RENIN GENE EXPRESSION IN INTRARENAL ARTERIOLES

肾内小动脉中肾素基因的表达

• 批准号: 3776793
• 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/3776793
• 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速率限制， 在控制心血管稳态中起重要作用。 定义 调节肾素基因表达的分子机制和 随后的细胞内合成，分选，运输， 活性肾素的加工、储存和最终释放是必需的 来了解肾素血管紧张素系统。 大多数分子 调节细胞内处理邻近其的肾素的机制 分泌物仍有待确定。 在正常成人肾脏中， 由肾小球（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抑制， 分泌肾素基因的细胞沿着 肾小球前血管，类似于未成熟动物的情况。 我们 假设来自肾脏的平滑肌细胞（SMC）的能力 动脉树开启和关闭肾素基因是由组织介导的， 与肾素5 '侧翼区结合的特异性反式作用因子 基因 我们建议隔离和描述这些因素，并确定 参与应答的顺式调节序列。 共转染 天然和突变的肾素启动子表达构建体的研究 交易因素将使我们能够评估的功能意义， 所涉及的序列。 负责的分子机制 还没有详细研究过肾素的细胞内加工。 肾素基因的前体、原体和肾素序列的突变将是可能的。 产生并引入AT-T20细胞，并使用EM显微镜， 将研究肾素的细胞内运输。 这些研究 将决定肾素基因中的特定序列是否 负责肾素分子的细胞内分选。 同样地， 使用针对特定结构域（前、原、肾素）的抗体， 分子，我们将确定特定的细胞器，其中肾素是 已处理。 进一步的研究将探讨肾素基因与 转录（通过插入序列探针测量）至肾素释放 来自通过反向溶血空斑试验检测的单个细胞。 对血管紧张素II反应的发育差异， 将评估钙浓度。 拟议的实验将 增加我们对调节肾素的分子过程的理解 生物合成、加工和释放。