Soluble (pro)renin receptor regulation of ENaC involved in aldosterone signaling in cultured collecting duct cells

We have previously shown that activation of (pro)Renin receptor (PRR) induces epithelial Na⁺ channel (ENaC) activity in cultured collecting duct cells. Here, we examined the role of soluble PRR (sPRR), the cleavage product of PRR in ENaC regulation, and further tested its relevance to aldosterone signaling. In cultured mpkCCD cells, administration of recombinant histidine-tagged sPRR (sPRR-His) at 10 nM within minutes induced a significant and transient increase in the amiloride-sensitive short-circuit current as assessed using the Ussing chamber technique. The acute ENaC activation was blocked by the NADPH Oxidase 1/4 inhibitor GKT137892 and siRNA against Nox4 but not the β-catenin Inhibitor ICG-001. In primary rat inner medullary collecting duct cells, administration of sPRR-His at 10 nM for 24 h induced protein expression of the α-subunit but not β- or γ-subunits of ENaC, in parallel with upregulation of mRNA expression as well as promoter activity of the α-subunit. The transcriptional activation of α-ENaC was dependent on β-catenin signaling. Consistent results obtained by epithelial volt ohmmeter measurement of equivalent current and Ussing chamber determination of short-circuit current showed that aldosterone-induced transepithelial Na⁺ transport was inhibited by the PRR decoy inhibitor PRO20 and PF-429242, an inhibitor of sPRR-generating Enzyme site-1 protease, and the response was restored by the addition of sPRR-His. Medium sPRR was elevated by aldosterone and inhibited by PF-429242. Taken together, these results demonstrate that sPRR induces two phases of ENaC activation via distinct mechanisms and functions as a mediator of the natriferic action of aldosterone.