mTOR regulates mineralocorticoid receptor transcriptional activity by ULK1- dependent and independent mechanisms.

The mineralocorticoid receptor (MR) is a transcription factor for genes mediating diverse, cell-specific functions, including trophic effects as well as promoting fluid/electrolyte homeostasis. It was reported that in intercalated cells, phosphorylation of the MR at serine 843 (S843) by ULK1 inhibits MR activation and that phosphorylation of ULK1 by mTOR inactivates ULK1, and thereby prevents MR inactivation. We extended these findings with studies in M1 mouse cortical collecting duct cells stably expressing the rat MR and a reporter gene. Pharmacological inhibition of ULK1 dose-dependently increased ligand-induced MR transactivation, while ULK1 activation had no effect. Pharmacological inhibition of mTOR and CRISPR/gRNA gene knockdown of Raptor or Rictor decreased phosphorylated ULK1 and ligand-induced activation of the MR reporter gene, as well as transcription of endogenous MR-target genes. As predicted, ULK1 inhibition had no effect on aldosterone-mediated transcription in M1 cells with the mutated MR-S843A (alanine cannot be phosphorylated). In contrast, mTOR inhibition dose-dependently decreased transcription in the MR-S843A cells, though not as completely as in cells with the wt MR-S843. mTOR, Raptor, and Rictor co-precipitated with the MR and addition of aldosterone increased their phosphorylated, active, state. These results suggest that mTOR significantly regulates MR activity in at least two ways: by suppressing MR inactivation by ULK1, and by a yet ill-defined mechanism which involves direct association with MR. They also provide new insights into the diverse functions of ULK1 and mTOR, two key enzymes that monitor the cell's energy status.