Lahiri Debomoy K, Maloney Bryan, Ge Yuan-Wen
Laboratory of Molecular Neurogenetics, Department of Psychiatry, Institute of Psychiatric Research, Indiana University School of Medicine, Indianapolis, IN 46202, USA.
J Mol Neurosci. 2006;28(2):193-210. doi: 10.1385/JMN:28:2:193.
The amyloid-beta (Abeta) peptide, the proteolytic fragment of Abeta precursor protein (APP), aggregates and forms neuritic plaques, a major hallmark of Alzheimer's disease (AD). The limiting step in generating the Abeta peptide from APP is cleavage by the beta-secretase enzyme, BACE1. Regulation of the BACE1 gene is likely to play an important role in AD etiology and treatment. We therefore studied the activity of a 4.1-kb 5'-flanking region (-3765/+364, +1 being the transcription start site) of the BACE1 gene, both in 5'- and 3'-deletion series and through Northern blotting. We show that the BACE1 promoter has regulatory activity throughout the 4.1-kb length, both positive and negative, and that this activity can be quantitatively modeled according to promoter sequence length, with the specific model depending on the presence of negative regulatory elements as the 5'- most portion of the sequence. We also examined a previously identified 141-bp proximal fragment (+224/+364) of the BACE1 promoter and two constituent (91- and 50-bp) subfragments. We report that the 91-bp fragment (+224/+314) is the most likely seat of neuronal expression of the BACE1 gene and that it is the portion of the 141-bp fragment that accounts for observed DNA-protein interactions in brain extracts. The 50-bp fragment (+315/+364), which showed significant reporter gene activity from the empty vector, binds nuclear proteins in a cell type-specific manner and contains the AP2 site as shown by the electrophoretic mobility shift assay. Overall, the 141-bp fragment had no strong matches within GenBank, and the 91-bp fragment is predicted to have several potential stem-loop sites. Taken together, BACE1 gene promoter activity is differentially regulated, and the 91-bp fragment represents a novel promoter region for cell type-specific regulation. This fragment might be a useful target to regulate BACE1 expression leading to Abeta production and to understand the neuropathogenesis of AD.
淀粉样β蛋白（Aβ）肽是Aβ前体蛋白（APP）的蛋白水解片段，会聚集并形成神经炎性斑块，这是阿尔茨海默病（AD）的主要标志。从APP生成Aβ肽的限速步骤是由β-分泌酶BACE1进行切割。BACE1基因的调控可能在AD的病因和治疗中发挥重要作用。因此，我们通过5'端和3'端缺失系列以及Northern印迹法研究了BACE1基因4.1kb 5'侧翼区域（-3765 / +364，+1为转录起始位点）的活性。我们发现BACE1启动子在整个4.1kb长度上都具有调控活性，包括正向和负向活性，并且这种活性可以根据启动子序列长度进行定量建模，具体模型取决于序列5'端最末端的负调控元件的存在情况。我们还检测了先前鉴定的BACE1启动子141bp近端片段（+224 / +364）及其两个组成的（91bp和50bp）亚片段。我们报告91bp片段（+224 / +314）最有可能是BACE1基因神经元表达的位点，并且它是141bp片段中负责在脑提取物中观察到的DNA - 蛋白质相互作用的部分。50bp片段（+315 / +364）在空载体中显示出显著的报告基因活性，以细胞类型特异性方式结合核蛋白，并且如电泳迁移率变动分析所示含有AP2位点。总体而言，141bp片段在GenBank中没有强匹配项，91bp片段预计有几个潜在的茎环结构位点。综上所述，BACE1基因启动子活性受到差异调控，91bp片段代表了一个用于细胞类型特异性调控的新型启动子区域。该片段可能是调节BACE1表达从而导致Aβ产生以及理解AD神经发病机制的有用靶点。
