MOLECULAR MECHANISMS REGULATING THE HUMAN RENIN GENE

肾素-血管紧张素系统（RAS）在调节中起主要作用， 血压和电解质平衡的影响。 虽然RAS已经在生理水平上进行了深入的研究， 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， RAS引起高血压，获得这些明确的认识 重要机制变得至关重要。我们的调查策略 调控HuRen基因表达的分子机制有两个方面。 首先，Calu-6和As4.1细胞将用于转染分析， 作为检测DNA-蛋白质的转录因子的来源 交互.其次，转基因小鼠将被用作模型， 研究组织特异性和细胞特异性表达，以及生理学 和基因的药理学调节。我们假设基础 HuRen基因的转录调控由一组 在5'侧翼区域内紧密聚集的弱调控元件 和存在于第一内含子内的沉默子。的高效表达 该基因由基因特异性增强子控制。感应器 Calu-6细胞内cAMP的增加主要是通过后 HuRen mRNA周转减少的转录机制 导致增强的消息稳定性。组织和细胞特异性 转基因小鼠中的基因是由位于近端的区域赋予的。 启动子，增强子，以及基因本身的体内。为了 为了进一步验证这些假设，我们建议：1）确定具体的 DNA序列和转录因子接触位点调节基底膜的形成 HuRen基因的转录活性，2）表征HuRen基因的转录活性， 增强子序列，3）鉴定调节适当组织的序列- 和转基因小鼠中HuRen基因的细胞特异性，以及4） 研究引起cAMP介导的 Calu-6细胞内源性HuRen mRNA表达。使用真正的肾素 表达细胞系和转基因动物将为我们提供 无与伦比的工具，将导致有意义的， 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.