Regulation of ENaC by phosphatidylinositide 3-kinase and phospholipids

描述（由申请人提供）： 在远端肾单位受调节的Na+重吸收，部分地控制人类和其他陆生脊椎动物的血压。醛固酮敏感性上皮Na+通道（ENaC）的活性限制了Na+的转运。ENaC的功能障碍和不适当的调节导致血压紊乱和肾脏、结肠和肺的盐处理不当。该提案是R 01研究ENaC的醛固酮调节的竞争性更新，该研究在其最初的资助期间取得了很大的成功。来自该初始资助期的结果以及更新的初步结果强烈表明，磷脂激酶，包括磷脂酰肌醇3-激酶（PI 3-K）及其产物磷脂酰肌醇是ENaC活性的关键决定因素。增加ENaC活性的两种生理学上重要的激素，醛固酮和胰岛素，增加远端肾单位上皮中P3-K活性及其产物磷脂酰肌醇3，4，5-三磷酸（PI（3，4，5）P3）的合成。支持这种增加的细胞机制仍然未知。此外，PI 3-K在多大程度上是醛固酮和胰岛素对ENaC作用的主要仲裁者，仍有待研究。此外，PI（3，4，5）P3调节ENaC的可能机制和生理后果尚未完全理解。类似地，ENaC内响应PI（3，4，5）P3信号传导的推定分子决定簇仍有待鉴定。PI 3-K及其磷脂酰肌醇产物是ENaC调节的核心的一般假设统一了该提议的实验和想法。这条调查路线是我早期研究的逻辑延伸，我希望测试它会产生新颖而重要的发现。在此，我提出了三个具体的目标：1）描绘并分配针对ENaC的PI 3-K和PI（3，4，5）P3信号通路的生理意义; 2）确定PI（3，4，5）P3调节ENaC的细胞机制; 3）建立PI（3，4，5）P3调节的ENaC内的分子决定簇。我通过由互补实验组成的全面实验设计来测试我的假设，以提供对PI 3-K从分子到整个组织对ENaC调节的综合理解。在人类和其他陆生脊椎动物中，远端肾肾单位的适当Na+重吸收对于调节全身Na+平衡以及血压至关重要。醛固酮敏感性上皮Na+通道（ENaC）的活性限制了Na+的重吸收。目前的建议继续我们的调查控制ENaC活性的细胞和分子机制，以响应醛固酮。

DESCRIPTION (provided by applicant): Regulated Na+ reabsorption at the distal nephron, in part, controls blood pressure in humans and other terrestrial vertebrates. Activity of the aldosterone-sensitive epithelial Na+ channel (ENaC) is limiting for Na+ transport here. Dysfunction and inappropriate regulation of ENaC result in blood pressure disorders and improper salt handling by the kidney, colon and lungs. This proposal is a competitive renewal of an R01 investigating aldosterone regulation of ENaC that has had much success during its initial funding period. Results from this initial funding period, as well as, newer preliminary results strongly suggest that phospholipid kinases, including phosphatidylinositide 3-kinase (PI3-K), and their product phosphatidylinositides are critical determinants of ENaC activity. Two physiologically important hormones that increase ENaC activity, aldosterone and insulin, increase P3-K activity and synthesis of its product phosphatidylinositol, phosphatidylinositol 3,4,5-trisphosphate (PI(3,4,5)P3), in distal nephron epithelia. The cellular mechanism underpinning this increase remains unknown. The extent to which PI3-K is the primary arbiter of aldosterone and insulin actions on ENaC, in addition, remains unexplored. Moreover, possible mechanisms and physiological consequences of PI(3,4,5)P3 modulation of ENaC are not fully appreciated. Putative molecular determinants within ENaC responsive to PI(3,4,5)P3 signaling, similarly, remain to be identified. The general hypothesis that PI3-K and its phosphatidylinositide products are central to regulation of ENaC unites the experiments and ideas of this proposal. This line of inquiry is a logical extension of my earlier studies, and I expect testing it to result in novel and significant findings. Here, I address three specific aims: 1) Delineate and assign physiological significance to PI3-K and PI(3,4,5)P3 signaling pathways targeting ENaC; 2) Determine the cellular mechanism of PI(3,4,5)P3 regulation of ENaC; and 3) Establish the molecular determinants within ENaC of PI(3,4,5)P3 regulation. I test my hypotheses with a comprehensive experimental design structured with complementary experiments to provide an integrative understanding of ENaC regulation by PI3-K from the molecule to the whole tissue. Proper Na+ reabsorption at the distal renal nephron in humans and other terrestrial vertebrates is central to regulation of systemic Na+ balance and thus, blood pressure. Activity of the aldosterone-sensitive epithelial Na+ channel (ENaC) is limiting for Na+ reabsorption here. The current proposal continues our investigation of the cellular and molecular mechanisms controlling ENaC activity in response to aldosterone.