Regulation of steroid receptor action temporal and structural aspects of specificity

Specificity of steroid receptor action is investigated in the present work, by analysis of the transcriptional activity of the androgen receptor (AR) through the cell cycle in contrast with the activity of the glucocorticoid receptor (GR) and by the construction and characterization of chimeras between the GR and the mineralocorticoid receptor (MR). We demonstrate that in G0, the AR potently enhances transcription. This activity is strongly reduced or abolished at the G1/S boundary. In S-phase, the receptor regains activity, indicating that there is a transient regulatory event that inactivates the AR at the G1/S transition. This regulation is specific for the AR, since the related GR is transcriptionally active in G1/S. Not all of the effects of androgens are blocked, however, since androgens retain the ability to increase AR protein levels at the G1/S junction. Inhibition of histone deacetylases partly rescues AR activity, demonstrating the involvement of acetylation pathways in the cell cycle regulation of AR transcriptional activity. Finally, a model is proposed which explains the inactivity of the AR at the G1/S transition by integrating the action of cell-cycle regulators and histone acetyltransferase-containing coactivators.; The GR/MR chimeras presented in this study are novel proteins that, contrary to our expectations, show increased ligand specificity compared to the parental receptors. We have described a receptor that is specific for the potent synthetic glucocorticoid TA and does not bind aldosterone. Another analyzed protein acts as a general corticoid receptor, responding to glucocorticoids and mineralocorticoids with similar potency and efficacy. A further set of chimeras have an increased ability to discriminate between ligands, responding potently to mineralocorticoids and only very weakly to synthetic glucocorticoids. Finally, a hinge chimera highlights the role of this domain as it only responds to mineralocorticoids unlike both the GR and the MR. Our data suggests that regions of the GR and the MR HBD are functionally non-reciprocal and that the determinants of hormone binding and transcriptional specificity are not found in equivalent receptor subdomains. We have presented evidence that suggests that receptor-specific interactions within domains, across receptor domains and with coactivators mediate aspects of specificity.