Regulation of NKCC2 and renal NaCl transport by protein-protein interactions

摘要 在肾脏，Henle环的粗大升支(TAL)对氯化钠稳态和血液至关重要。 压力调节。在人类和盐敏感型高血压动物模型中，盐的吸收是异常的 在TAL中增加，其中氯化钠的吸收取决于肾脏转运体NKCC2，一个顶端的Na/K/2Cl- 联运公司。我们发现，NKCC2在TAL顶膜上的存在控制了食盐的吸收 在这个上皮组织中。控制顶膜NKCC2水平的分子机制包括内吞作用， 回收和胞外插入。抑制内吞检索导致NKCC2在膜上积聚 增加了对氯化钠的吸收。任何影响NKCC2内吞作用的基因或蛋白都可能影响 NKCC2的活性和肾脏的盐分转运，但只有少数已知的蛋白质结合NKCC2。使用目标 蛋白质组学筛选，我们发现ALMS1(阿尔斯特伦综合征1)和ACTN4(α-肌动蛋白4)相互作用 NKCC2的合作伙伴。我们还发现ALMS1和ACTN4相互作用，增加了 这些蛋白质形成一个复合体。ALMS1和ACTN4的单核苷酸多态与 高血压和肾功能减退。我们发现ALMS1基因敲除的大鼠有更高的表面NKCC2 还有高血压。我们发现ACTN4，一种与足细胞生物学有关的蛋白质，也有表达 整个肾单位，包括TAL。ALMS1和ACTN4在TAL处理肾脏氯化钠中的作用 它们在血压调节中的作用尚不清楚。我们假设ALMS1控制表面NKCC2 NKCC2和ACTN4的羧基末端结合介导NKCC2的水平和NKCC2介导的盐吸收 NKCC2从顶膜内吞。TAL中ALMS1或ACTN4表达的降低 增加表面NKCC2、NKCC2介导的氯化钠重吸收、肾小管肾小球反馈(TGF)敏感性 并导致盐敏性高血压。我们的长期目标是增加我们对 ALMS1和ACTN4在肾脏盐转运中的作用。

Abstract In the kidney, the thick ascending limb (TAL) of the loop of Henle is critical for NaCl homeostasis and blood pressure regulation. In humans and animal models of salt-sensitive hypertension, NaCl absorption is abnormally increased in the TAL, where NaCl absorption depends on the renal transporter NKCC2, an apical Na+/K+/2Cl- co-transporter. We showed that the presence of NKCC2 at the TAL apical membrane controls NaCl absorption in this epithelium. The molecular mechanisms that control apical membrane NKCC2 levels involve endocytosis, recycling and exocytic insertion. Inhibition of endocytic retrieval causes NKCC2 accumulation at the membrane and increased NaCl absorption. Any gene or protein affecting NKCC2 endocytosis could potentially influence NKCC2 activity and renal salt transport but only few proteins are known to bind NKCC2. Using a targeted proteomics screen, we identified ALMS1 (Alström Syndrome 1) and ACTN4 (alpha-actinin 4) as interacting partners of NKCC2. We also found that ALMS1 and ACTN4 interact with each other, raising the possibility that these proteins form a complex. Single nucleotide polymorphisms in ALMS1 and ACTN4 are associated with hypertension and decreased kidney function. We found that ALMS1 knockout rats have higher surface NKCC2 and high blood pressure. We found that ACTN4, a protein involved in podocyte biology, is also expressed throughout the nephron, including the TAL. The roles of ALMS1 and ACTN4 in renal NaCl handling by the TAL and their role in blood pressure regulation are unknown. We hypothesize that ALMS1 controls surface NKCC2 levels and NKCC2-mediated NaCl absorption by binding the carboxyl-terminus of NKCC2 and ACTN4 to mediate NKCC2 endocytosis from the apical membrane. A decrease in ALMS1 or ACTN4 expression in the TAL increases surface NKCC2, NKCC2-mediated NaCl reabsorption, tubulo-glomerular feedback (TGF) sensitivity and leads to salt-sensitive hypertension. Our long-term goal is to increase our understanding of the role of ALMS1 and ACTN4 in kidney NaCl transport.