Glucocorticoid receptor Thr524 phosphorylation by MINK1 induces interactions with 14-3-3 protein regulators

Glucocorticoid receptor (GR) is a ligand-dependent transcription factor that plays a central role in inflammation. GR activity is also modulated via protein-protein interactions, including binding of 14-3-3 proteins induced by GR phosphorylation. However, the specific phosphorylation sites on GR that trigger these interactions and their functional consequences are less clear. Hence, we sought to examine this system in more detail. We used phosphorylated GR peptides, biophysical studies and X-ray crystallography to identify key residues within the ligand binding domain of GR, T524 and S617, whose phosphorylation results in binding of the representative 14-3-3 protein 14-3-3ζ. A kinase screen identified MINK1 as responsible for phosphorylating T524 and ROCK1 for phosphorylating S617; cell-based approaches confirmed the importance of both GR phosphosites and MINK1 but not ROCK1 alone in inducing GR-14-3-3 binding. Together our results provide molecular-level insight into 14-3-3-mediated regulation of GR and highlight both MINK1 and the GR-14-3-3 axis as potential targets for future therapeutic intervention.