BACKGROUND The renal epithelial sodium channel (ENaC) is essential for sodium balance and blood pressure control. ENaC undergoes complex proteolytic activation by not yet clearly identified tubular proteases. Here, we examined a potential role of transmembrane serine protease 2 (TMPRSS2). METHODS Murine ENaC and TMPRSS2 were (co-)expressed in Xenopus laevis oocytes. ENaC cleavage and function were studied in TMPRSS2-deficient murine cortical collecting duct (mCCDcl1) cells and TMPRSS2-knockout (Tmprss2-/-) mice. Short-circuit currents (ISC) were measured to assess ENaC-mediated transepithelial sodium transport of mCCDcl1 cells. The mCCDcl1 cell transcriptome was studied using RNA sequencing. The effect of low-sodium diet with or without high potassium were compared in Tmprss2-/- and wildtype mice using metabolic cages. ENaC-mediated whole-cell currents were recorded from microdissected tubules of Tmprss2-/- and wildtype mice. RESULTS In oocytes, co-expression of murine TMPRSS2 and ENaC resulted in fully cleaved γ-ENaC and ∼2-fold stimulation of ENaC currents. High baseline expression of TMPRSS2 was detected in mCCDcl1 cells without a stimulatory effect of aldosterone on its function or transcription. TMPRSS2 knockout in mCCDcl1 cells compromised γ-ENaC cleavage and reduced baseline and aldosterone-stimulated ISC which could be rescued by chymotrypsin. A compensatory transcriptional upregulation of other proteases was not observed. Tmprss2-/- mice kept on standard diet exhibited no apparent phenotype, but renal γ-ENaC cleavage was altered. In response to a low-salt diet, particularly with high potassium intake, Tmprss2-/- mice increased plasma aldosterone significantly more than wildtype mice to achieve a similar reduction of renal sodium excretion. Importantly, the stimulatory effect of trypsin on renal tubular ENaC currents was much more pronounced in Tmprss2-/- mice than that in wildtype mice. This indicated the presence of incompletely cleaved and less active channels at the cell surface of TMPRSS2-deficient tubular epithelial cells. CONCLUSIONS TMPRSS2 contributes to proteolytic ENaC activation in mouse kidney in vivo.

中文翻译：

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小鼠肾脏中跨膜丝氨酸蛋白酶 2 和上皮钠通道的蛋白水解激活。


背景 肾上皮钠通道 （ENaC） 对于钠平衡和血压控制至关重要。ENaC 通过尚未明确鉴定的肾小管蛋白酶进行复杂的蛋白水解激活。在这里，我们检查了跨膜丝氨酸蛋白酶 2 （TMPRSS2） 的潜在作用。方法 小鼠 ENaC 和 TMPRSS2 在非洲爪蟾卵母细胞中 （共） 表达。在 TMPRSS2 缺陷小鼠皮质集合管 （mCCDcl1） 细胞和 TMPRSS2 敲除 （Tmprss2-/-） 小鼠中研究 ENaC 切割和功能。测量短路电流 （ISC） 以评估 ENaC 介导的 mCCDcl1 细胞跨上皮钠转运。使用 RNA 测序研究 mCCDcl1 细胞转录组。使用代谢笼比较了低钠饮食加或不加高钾的 Tmprss2-/- 和野生型小鼠的效果。从 Tmprss2-/- 和野生型小鼠的显微解剖小管中记录 ENaC 介导的全细胞电流。结果 在卵母细胞中，小鼠 TMPRSS2 和 ENaC 的共表达导致 γ-ENaC 完全裂解和 ENaC 电流的 ∼2 倍刺激。在 mCCDcl1 细胞中检测到 TMPRSS2 的高基线表达，醛固酮对其功能或转录没有刺激作用。mCCDcl1 细胞中的TMPRSS2敲除损害了 γ-ENaC 切割，降低了基线和醛固酮刺激的 ISC，这可以通过胰凝乳蛋白酶来挽救。未观察到其他蛋白酶的代偿性转录上调。标准饮食中的 Tmprss2-/-小鼠没有表现出明显的表型，但肾 γ-ENaC 切割发生了改变。作为对低盐饮食的反应，特别是高钾摄入量，Tmprss2 - / - 小鼠比野生型小鼠显着增加血浆醛固酮，以实现类似的肾钠排泄减少。 重要的是，胰蛋白酶对肾小管 ENaC 电流的刺激作用在 Tmprss2 - / - 小鼠中比在野生型小鼠中更明显。这表明 TMPRSS2 缺陷肾小管上皮细胞的细胞表面存在未完全裂解且活性较低的通道。结论 TMPRSS2 有助于小鼠体内小鼠肾脏的蛋白水解 ENaC 激活。