RENIN GENE EXPRESSION IN INTRARENAL ARTERIOLES

肾内小动脉中肾素基因的表达

• 批准号: 3840393
• 负责人: R ARIEL GOMEZ
• 金额: --
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3840393
• 关键词: angiotensin II; arterioles; calcium flux; electron microscopy; gene expression; genetic promoter element; genetic regulatory element; homeostasis; kidney circulation; laboratory rabbit; laboratory rat; molecular cloning; plaque assay; posttranslational modifications; protein transport; renin; tissue /cell culture; transcription factor; transfection

As the first and rate limiting of the renin angiotensin system, renin has an important role in control of cardiovascular homeostasis. Definition of the molecular mechanisms regulating renin gene expression and the subsequent intracellular events of synthesis, sorting, trafficking, processing, storage, and finally release of the active renin is essential for understanding the renin angiotensin system. Most of the molecular mechanisms regulating intracellular handling of renin proximal to its secretion remain to be defined. In the normal adult kidney, renin is synthesized by the juxtaglomerular (JG) cells. However, during early development, renin is expressed in arcuate and interlobular arteries. If an adult animal is subjected to converting enzyme inhibition, there is a recruitment of renin gene expressing and releasing cells along the preglomerular vessels, resembling the situation in the immature animal. We hypothesize that the ability of smooth muscle cells (SMCs) from the renal arterial tree to turn on and off the renin gene is mediated by tissue- specific transacting factors binding to the 5' flanking region of the renin gene. We propose to isolate and characterize those factors and to identify the Cis regulatory sequences involved in the response. Cotransfection studies of native and mutated renin promoter expression constructs with transacting factors will allow us to assess the functional significance of sequences involved. The molecular mechanisms responsible for the intracellular processing of renin have not been investigated in detail. Mutations of the pre-, pro-, and renin sequences of the renin gene will be produced and introduced into AT-T20 cells and, using EM microscopy, the intracellular trafficking of renin will be investigated. These studies will determine whether specific sequences within the renin gene are responsible for intracellular sorting of the renin molecule. Similarly, using antibodies directed against specific domains (pre-, pro-, renin) of the molecule, we will determine the specific organelles where renin is processed. Additional studies will examine the relationship of renin gene transcription (measured by intervening sequence probes) to renin release from individual cells detected by the reverse hemolytic plaque assay. Developmental differences in response to angiotensin II, and changes in the calcium concentration will be evaluated. The proposed experiments will increase our understanding of the molecular processes regulating renin biosynthesis, processing, and release.

作为肾素血管紧张素系统的第一个和限速因子，肾素具有 在控制心血管内稳态方面起着重要作用。定义 肾素基因表达调控的分子机制 随后的细胞内合成、分类、贩运、 活性肾素的加工、储存和最终释放是必不可少的 了解肾素血管紧张素系统。大多数分子 调节ITS近端肾素细胞内处理的机制 分泌物的定义尚不明确。在正常成人肾脏中，肾素是 由肾小球旁(JG)细胞合成。然而，在早期， 在发育过程中，肾素在弓状动脉和小叶间动脉中表达。如果 成年动物受到转化酶的抑制，有一种 肾素基因表达和释放细胞沿血管的募集 肾小球前血管，类似于未成熟动物的情况。我们 假设肾脏的平滑肌细胞(SMC)的能力 开启和关闭肾素基因的动脉树是由组织介导的 与肾素5‘侧翼区结合的特异性交易因子 吉恩。我们建议分离和表征这些因素，并确定 参与反应的顺式调控序列。共转染 天然和突变肾素启动子表达载体的研究 交易因素将使我们能够评估 涉及的序列。导致这一现象的分子机制 肾素在细胞内的处理还没有详细的研究。 肾素基因的前、前和肾素序列的突变将是 产生并导入AT-T20细胞，并使用EM显微镜观察 将对肾素在细胞内的运输进行调查。这些研究 将决定肾素基因内的特定序列是否 负责肾素分子的细胞内分选。同样， 使用针对特定结构域(前、前、肾素)的抗体 分子，我们将确定肾素所在的特定细胞器 已处理。更多的研究将检查肾素基因之间的关系 转录(通过插入序列探针测量)以释放肾素 从反向溶血空斑试验检测到的单个细胞。 血管紧张素II反应的发育差异，以及 将对钙浓度进行评估。拟议中的实验将 加深对肾素调节分子过程的了解 生物合成、加工和释放。