ROLE OF GLUCOSE IN hAT1 GENE EXPRESSION

描述（申请人提供）：高血压和糖尿病是糖尿病肾病发病的两大危险因素。血管紧张素转换酶抑制剂治疗在糖尿病肾病患者中广泛有效，提示肾素血管紧张素系统在该疾病的进展中起重要作用。血管紧张素II是肾素血管紧张素系统的活性成分，主要通过血管紧张素1型（AT1）受体起作用。与大鼠中AT1受体由两个基因（AT1A, AT1B）编码相比，人类中只有一个基因编码血管紧张素1型（hAT1），提示该受体具有物种特异性表达和差异调控。在糖尿病中，从发病到终末期肾病，hAT1受体的表达在肾脏的不同区域有所不同，但有一个例外，即近端肾小管，该受体在近端肾小管中一直下调，提示该受体有小管特异性调节。ACE抑制剂不能逆转hAT1受体的减少，表明受体的下调不是由血管紧张素II的上调介导的。糖尿病中导致hAT1受体下调的分子机制尚不清楚。近端小管中AT1基因表达的任何改变（增加/减少）都具有显著的病理生理后果。我们假设在正常生理中，hAT1受体的表达是通过葡萄糖和胰岛素对hAT1基因转录的规范化相互作用实现的。另外，在糖尿病中，当细胞外葡萄糖水平高而胰岛素水平低时，葡萄糖和胰岛素之间的平衡相互作用将发生改变，最终结果将是hAT1基因表达减少。最近，我们已经在hAT1基因启动子中确定了其基础转录所需的特定序列，并作为胰岛素反应（增强子）元件发挥作用。进一步的研究揭示了增强子上游的一个抑制元件，可以对正常/高水平的细胞外葡萄糖做出反应。我们的观察是，在葡萄糖存在的情况下（正常/高），胰岛素对hAT1转录抑制没有增强作用，而在没有葡萄糖或低糖存在的情况下，胰岛素会增强hAT1基因转录。此外，我们有证据表明这些调控元件识别葡萄糖和胰岛素诱导的特定核交易因子。我们的观察是hAT1基因转录的生理水平可能通过葡萄糖和胰岛素之间的相互作用受到抑制元件控制的第一个证据。虽然我们认识到葡萄糖和胰岛素都是hAT1转录的重要调节因子，但在本提案中，我们将主要关注葡萄糖介导的hAT1基因转录。因此，本申请的总体目标是确定葡萄糖在控制hAT1基因转录中的功能意义，并确定与葡萄糖信号相关的特定交易因子的作用，以了解hAT1基因在糖尿病、高血压等病理生理中的调控分子生化机制。

DESCRIPTION (provided by applicant): Hypertension and diabetes are two major risk factors in the pathogenesis of diabetic nephropathy. Angiotensin converting enzyme inhibitor therapy is broadly effective in patients with diabetic nephropathy suggesting an important role for renin angiotensin system in the progression of this disorder. Angiotensin II, the active component of the renin angiotensin system, acts primarily through angiotensin type 1 (AT1) receptors. Compared to rat in which two genes (AT1A, AT1B) code for AT1 receptor, in humans, a single gene code for angiotensin type 1 (hAT1), suggesting species specific expression and differential regulation of the receptor. In diabetes, from the onset to end-stage nephropathy, hAT1 receptor expression varies in different regions of the kidney with one exception, the proximal tubule in which, this receptor is down regulated at all times suggesting tubule specific regulation of the receptor. Reduction in hAT1 receptors could not be reversed by ACE inhibitors demonstrating that the receptor downregulation was not mediated by the up-regulation of angiotensin II. The molecular mechanisms leading to hAT1 receptor down-regulation in diabetes are not known. Any alterations (increase/decrease) in AT1 gene expression in proximal tubule have significant pathophysiological consequences. We hypothesize that in normal physiology, expression of the hAT1 receptor is achieved by normalized interactions between glucose and insulin on hAT1 gene transcription. Alternatively, in diabetes, when extracellular glucose levels are high and insulin levels are low, the equilibrium interaction between glucose and insulin will shift and the end-result will be decreased expression of hAT1 gene. Recently we have identified a specific sequence in the hAT1 gene promoter required for its basal transcription and functions as an insulin response (enhancer) element. Additional studies revealed a represser element upstream of the enhancer that can respond to normal/high levels of extracellular glucose. Our observation is that in the presence of glucose (normal/high), insulin has no enhancer effect on hAT1 transcriptional repression, where as in the absence of glucose or presence of low glucose insulin enhances the hAT1 gene transcription. In addition, we have evidence that these regulatory elements recognize specific nuclear transacting factors induced by glucose and insulin. Our observation is the first evidence that physiological levels of hAT1 gene transcription is controlled by a represser element perhaps through an interplay between glucose and insulin. Although we recognize that both glucose and insulin are important regulators of hAT1 transcription, in this proposal we will focus primarily on glucose mediated hAT1 gene transcription. Therefore, the overall goal of this application is to determine the functional significance of glucose in the control of hAT1 gene transcription and identify the role of specific transacting factors associated with glucose signaling in order to understand the molecular and biochemical mechanisms involved in the regulation of hAT1 gene in pathophysiology such as diabetes and hypertension. Non Technical description: Hormone angiotensin II through its receptor protein plays a very important role in the development of diabetes mediated kidney diseases. In this study we are proposing to identify the regulation of the receptor gene and factors that are controlled by high glucose in the kidney. With this information we are hoping to obtain a better understanding of the receptor action, and also a direction in the management and treatment of kidney diseases in diabetes.