Transcriptional Regulation of BACE and the Components of the gamma-Secretase Comp

描述(由申请人提供)：淀粉样蛋白(A)的过量生产被广泛认为会引发一连串的神经变性，导致阿尔茨海默病(AD)。淀粉样前体蛋白(APR)是由淀粉样前体蛋白(APR)通过分泌酶(BACE1)和分泌酶复合体(由早老素、尼卡斯特素、APH-1和PEN-2四种成分组成)依次切割而成。在翻译后水平上对分泌酶活性的调控已做了大量的研究工作，而对任何蛋白质生物发生的第一步--分泌酶成分基因表达的转录调控的研究却很少。最近，我们和其他人已经确定了BACE1、早老素、PEN-2和APH-1的基本启动子序列，以及几个已识别的转录因子。然而，更多的转录因子和相关的调控机制/途径仍有待确定。诸如转录事件如何在不同类型的细胞/组织中不同地影响分泌酶活性，以及分泌酶成分的转录调控在AD发病机制中的重要性等问题是非常重要的，仍有待解决。基于已有的信息和我们的初步结果，我们建议研究以下目标：具体目标1：进一步研究BACE1基因转录调控在缺氧/缺氧和A？中的作用，确定BACE1基因启动子利用的细胞类型特异性，以及BACE1基因转录调控在AD发病机制中的重要性。我们将(1)确定BACE1的转录是否受HIF-1的调控，以及体内HIF-1缺乏是否影响BACE1的表达；(2)确定BACE1转录因子(即SP1和HIF-1)在AD脑中的表达及其活性是否发生变化；(3)确定BACE1在神经元和非神经元细胞中的转录调控；以及(4)研究Aβ和BACE1之间的潜在相互调节，即确定Aβ处理是否调节BACE1的转录。具体目的2：明确尼古丁的启动子和转录调控，确定不同β-分泌酶组成基因启动子利用的细胞类型特异性和A？对它们的转录调控，以及这些基因转录调控在阿尔茨海默病发病和调控β-分泌酶底物特异性中的重要性。我们将：(1)确定尼古丁基因启动子和转录调控的特征；(2)确定AD大脑中调控β-分泌酶成分表达的转录因子水平是否发生改变；(3)确定编码四种β-分泌酶成分的基因的细胞类型特异性转录调控；(4)确定A？处理是否促进β-分泌酶成分的表达；以及(5)确定转录调控对不同底物的β-分泌酶活性的影响。研究这些分泌酶成分的转录调控对于开发有效的AD治疗方法具有重要意义。

DESCRIPTION (provided by applicant): Overproduction of ¿-amyloid (A¿) is widely believed to trigger a cascade of neurodegeneration leading to Alzheimer's disease (AD). A¿ is proteolytically derived from ¿-amyloid precursor protein (APR) through sequential cleavages by ¿-secretase (BACE1) and ?-secretase complex (consisting of four components: presenilin, nicastrin, APH-1 and PEN-2). Much research effort has gone into studying regulation of secretase activity at the post-translation level, transcriptional regulation of gene expression of secretase components, the first step of any protein's biogenesis, has been much less explored. Recently we and others have characterized the basic promoter sequence for BACE1, presenilins, PEN-2 and APH-1, and several identified transcriptional factors. However, additional transcriptional factors and related regulatory mechanisms/pathways still remain to be identified. Questions such as how transcriptional events may differentially affect the secretase activity in different cell/tissue types, and the importance of the transcriptional regulation of the secretase components in AD pathogenesis are highly important and remain to be addressed. Based on the available information and our preliminary results, we propose to study the following aims: Specific Aim 1: to further characterize the transcriptional regulation of BACE1 by HIF/hypoxia and A¿, to determine the cell type specificity of promoter utilization for BACE1, and to correlate the importance of the transcriptional regulation of BACE1 gene in AD pathogenesis. We will (1) determine whether transcription of BACE1 is regulated by HIF-1 and whether HIF-1 deficiency affects BACE1 expression in vivo; (2) determine whether expression of BACE1 transcription factors (i.e. SP1 and HIF-1) and their activity are altered in AD brains; (3) determine transcriptional regulation of BACE1 in neuronal and non-neuronal cells; and (4) study a potential reciprocal regulation between A¿ and BACE1, i.e. to determine whether A¿ treatment regulates transcription of BACE1. Specific Aim 2: to define the promoter and transcriptional regulation of nicastrin, to determine the cell type specificity of promoter utilization for various ?-secretase component genes and their transcriptional regulation by A¿, and to correlate the importance of the transcriptional regulation of these genes in AD pathogenesis and in controlling the ?-secretase substrate specificity. We will (1) characterize promoter and transcriptional regulation of nicastrin gene; (2) determine whether the levels of transcription factors regulating expression of ?-secretase components are altered in AD brains; (3) define the cell type specific transcriptional regulation of genes encoding the four ?-secretase components; (4) determine whether A¿ treatment promotes the expression of ?-secretase components; and (5) determine the effects of transcriptional regulation on the ?-secretase activity to different substrates. Studying transcriptional regulation of these secretase components should be instrumental for developing effective therapies for AD.