Mechanistic studies have defined several molecular events associated with estrogen-dependent activation of the ER, including changes in receptor conformation, increased interaction with coactivators, increased receptor phosphorylation, and alterations in the ability of receptor to bend DNA. Although the mechanism(s) by which steroid ligands increase receptor phosphorylation are poorly defined at the present time, ligand-dependent activation of all steroid receptors studied to date, including the ER, is accompanied by an increase in receptor phosphorylation. The correlation between receptor phosphorylation and transcriptional activity suggests that these two events are functionally linked. buy ortho tri-cyclen
In the first transient transfection studies demonstrating ligand-independent activation of a steroid receptor, 8-bro-mo-cAMP (an activator of protein kinase A) or okadaic acid (an inhibitor of protein phosphatases 1 and 2A) was used to activate chicken PR-dependent expression of a synthetic target gene. Similarly, human ER can be ligand-independently activated by okadaic acid in CV, (green monkey kidney) cells, and rat ER is transcriptionally activated by cholera toxin and 3-isobutyl-1-methylxanthine (ШМХ, a phosphodiesterase inhibitor), or 8-bromo-cAMP in primary rat uterine cell cultures. In other cell types (MCF-7 and Chinese hamster ovary cells), neither cholera toxin/IBMX nor 12-O-tetradecanoylphorbol 13-acetate (TPA; an activator of protein kinase C) were able to increase ER-dependent reporter gene expression in the absence of estradiol. The variability in the ability of phosphorylating agents to modulate ER activity in a ligand-independent manner is likely to reflect promoter or cell-type specific differences in ER function. These differences also have been noted to influence basal and estrogen-stimulated ER-dependent transcription. Indeed, the ability of TPA to synergize with estradiol in the activation of ER-dependent transcription varies with these two parameters.