Effects of hormone and cellular modulators of protein phosphorylation on transcriptional activity, DNA binding, and phosphorylation of human progesterone receptors.

Although protein phosphatase magnesium-dependent 1 ␦ (PPM1D) was initially characterized as a p53-regulated phosphatase responsible for inactivation of p38 MAPK and consequent inactivation of p53, its overexpression and amplification in human breast cancers led us to assess its role in steroid hormone action. We found that PPM1D stimulated the activity of several nuclear receptors including the progesterone receptor (PR) and estrogen receptor. Although p38 MAPK inhibited PR activity, PPM1D stimulation of PR activity was greater than that achieved by a chemical inhibitor of p38 MAPK, SB202190. This suggests an additional novel function for PPM1D. Consistent with this, the transcriptional activity of endogenous PR in MCF-7 breast cancer cells was preferentially inhibited by small interfering RNA for PPM1D; SB202190 failed to reverse the inhibition. Although PPM1D phosphatase activity was required for stimulation of transcriptional activity, the activity of a PR phosphorylation site null mutant was enhanced by PPM1D, indicating that PR is not the direct target. Additional studies revealed that PPM1D enhanced the intrinsic activity of p160 coactivators such as steroid receptor coactivator-1 and promoted the interaction between PR and steroid receptor coactivator-1 in a mammalian two-hybrid assay. Neither activity was induced by SB202190. Although PPM1D stimulated PR activity in part through inhibition of p38 MAPK, its primary action is novel and independent of p38 MAPK. Thus, we speculate that PPM1D promotes breast tumor growth both by inhibiting p53 activity and by enhancing steroid hormone receptor action.

Section: Discussionmentioning

confidence: 99%

Dual Roles for the Phosphatase PPM1D in Regulating Progesterone Receptor Function

Proia

Nannenga

Donehower

et al. 2006

“…Chicken PR-A are activated in a ligand-independent manner by cAMP and epidermal growth factor (36,37). Despite attempts to do so, this phenomenon has not been reliably demonstrated for human PRs (38). In addition to chicken PRs, several other nuclear receptors can be activated by dopamine through D1 receptors (39), and dopamine placed directly into the third ventricle of the brain increases female rat sexual behavior in a progesterone-independent, but PR-dependent manner (40).…”

Section: Discussionmentioning

confidence: 99%

New Human Breast Cancer Cells to Study Progesterone Receptor Isoform Ratio Effects and Ligand-independent Gene Regulation

Jacobsen

Richer

Schittone

et al. 2002

(iii) inducible PR-B cells (Y iB), (iv) stable PR-B plus inducible PR-A cells (B iA), and (v) stable PR-A plus inducible PR-B cells (A iB). Expression levels of each isoform and/or the PR-A/PR-B ratios could be tightly controlled by the dose of induceras demonstrated by immunoblotting and transcription studies. Induced PRs underwent normal progestin-dependent phosphorylation and down-regulation and regulated exogenous promoters as well as endogenous gene expression. Transcription of exogenous promoters was dependent on the PR-A/PR-B ratio, whereas transcription of endogenous genes was more complex. Finally, we have described several genes that are regulated by induced PR-A even in the absence of ligand.

“…For example, the chicken PR (23), androgen receptor (24), retinoic acid receptor (25), retinoid X receptor (26), and peroxisome proliferator-activated receptor-␦ (27) can be activated by cAMP signaling, demonstrating that this mode of ligand-independent activation is not exclusive to ER␣. However, human PR (28) cannot be activated ligand-independently via this mechanism, nor can the unliganded glucocorticoid receptor, although cAMP stimulation increases the hormone-dependent responses of these receptors (29,30). Collectively, these reports demonstrate the specific nature by which the cAMP signaling pathway can cross-talk with different nuclear receptors.…”

mentioning

confidence: 86%

Mechanistic Differences in the Activation of Estrogen Receptor-α (ERα)- and ERβ-dependent Gene Expression by cAMP Signaling Pathway(s)

Coleman

Dutertre

El-Gharbawy

et al. 2003

Although increases in intracellular cAMP can stimulate estrogen receptor-␣ (ER␣) activity in the absence of exogenous hormone, no studies have addressed whether ER␤ can be similarly regulated. In transient transfections, forskolin plus 3-isobutyl-1-methylxanthine (IBMX), which increases intracellular cAMP, stimulated the transcriptional activities of both ER␣ and ER␤. This effect was blocked by the protein kinase A inhibitor H89 (N-(2-(p-bromocinnamylamino)-ethyl)-5-isoquinolinesulfonamide) and was dependent on an estrogen response element. A 12-O-tetradecanoylphorbol-13-acetate response element (TRE) located 5 to the estrogen response element was necessary for cAMP-dependent activation of gene expression by ER␤ but not ER␣, indicating that the former subtype requires a functional interaction with TRE-interacting factor(s) to stimulate transcription. Both p160 and CREB-binding protein coactivators stimulated cAMP-induced ER␣ and ER␤ transcriptional activity. However, mutation of the two cAMP-inducible SRC-1 phosphorylation sites important for cAMP activation of chicken progesterone receptor or all seven known SRC-1 phosphorylation sites did not specifically impair cAMP activation of ER␣. The E/F domains of ER␣ are sufficient for activation by forskolin/IBMX, and this is accompanied by an increase in receptor phosphorylation. In contrast, cAMP signaling reduces the phosphorylation of the corresponding region of ER␤, and this correlates with the lack of forskolin/IBMX stimulated transcriptional activity. Our data suggest that cAMP activation of ER␣ transcriptional activity is associated with receptor instead of SRC-1 phosphorylation. Moreover, differences in the cofactor requirements, domains of ER␣ and ER␤ sufficient for forskolin/IBMX activation, and the effect of cAMP on receptor phosphorylation indicate that this signaling pathway utilizes distinct mechanisms to stimulate ER␣ and ER␤ transcriptional activity.