Question:

The mineralocorticoid receptor (MR), a ligand-dependent transcription factor, classically contributes to electrolyte homeostasis and blood pressure control but independently mediates pathophysiological processes in the reno-cardiovascular system like inflammation, endothelial dysfunctions and vascular remodeling. Mechanistically, the MR resides in the cytosol in its unligand form. Upon binding of its natural ligand aldosterone, the MR translocates into the nucleus and binds at hormone response elements and regulates target gene expression. Interestingly, aldosterone stimulation mediates post-translational modifications (PTMs) of the MR which are implicated to alter MR functions. A common PTM which detailed analyses are lacking is MR phosphorylation. Based on sequence analyses, we identified casein kinase II (CK2) as prominent candidate.

Methods:

To identify CK2-induced MR phosphorylation sites we conducted peptide micro arrays. Afterwards, we investigated the influence of CK2 on MR functions like nuclear translocation, transactivation activity conducting reporter gene assays and mutagenesis experiments in HEK-293 cells.

Results and Conclusion:

To analyze the exact position of CK2-induced MR phosphorylation, peptide microarrays were conducted which showed that CK2 phosphorylates the MR at position serine 111 and 459 in the N-terminal domain. To verify the importance of this two CK2 phosphorylated serine residues mutagenesis experiment will be conducted. In the presence of the specific CK2 inhibitor TBCA aldosterone-induced MR transactivation activity was time- and concentration dependently reduced without affecting cell viability. These results show that the genomic MR activity is dependent on CK2-mediated MR phosphorylation. Mechanistically, the nuclear MR translocation after TBCA and aldosterone treatment was unchanged compared to aldosterone treatment alone suggesting the involvement of other mechanisms like altered MR-DNA interactions.