MOLECULAR MECHANISMS REGULATING THE HUMAN RENIN GENE

肾素-血管紧张素系统(RAS)在这一调节过程中起着重要作用 人类和其他哺乳动物的血压和电解质平衡。 尽管已经在生理水平上对RAS进行了深入的研究 调控系统中基因表达的分子机制 才刚刚开始被发现。基因研究表明 RAS基因与动物和人类高血压之间的联系，以及 转基因研究表明，人的心脏可能会出现异常 RAS导致高血压，对这些有清楚的认识 重要的机制变得至关重要。我们的战略是调查 调控HuRen基因表达的分子机制有两个方面。 首先，将使用CALU-6和AS4.1细胞进行转染分析和 作为检测DNA-蛋白质的转录因子来源 互动。第二，转基因小鼠将作为模型用于 研究特定组织和特定细胞的表达，以及生理 以及对该基因的药理学调节。我们假设基本的 HuRen基因的转录调控是由一组 在5‘侧翼区域内紧密聚集的弱监管因素 以及存在于第一内含子内的消音器。的高水平表达 该基因由一种基因特异性增强子控制。感应性由 CALU-6细胞内cAMP的升高主要是通过 涉及HuRen mRNA成交量降低的转录机制 从而增强了消息稳定性。组织和细胞的特异性 转基因小鼠的基因是由位于近端的区域分配的 启动子、增强子和基因本身的体内。按顺序 为了进一步检验这些假设，我们建议：1)确定具体的 DNA序列和转录因子接触位点调节碱基 Huen基因的转录活性，2)Huen基因的特征 增强子序列，3)识别调节适当组织的序列- 和HuRen基因在转基因小鼠中的细胞特异性，以及4) 探讨cAMP介导的血管紧张素转换酶升高的机制 CALU-6细胞内源性HuRen基因的表达真正的肾素的使用 表达细胞系和转基因动物将为我们提供 无与伦比的工具，将产生有意义和生理上的 相关数据。

The renin-angiotensin system (RAS) plays a major role in the regulation of blood pressure and electrolyte balance in humans and other mammals. Although the RAS has been intensively examined at the physiological level the molecular mechanisms regulating the expression of genes in the system are only beginning to be uncovered. With genetic studies suggesting a link between the RAS genes and hypertension in animals and in humans, and transgenic studies demonstrating the potential for abnormalities in the RAS to cause hypertension, gaining a clear understanding of these important mechanisms becomes essential. Our strategy to investigate the molecular mechanisms regulating HuRen gene expression is two pronged. First, Calu-6 and As4.1 cells will be used for transfection analysis and as a source of transcription factors for examining DNA-protein interactions. Second, transgenic mice will be used as a model to investigate tissue-specific and cell-specific expression, and physiologic and pharmacologic regulation of the gene. We hypothesize that basal transcriptional regulation of the HuRen gene is conferred by a group of closely clustered weak regulatory elements within the 5' flanking region and a silencer present within the first intron. High level expression of the gene is controlled by a gene-specific enhancer. Inducibility by increased intracellular cAMP in Calu-6 cells occurs largely by a post- transcriptional mechanism involving a decrease in HuRen mRNA turnover leading to enhanced message stability. Tissue and cell-specificity of the gene in transgenic mice is conferred by regions located in the proximal promoter, the enhancer, and within the body of the gene itself. In order to further test these hypotheses we propose to: 1) identify the specific DNA sequences and transcription factor contact sites regulating basal transcriptional activity of the HuRen gene, 2) characterize the HuRen enhancer sequence, 3) identify sequences regulating appropriate tissue- and cell-specificity of the HuRen gene in transgenic mice, and 4) investigate the mechanisms causing the cAMP-mediated increase in endogenous HuRen mRNA in Calu-6 cells. The use of bonafide renin expressing cell lines and transgenic animals will provide us with unparalleled tools that will result in meaningful and physiologically relevant data.