RENIN PROCESSING IN RENAL DEVELOPMENT

作为肾素-血管紧张素系统的第一限速酶， 肾素在心血管内稳态的控制中起着重要作用。 肾素基因表达调控的分子机制研究进展 以及随后的细胞内合成、分类、 活性肾素的贩运、加工、储存和最终释放 对于理解肾素-血管紧张素系统是至关重要的。大多数 调节肾素近端细胞内处理的分子机制 其分泌部位尚待确定。在正常的成人肾脏中， 肾素由肾小球旁(JG)细胞合成。然而，在 早期发育，肾素在弓形和小叶间表达 动脉。如果成年动物受到转化酶的影响 抑制肾素基因的表达和释放 沿着肾小球前血管的细胞，类似于 幼小的动物。我们假设，平滑肌细胞的能力 从肾动脉树(SMC)中开启和关闭肾素基因 由与5‘端结合的组织特异性横切因子介导 肾素基因的侧翼区域。我们建议隔离和 鉴定这些因子并鉴定顺式调节序列 参与了回应。天然基因和突变基因的共转染研究 带有横切因子的肾素启动子表达载体将允许 US来评估所涉及序列的功能意义。这个 细胞内加工的分子机制 肾素尚未得到详细研究。前或前的突变 将产生肾素基因的前-和肾素序列，并 导入AT-T20细胞，用电子显微镜观察细胞内 贩卖或肾素将被调查。这些研究将确定 肾素基因内的特定序列是否与 肾素分子的细胞内分选。同样，使用抗体 针对分子的特定结构域(前、前、肾素)，我们 将确定处理肾素的特定细胞器。 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。 导致肾素释放的细胞膜和胞浆事件将是 血管紧张素II、β受体激动剂和 拮抗剂和钙浓度的变化。建议数 实验将增加我们对分子过程的理解。 调节肾素的生物合成、运输和释放。

As the first and rate limiting enzyme of the renin-angiotensin system, renin has an important role in the 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 site 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 transecting 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. Co-transfection studies of native and mutated renin promoter expression constructs with transecting 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-, or pro- and renin sequences of the renin gene will be produced and introduced into AT-T20 cells and, using EM microscopy, the intracellular trafficking or 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. The cell membrane and cytosolic events leading to renin release will be investigated in response to angiotensin II, beta-receptor agonists and antagonists and changes in the calcium concentration. The proposed experiments will increase our understanding of the molecular processes regulating renin biosynthesis, trafficking and release.