Ankyrin 3 is regulated by aldosterone and mircoRNAs, and alters ENaC‐mediated sodium transport

Abstract

The role of microRNAs (miRs) in the physiologic regulation of sodium transport has yet to be fully appreciated. miRs are small non‐coding RNAs that selectively target and downregulate mRNAs to inhibit protein expression. Our previous studies indicated that aldosterone altered miR expression to directly modulate Na+ transport via the epithelial sodium channel (ENaC). In silico predicted protein targets were screened in cultured mouse cortical collecting duct (mCCD) cells to test for their involvement in ENaC regulation. One of the candidate miR‐regulated targets was ankyrin 3 (ank3). Ank3 expression increased in mCCD cells stimulated with 50nM aldosterone for 24hrs. By inhibiting the expression of miRs directly we increased ank3 protein levels indicating that ank3 was a miR target. To demonstrate the regulation of ank3 by miRs we transiently transfected the 3’‐untranslated region (UTR) of ank3 linked to a dual luciferase reporter into mCCD cells. Both aldosterone and miRs altered luciferase expression confirming the ank3‐UTR as a miR target. Knockdown of ank3 reduced ENaC short‐circuit currents (53.9 ± 5.13%, n=22) and overexpression of ank3 raised ENaC currents (140.6 ± 9.75%, n=12), independent of aldosterone or miR regulation, thereby linking expression of this protein with Na+ transport. Ank3 represents a novel aldosterone‐induced and miR‐regulated protein involved in altering Na+ transport in the CCD.