双拷贝APP/BACE/DPsn转基因果蝇模型的建立及基因功能的研究

Construction and functional study of a transgenic Drosophila model with two copies of APP/BACE/DPsn genes

阿尔茨海默症(Alzheimer's disease,AD),是一种以脑中β-淀粉样蛋白(β-amyloid peptide,Aβ)沉积为主要病理改变的神经退行性疾病。在果蝇Drosophila模型中建立淀粉样蛋白前体蛋白(amyloid precursor protein,APP)的剪切通路模拟Aβ的产生过程,有望建立一种快速筛选治疗AD药物的动物模型。我们利用经典的Gal4/UAS系统,将现有的APP/BACE/DPsn果蝇品系连续杂交,通过同源重组的方法构建表达两个拷贝的APP/BACE/DPsn稳定可遗传的转基因果蝇新品系。进一步的实验结果表明:与不表达APP/BACE/DPsn的对照果蝇w/y;APP/Cyo;BACE-DPsn/TM6BTb相比,表达两拷贝APP/BACE/DPsn的w/y;elav-APP;BACE-DPsn果蝇的最长寿命为52d,比对照组(69d)缩短了17d,为对照组果蝇的75%;中位生存时间为39d,比对照组(49d)缩短了10d,为对照组的80%;平均寿命为37d,比对照组(47d)缩短了10d,为对照组的79%。同时,表达两个拷贝APP/BACE/DPsn的果蝇所产卵的羽化时间比对照果蝇延长了3d;其羽化成虫的理论值为1∶9(11%),而实际羽化率仅为5.2%。结果提示,由elav-Gal驱动在果蝇泛神经元内过表达APP/BACE/DPsn,可以缩短果蝇寿命、干扰果蝇胚胎正常发育。该果蝇有可能作为初步筛选AD治疗药物的动物模型,为AD治疗新药的发现提供工具。

Alzheimer′s disease （AD） is a kind of cognitive dysfunction disease and β-amyloid （Aβ） generation is crucial for AD pathogenesis and plays a key role in disease progression. A transgenic fly expressing two copies of APP/BACE/DPsn to mimic the pathologic changes of AD might be useful for AD therapeutic drug screening. Using the classic Gal4/UAS system, we constructed the stable transgenic flies expressing two copies of APP/BACE/DPsn genes by consecutive crosses and homologus recombination. Further tests revealed that the lifespan and the medium survival time of flies expressing two copies of APP/BACE/DPsn genes were 52 d and 39 d, respectively, which were much shorter than the lifespan of 69 d and the medium survival time of 49 d of the control flies. Furthermore, the eclosion time of the flies expressing two copies of APP/BACE/DPsn genes was 3 d longer than that of the control flies, and the eclosion rate was 5.2% which was much less than the theoretical value 1∶9 （11%）. The results suggest that the elav-Gal driven neuronal expression of two copies of APP/BACE/DPsn genes in flies leads to shorter lifespan and decreases viability of the offspring. These phenotypes of the transgenic fly might be used as a preliminary drug screen model for AD therapy.