The progesterone receptor hinge region regulates the kinetics of transcriptional responses through acetylation, phosphorylation, and nuclear retention

Progesterone receptors (PRs) are critical regulators of mammary gland development and contributors to breast cancer progression. Posttranslational modifications of PR have been shown to alter hormone responsiveness. Site-directed mutagenesis demonstrated that upon hormone binding, PR is acetylated at the consensus sequence, KXKK (amino acids 638-641), located within the hinge region. We created an acetylation-deficient (K-A) mutant as well as acetylation mimics (K-Q or K-T). Interestingly, similar to K-A PR, PR acetylation mimics (K-Q or K-T) displayed delayed phosphorylation and nuclear entry relative to wild-type (wt) PR-B, indicative of disruption of PR nuclear-cytoplasmic shuttling. Wt PR-B, but not K-mutant PRs, induced c-myc at 1 h of progestin treatment. However, at 6 h of treatment, c-myc induction was comparable with levels induced by wt PR-B, suggesting that the precise timing of PR phosphorylation and nuclear retention are critical for cells to rapidly initiate robust transcriptional programs. In contrast to c-myc, progestin-induced serum- and glucocorticoid-regulated kinase (SGK) expression displayed sensitivity to PR acetylation but not nuclear entry. Namely, in the presence of progestin, acetylation-deficient (K-A) mutant PR-B up-regulated SGK mRNA relative to wt PR; progesterone response element-luciferase assays confirmed this result. However, K-Q and K-T acetylation mimics only weakly induced SGK expression independently of nuclear retention. These data reveal the ability of PR acetylation to alter the magnitude of transcriptional response at selected (slow response) promoters (SGK), whereas the hinge region dictates the kinetics of the transcriptional response to hormone at other (rapid response) promoters (c-myc). In sum, the PR hinge region is multifunctional. Understanding the ability of this region to couple acetylation, phosphorylation, and nuclear entry may provide clues to mechanisms of altered hormone responsiveness.