Relative contribution of molecular mechanisms to cumulative ligand-mediated downregulation of GRα

Central to the pharmacological use of glucocorticoids (GCs) is the availability of the glucocorticoid receptor alpha (GR alpha). However, chronic GC therapy often results in the ligand-mediated downregulation of the GR alpha, and the subsequent development of an acquired GC resistance. While studies have demon-strated the dimerization-dependent downregulation of GR alpha, as well as the molecular mechanisms through which ligand-mediated downregulation occurs, little is known regarding the relative contri-bution of these molecular mechanisms to the cumulative ligand-mediated downregulation of the receptor, especially within an endogenous system. Thus, to probe this, the current study evaluates the conformational-dependent regulation of GR alpha protein using mouse embryonic fibroblast (MEF) cells containing either wild type GR alpha (MEFwt) or the dimerization deficient GR alpha mutant (MEFdim) and in-hibitors of transcription, translation, and proteasomal degradation. Results show that the promotion of GR alpha dimerization increases the downregulation of the receptor via two main mechanisms, proteasomal degradation of the receptor protein, and downregulation of GRwt mRNA transcripts. In contrast, when receptor dimerization is restricted these two mechanisms play a lesser role and results suggest that stabilization of GR alpha protein by non-coding RNAs may potentially be the major regulatory mechanism. Together, these findings clarify the relative contribution of the molecular mechanisms involved in ligand-mediated downregulation of GR alpha and provides pharmacological targets for the development of GRa ligands with a more favourable therapeutic index. (C) 2022 Elsevier Inc. All rights reserved.