Aldosterone-induced Na+ Absorption is Regulated by Protein Acetylation in a Cellular Model of the Cortical Collecting Duct: Experimental Biology 2017

Fine-tuning of total body sodium content occurs within the aldosterone-sensitive distal nephron (ASDN). Here, the volume-regulating hormone aldosterone can promote Na+ retention to offset changes in extracellular fluid volume by binding the mineralocorticoid receptor (MR). The resulting transcription of key intracellular proteins, including serum and glucocorticoid-induced kinase 1 (SGK1), promote Na+ reabsorption via the epithelial sodium channel (ENaC). Acetylation, a post-translational modification involving attachment of acetyl groups to lysine residues, appears to control the transcriptional activity of the MR. Indeed recently it was demonstrated in rats that experimentally-induced hypertension could be ameliorated by inhibiting histone deacetylases (HDACs), which remove acetyl groups. The molecular mechanisms underpinning this response, however, remain unclear. The aim of this study was to explore the role of acetylation in mediating aldosterone-induced Na+ absorption using a highly differentiated cellular model of the cortical collecting duct, mCCDcl1 cells.ENaC-mediated Na+ absorption was quantified across monolayers of cells via electrometric measurements. Expression of HDAC transcripts was determined by RT-PCR. The acetylation status of intracellular proteins as well as the abundance and activity of SGK1 were monitored by Western blot analysis.mCCDcl1 cells expressed all known nuclear class I and cytoplasmic class II HDACs, including HDAC3 (class I) previously suggested to modulate MR functionality. Trichostatin A (TSA), a broad-spectrum inhibitor of histone deacetylases, increased acetylation of histones 3 and 4 (class I HDAC activity), and α-tubulin (class II HDAC6-mediated) in a time- and concentration-dependent manner. A class I HDAC3-selective inhibitor, MI192, also increased acetylation of intracellular proteins although with a delayed effect relative to TSA. Aldosterone (3nM) stimulated ENaC-mediated Na+ transport by ~3-fold compared to control groups, associated with increased SGK1 abundance and activity. Pre-incubation with TSA abolished aldosterone-induced Na+ absorption and the associated increase in SGK1 abundance and activity. However, MI192 did not alter the effects of aldosterone in mCCDcl1 cells, implying a lack of involvement for HDAC3. Additionally, despite TSA inhibiting aldosterone-induced Na+ absorption, it did not modify insulin-stimulated ENaC currents demonstrating that insulin, which promotes ENaC activity via post-transcriptional regulation of SGK1, acts independently of acetylation.Together these data suggest that acetylation of intracellular proteins is critical for mediating aldosterone-induced ENaC-mediated Na+ absorption, however class I HDAC3 does not appear to play a role. Further work is required to elucidate the identity of the histone deacetylases involved in permitting this key hormonal response in the ASDN.