Journal of the American Society of Nephrology : JASN · 2024
Transmembrane Serine Protease 2 and Proteolytic Activation of the Epithelial Sodium Channel in Mouse Kidney.
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Sure Florian, Afonso Sara, Essigke Daniel, Schmidt Paul, Kalo M Zaher, Nesterov Viatcheslav, Kißler Alicia, Bertog Marko, Rinke Ralf, Wittmann Sabine, Broeker Katharina A E, Gramberg Thomas, Artunc Ferruh, Korbmacher Christoph, Ilyaskin Alexandr V
Аннотация
KEY POINTS: Proteolytic activation of the epithelial sodium channel (ENaC) was compromised by transmembrane serine protease 2 deficiency in murine cortical collecting duct cells and native mouse kidney. To compensate for impaired ENaC activation, rise in plasma aldosterone in response to low-salt diet was enhanced in Tmprss2−/− mice. Transmembrane serine protease 2 may be a potential drug target to limit proteolytic ENaC activation in disorders with increased renal ENaC activity.
BACKGROUND: The renal epithelial sodium channel (ENaC) is essential for sodium balance and BP 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 wild-type mice using metabolic cages. ENaC-mediated whole-cell currents were recorded from microdissected tubules of Tmprss2−/− and wild-type mice.
RESULTS: In oocytes, coexpression of murine TMPRSS2 and ENaC resulted in fully cleaved γ-ENaC and approximately two-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 wild-type 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 wild-type 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.
BACKGROUND: The renal epithelial sodium channel (ENaC) is essential for sodium balance and BP 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 wild-type mice using metabolic cages. ENaC-mediated whole-cell currents were recorded from microdissected tubules of Tmprss2−/− and wild-type mice.
RESULTS: In oocytes, coexpression of murine TMPRSS2 and ENaC resulted in fully cleaved γ-ENaC and approximately two-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 wild-type 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 wild-type 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.