Lipid rafts, dopamine 1 receptor, and hypertension

项目摘要 D1R在血压调节中起重要作用。DRD1 小鼠生殖系缺失会导致高血压。G蛋白偶联受体激酶4型 (GRK4)在D1R的正常细胞循环和功能中起重要作用。然而，一些 回收的D1R必须针对脂筏才能发挥作用。因此，突变体D1Rs 347C&gt；A和 351C&gt；A仍然以质膜为靶标，但不能在脂筏中定位；这些突变 阻止D1R介导的肾近端小管cAMP生成增加 细胞(RPTCs)和损害(如347C和gt；A)小鼠正常的血压调节。D1R中的C347和C351 是棕榈酰化位点；SNX19作为D1R棕榈酰化的支架蛋白。 高尔基人，通过高尔基人特有的锌指蛋白。脂筏也是需要的适当的 腺酰环化酶5和6的膜分布和维持以及D1R的调节。 棕榈酰化的抑制或分子生物学干扰可防止D1R靶向脂筏， 损害D1R功能，并导致高血压。沉默SNX19损害D1R介导的信号转导 RPTCs内cAMP含量增加，Na转运减少，但远曲小管上皮细胞内cAMP含量增加，Na转运减少。 肾SNX19对血压的调节很重要；肾限制的SNX19沉默增加 C57BL/6小鼠血压正常但不含低盐饮食。小鼠中SNX19的种系缺失 在正常食盐饮食中增加血压。SNX19位于D1R的上游。SNX19 rs2298566已关联 人类排钠和高血压能力降低，人类D1R功能受损 (H)RPTCs。高血压患者hRPTCs中SNX19蛋白表达降低。我们将测试 总体假设SNX19在D1R向RPT中的脂筏运输中起重要作用 正常D1R功能的质膜。肾脏D1R功能受损，引起 由于其在RPT质膜上向脂筏的转运受阻，导致对盐敏感 高血压。特异靶1将测试SNX19将D1R靶向到脂质中的假设 RPTC质膜筏对正常的D1R功能至关重要。SNX19和GRK4相互作用 在脂筏中调节D1R功能，包括D1R介导的对Na转运的抑制 RPTCS。特定目标2将测试假设，棕榈酰化位点的突变在 D1R基因阻止SNX19靶向RPTC血浆中脂筏的能力 薄膜。C57BL/6小鼠SNX19生殖系缺失或肾脏限制性缺失导致盐中毒 敏感性高血压。突变DRD1阻止D1R靶向RPTC血浆脂筏 膜也会引起盐敏感型高血压。这些基因及其蛋白质可能是 治疗人类原发性高血压的靶点。

Project Summary The D1 dopamine receptor (D1R) is important in the regulation of blood pressure (BP). Drd1 germline deletion in mice causes hypertension. G protein-coupled receptor kinase type 4 (GRK4) is important in the normal cellular recycling and function of D1R. However, some of the recycled D1R has to be targeted to lipid rafts to be functional. Thus, mutant D1Rs 347C>A and 351C>A still target to the plasma membrane but fail to localize in lipid rafts; these mutations prevent the increase in D1R-mediated stimulation of cAMP production in renal proximal tubule cells (RPTCs) and impair (e.g., 347C>A) normal BP regulation in mice. C347 and C351 in D1R are palmitoylation sites; SNX19 functions as a scaffold protein for the palmitoylation of D1R at the Golgi, via Golgi-specific zinc finger protein. Lipid rafts are also needed for the proper membrane distribution and maintenance of adenylyl cyclases 5 and 6 and regulation by D1R. Inhibition or molecular biological disruption of palmitoylation prevents D1R targeting to lipid rafts, impairs D1R function, and causes hypertension. Silencing SNX19 impairs D1R-mediated increase in cAMP and decrease in Na+ transport in RPTCs but not distal convoluted tubule cells. Renal SNX19 is important in the regulation of BP; renal-restricted silencing of Snx19 increases BP in C57Bl/6 mice on normal but not low salt diet. Germline deletion of SNX19 in mice also increases BP on normal salt diet. SNX19 is upstream of D1R. SNX19 rs2298566 is associated with decreased ability to excrete Na+ and high BP in humans and impairs D1R function in human (h)RPTCs. SNX19 protein is decreased in hRPTCs from hypertensive humans. We will test the overall hypothesis that SNX19 is important in the trafficking of D1R to lipid rafts in the RPT plasma membrane for normal D1R function. Impaired functioning of D1R in the kidney, caused by its impaired trafficking to lipid rafts in RPT plasma membranes, causes salt-sensitive hypertension. Specific Aim 1 will test the hypothesis that SNX19 targeting of D1R into the lipid rafts of RPTC plasma membrane is crucial for normal D1R function. SNX19 and GRK4 interact in lipid rafts to regulate D1R function, including D1R-mediated inhibition of Na+ transport in RPTCs. Specific aim 2 will test the hypothesis that mutations of the palmitoylation sites in the D1R gene prevent the ability of SNX19 to target the D1R to lipid rafts in the RPTC plasma membrane. Germline deletion or renal-restricted deletion of SNX19 in C57Bl/6 mice causes salt- sensitive hypertension. Mutating DRD1 to prevent D1R targeting to lipid rafts of RPTC plasma membrane also causes salt-sensitive hypertension. These genes and their proteins could be targets in the treatment of human essential hypertension.