Mechanisms of Hormonal Regulation of the Sodium Chloride Cotransporter

描述(申请人提供)：硫氮化物敏感的氯化钠共转运体(NCC)是盐平衡和全身血压的关键决定因素之一。然而，与其他盐平衡的重要影响因素相比，我们对激素和生理信号如何改变这种共转运蛋白的活性知之甚少。初步数据表明，ERK1/2 MAPK(细胞外信号调节蛋白激酶1和2丝裂原激活蛋白激酶)通路在NCC的泛素化和内吞作用中发挥作用。与-非赖氨酸激酶1和4(WNK1和WNK4)一起，Ste20相关的富含丙氨酸的丙氨酸激酶(SPAK)和氧化应激反应-1(OSR1)也是NCC的重要调节因子，有数据表明一些激素对NCC起调节作用。然而，激素与调节激素对NCC影响的细胞内途径之间的联系尚未建立。这项拟议研究的广泛假设是，NCC受到激素调节，激素调节通过调节ERK1/2、SPAK/OSR和WNKS来进行。目标1将通过激活ERK1/2 MAPK来确定NCC的激素调节。ERK1/2MAPK将被作为表皮生长因子(EGF)和钙敏感受体(CaSR)对NCC的激素/生理效应的中央中介。每种激素的ERK1/2激活机制以及泛素化和内吞作用在这一过程中的作用将被彻底研究，追踪从激素刺激到功能效应的途径。目的2研究醛固酮通过ERK1/2 MAPK和SPAK/OSR对NCC的激素调节作用及其机制。醛固酮对ERK1/2的抑制和Spak/OSR的激活可能导致NCC活性升高。WNKS在这些途径中的作用将得到彻底研究。目的3将确定血管紧张素II对NCC进行激素调节的潜在分子机制。将重点研究WNKS中每一条激酶通路的作用。将利用哺乳动物细胞培养、离体管微灌流和整体动物实验来检查细胞/分子、肾小管和机体水平的影响。对血压激素调节的了解在开发治疗高血压的新疗法方面发挥了关键作用，高血压是导致全球死亡率最大的单一危险因素。这些研究将为调节血压动态平衡的这一重要效应因子提供重要信息。 公共卫生相关性：氯化钠共转运体(NCC)是哺乳动物肾脏中一条重要的盐吸收途径，是最有效的抗高血压药物之一的作用部位，在高血压的遗传性疾病中发挥关键作用。尽管这种共转运蛋白在人类疾病中很重要，但我们对其激素调节知之甚少。对这一影响血压的关键因素的激素调节的研究将为高血压的发病机制提供宝贵的见解。

DESCRIPTION (provided by applicant): The thiazide-sensitive sodium chloride cotransporter (NCC) is one of the key determinants of salt balance and thus systemic blood pressure. However, in contrast to other important effectors of salt balance we know little about how hormonal and physiological signals alter activity of this cotransporter. Preliminary data demonstrates the role of the ERK1/2 MAPK (Extra-cellular signal-Regulated Kinases 1 and 2 Mitogen-Activated Protein Kinases) pathway in the ubiquitination and endocytosis of NCC. With-No-Lysine Kinase 1 and 4 (WNK1 and WNK4), Ste20-related proline alanine-rich kinase (SPAK) and Oxidative Stress Response-1 (OSR1) are also important regulators of NCC, and there is data that some hormones regulate NCC. However, a link between hormones and the intracellular pathways that mediate the effects of hormones on NCC has not been established. The broad hypothesis of the proposed investigation is that NCC is subject to hormonal regulation that proceeds through modulation of the kinases ERK1/2, SPAK/OSR and WNKs. Aim 1 will define the hormonal regulation of NCC through activation of ERK1/2 MAPK. ERK1/2 MAPK will be examined as a central mediator of the hormonal/physiological effects of Epidermal Growth Factor (EGF) and the Calcium-sensing Receptor (CaSR) on NCC. The mechanism of ERK1/2 activation and the roles of ubiquitination and endocytosis in this process will be thoroughly examined for each hormone, tracing a pathway from hormonal stimulus to functional effect. Aim 2 will determine the hormonal regulation of NCC by Aldosterone through ERK1/2 MAPK and SPAK/OSR and the mechanisms underlying these effects. ERK1/2 inhibition and SPAK/OSR activation by Aldosterone may lead to increased NCC activity. The role of WNKs in these pathways will be thoroughly examined. Aim 3 will determine the underlying molecular mechanisms mediating hormonal regulation of NCC by Angiotensin II. The roles of each of these kinase pathways will be examined with a particular emphasis on WNKs. Mammalian cell culture, isolated tubule microperfusion and whole animal experimentation will be utilized to examine effects at the cellular/molecular, tubular and organismal level. Knowledge of hormonal regulation of blood pressure has played a critical role in the development of new therapies to treat hypertension, the single risk factor that contributes the most to worldwide mortality. These studies will provide vital information on the regulation of this important effector of blood pressure homeostasis. PUBLIC HEALTH RELEVANCE: The sodium chloride cotransporter (NCC) is an important salt absorptive pathway in the mammalian kidney that is the site of action of one of the most effective classes of anti- hypertensive medications and plays a key role in genetic disorders of hypertension. Despite the importance of this cotransporter in human disease, we know little about its hormonal regulation. This investigation of the hormonal regulation of this key effector of blood pressure will provide invaluable insight into the pathogenesis of Hypertension.