Regulation of ENaC by Casein Kinase 2

高血压影响着美国和全世界数以百万计的人。更好地理解 高血压的分子和细胞起源将提高治疗和诊断的效果。这个 上皮性钠离子通道(ENaC)是肾脏钠离子排泄的最终仲裁者。因此，自由裁量控制 ENaC微调肾脏排泄。适当的肾脏排泄是正常调节的关键因素。 动脉血压。因此，ENaC及其上游调节因子的功能障碍导致调节失调 由异常排泄引起的血压升高。酪蛋白激酶2(CK2)具有磷酸化ENaC的功能。这个 然而，这在生理上的重要性是模糊的。我们的初步结果表明，磷酸化 CK2是正常的ENaC活动和肾脏Na+排泄所必需的。CK2内的磷酸化位点 ENAC位于一个典型的“锚定”锚定蛋白结合基序中。ENaC中的此站点与CK2有相似之处 不相关的NAV和KCNQ通道中的磷酸化位点，也位于“锚”基序中。 CK2对NAV和KCNQ通道的磷酸化作用是有利于结合的分子“开关” Anyrin-3(Ank-3)。Ank-3的结合促进了这些通道的适当膜定位 增加它们的活跃度。主细胞(PC)Ank-3的靶向缺失显著降低ENaC 我们的PC-Ank-3 KO小鼠的活动。考虑到我们强劲的初步结果和可能的 CK2对ENaC、NAV和KCNQ的收敛进化塑造调节，我们提出检验的前提是 锚定基序中CK2对ENaC的磷酸化是Ank-3与 通道，这是正常的通道位置和功能所必需的，以及肾脏Na+的适当调节 排泄物。我们将使用包括新思维和工具的多学科方法来测试这些想法， 包括PC特有的CK2和Ank-3KO小鼠，以及与一项研究相结合的高度严谨 设计范围广泛，对分子和细胞机制以及整体提出问题 动物生理后果。以下具体目标将用来检验我们的想法：1)确定 CK2调节ENaC活性的细胞和分子机制；2)定量 CK2调节ENaC的生理功能；3)确定CK2对ENaC的调节是否 保存在人类体内。如果CK2对ENaC的调节具有临床和生理重要性 监管必须在整个门类中保持保守，特别是在人类身上。我们在量化ENaC方面的开创性努力 健康人体肾脏小管中的活动使我们能够在最相关的环境中检验我们的想法 可能：天然集合管内的人类主细胞。在实现这些目标之后，我们将 了解ENaC的CK2磷酸化是否、如何以及何时起“开关”作用，促进Ank-3结合增加 渠道活动包括对调解这一法规的机制有详细的了解，以及 对这种监管的生理后果有丰富的认识。

Hypertension affects millions of people in the United States and worldwide. Improved understanding of the molecular and cellular origins of hypertension will improve the efficacy of treatment and diagnosis. The Epithelial Na+ Channel, ENaC, is the final arbiter of Na+ excretion in the kidneys. As such, discretionary control of ENaC fine-tunes renal excretion. Appropriate renal excretion is a key factor in the normal regulation of arterial blood pressure. Consequently, dysfunction of ENaC and its upstream modulators cause dysregulation of blood pressure due to abnormal excretion. Casein kinase 2 (CK2) is known to phosphorylate ENaC. The physiological importance of this, though, is obscure. Our preliminary results demonstrate that phosphorylation by CK2 is necessary for normal ENaC activity and renal Na+ excretion. The CK2 phosphorylation site within ENaC resides within a canonical “anchor” ankyrin binding motif. This site in ENaC shares similarity to CK2 phosphorylation sites in the unrelated NaV and KCNQ channels, which also lie within “anchor” motifs. Phosphorylation of NaV and KCNQ channels by CK2 acts as a molecular “switch” favoring the binding of ankyrin-3 (Ank-3). The binding of Ank-3 facilitates the proper membrane localization of these channels increasing their activity. The targeted deletion of Ank-3 in principal cells (PC) significantly decreased ENaC activity in our PC-Ank-3 KO mouse. In consideration of our strong preliminary results and the possible convergent evolution shaping regulation of ENaC, NaV and KCNQ by CK2, we propose testing the premise that phosphorylation of ENaC by CK2 within “anchor” motifs is necessary and sufficient for Ank-3 binding to the channel, which is required for normal channel locale and function, and the proper regulation of renal Na+ excretion. We will test these ideas using a multidisciplinary approach that includes novel thinking and tools, including PC-specific CK2 and Ank-3 KO mice, and a high degree of rigor in conjunction with a research design that is broad in scope asking questions about molecular and cellular mechanisms as well as whole animal physiological consequences. The following specific aims will be used to test our ideas: 1) To determine the cellular and molecular mechanisms of CK2 regulation of ENaC activity; 2) To quantify the physiological function of CK2 regulation of ENaC; and 3) To determine if CK2 regulation of ENaC is conserved in humans. If CK2 regulation of ENaC is to be of clinic and physiological importance such regulation must be conserved across phyla particularly in humans. Our pioneering efforts to quantify ENaC activity in tubules from healthy donor human kidneys allows us to test our ideas in the most relevant setting possible: the human principal cell within the native collecting duct. After accomplishing these aims, we will know if, how and when CK2 phosphorylation of ENaC functions as a “switch” to favor Ank-3 binding to increase channel activity to include having a detailed understanding of the mechanisms mediating this regulation, and a rich appreciation of the physiological consequences of such regulation.