PS-1和PS-2突变的家族的成纤维细胞中的β-淀粉样蛋白产生异常，线虫中的一种同源蛋白影响Notch信号转导，但早老素基因的功能目前尚不清楚。因此，这一应用的长期目标是确定早老素蛋白和其他早老素相互作用蛋白的功能。果蝇是确定未知基因功能的理想模式生物，在这种情况下，这一点更加理想，因为大多数关于Notch信号的研究都是在果蝇中进行的。利用早老素基因之间的高度保守性，我们实验室成功地在果蝇中克隆了该基因。该项目的第一个具体目标将是确定果蝇中该基因的特征，研究其基因结构、发育转录模式、在胚胎、幼虫、蛹和成虫中的定位以及DNA测序。将使用南方和北方印迹技术和光学显微镜来进行这一部分的项目。第二个具体目标是产生该基因的零突变，并研究它们对果蝇的影响。将使用P-元件插入突变来产生突变果蝇，并对这些果蝇进行遗传特征分析。第三个具体目标是确定哪些基因与早老素基因相互作用。苍蝇遗传学和酵母双杂交方法将被用来寻找这些与早老素相互作用的基因。这个项目将使人们更好地了解早老素基因的生物学和功能，早老素基因是早发性家族性阿尔茨海默病的主要原因。通过确定早老素的功能，以及它们与其他基因的相互作用，将对AD的发病机制有更多的了解，这最终可能导致改进对该疾病的治疗。此外，对这些早老素相互作用蛋白的长期研究将有助于我们确定在早发和晚发病例中可能导致或影响AD的其他人类基因座。

Genetic linkage studies in early onset-familial Alzheimer's disease (AD) families have determined that mutations in at least three loci will cause the disease: the amyloid beta-protein precursor gene, the presenilin-1 gene (PS-1), and the presenilin-2 gene (PS-2). While it has been reported that beta-amyloid production is abnormal in fibroblasts from families carrying PS-1 and PS-2 mutations, and a homologues protein in C. elegans affects Notch signaling, the function of the presenilin gene is presently unknown. Therefore, the long-term objective of this application is to determine the function of the presenilin protein and other presenilin- interacting proteins. Drosophila melanogaster is an ideal model organism for determining the function of uncharacterized genes, which in this case is made even more ideal as most studies of Notch signaling have been carried out in Drosophila. Using the high conservation among conservation among presenilin genes, our lab has successfully coned the gene in Drosophila. The first specific aim of this project will be to characterize this gene in Drosophila, studying its gene structure, its developmental transcription pattern, and its localization in embryos, larvae, pupae, and adult flies, DNA sequencing. Southern and Northern blotting techniques, and light microscopy will be used to carry out this part of the project. The second specific aim will be to produce null mutations of this gene and study their effects in Drosophila. P-element insertion mutagenesis will be used to produce the mutant flies, and the flies will be characterized genetically. The third specific aim will be to determine which genes interact with the presenilin gene. Fly genetics and yeast two-hybrid approaches will be used to find these presinilin- interacting genes. This project will lead to a greater understanding about the biology and function of the presenilin genes, the major cause of early- onset familial Alzheimer's disease. By determining the functions of the presenilins, and their interactions with other genes, a greater understanding of the pathogenesis of AD will be gained, which may lead eventually to improved treatment of the disease. Additionally, a long- term study of these presenilin-interacting proteins will help us to identify other human loci which may cause or influence AD in both early-onset and late-onset cases.