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Progesterone receptor membrane component 1 is phosphorylated upon progestin treatment in breast cancer cells.

ABSTRACT: Menopausal hormone therapy, using estrogen and synthetic progestins, is associated with an increased risk of developing breast cancer. The effect of progestins on breast cells is complex and not yet fully understood. In previous in vitro and in vivo studies, we found different progestins to increase the proliferation of Progesterone Receptor Membrane Component-1 (PGRMC1)-overexpressing MCF7 cells (MCF7/PGRMC1), suggesting a possible role of PGRMC1 in transducing membrane-initiated progestin signals. Understanding the activation mechanism of PGRMC1 by progestins will provide deeper insights into the mode of action of progestins on breast cells and the often-reported phenomenon of elevated breast cancer rates upon progestin-based hormone therapy. In the present study, we aimed to further investigate the effect of progestins on receptor activation in MCF7 and T47D breast cancer cell lines. We report that treatment of both breast cancer cell lines with the progestin norethisterone (NET) induces phosphorylation of PGRMC1 at the Casein Kinase 2 (CK2) phosphorylation site Ser181, which can be decreased by treatment with CK2 inhibitor quinalizarin. Point mutation of the Ser181 phosphorylation site in MCF7/PGRMC1 cells impaired proliferation upon NET treatment. This study gives further insights into the mechanism of differential phosphorylation of the receptor and confirms our earlier hypothesis that phosphorylation of the CK2-binding site is essential for activation of PGRMC1. It further suggests an important role of PGRMC1 in the tumorigenesis and progression of breast cancer in progestin-based hormone replacement therapy.

SUBMITTER: Willibald M

PROVIDER: S-EPMC5641147 | biostudies-other | 2017 Sep

REPOSITORIES: biostudies-other

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Progesterone receptor membrane component 1 is phosphorylated upon progestin treatment in breast cancer cells.

Willibald Marina M Bayer Giuliano G Stahlhut Vanessa V Poschmann Gereon G Stühler Kai K Gierke Berthold B Pawlak Michael M Seeger Harald H Mueck Alfred O AO Niederacher Dieter D Fehm Tanja T Neubauer Hans H

Oncotarget 20170802 42

Menopausal hormone therapy, using estrogen and synthetic progestins, is associated with an increased risk of developing breast cancer. The effect of progestins on breast cells is complex and not yet fully understood. In previous <i>in vitro</i> and <i>in vivo</i> studies, we found different progestins to increase the proliferation of Progesterone Receptor Membrane Component-1 (PGRMC1)-overexpressing MCF7 cells (MCF7/PGRMC1), suggesting a possible role of PGRMC1 in transducing membrane-initiated ...[more]

PMID: 29069804

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Project description:BackgroundPGRMC1 (progesterone receptor membrane component 1) is a highly conserved heme binding protein, which is overexpressed especially in hormone receptor-positive breast cancer and plays an important role in breast carcinogenesis. Nevertheless, little is known about the mechanisms by which PGRMC1 drives tumor progression. The aim of our study was to investigate the involvement of PGRMC1 in cholesterol metabolism to detect new mechanisms by which PGRMC1 can increase lipid metabolism and alter cancer-related signaling pathways leading to breast cancer progression.MethodsThe effect of PGRMC1 overexpression and silencing on cellular proliferation was examined in vitro and in a xenograft mouse model. Next, we investigated the interaction of PGRMC1 with enzymes involved in the cholesterol synthesis pathway such as CYP51, FDFT1, and SCD1. Further, the impact of PGRMC1 expression on lipid levels and expression of enzymes involved in lipid homeostasis was examined. Additionally, we assessed the role of PGRMC1 in key cancer-related signaling pathways including EGFR/HER2 and ERα signaling.ResultsOverexpression of PGRMC1 resulted in significantly enhanced proliferation. PGRMC1 interacted with key enzymes of the cholesterol synthesis pathway, alters the expression of proteins, and results in increased lipid levels. PGRMC1 also influenced lipid raft formation leading to altered expression of growth receptors in membranes of breast cancer cells. Analysis of activation of proteins revealed facilitated ERα and EGFR activation and downstream signaling dependent on PGRMC1 overexpression in hormone receptor-positive breast cancer cells. Depletion of cholesterol and fatty acids induced by statins reversed this growth benefit.ConclusionPGRMC1 may mediate proliferation and progression of breast cancer cells potentially by altering lipid metabolism and by activating key oncogenic signaling pathways, such as ERα expression and activation, as well as EGFR signaling. Our present study underlines the potential of PGRMC1 as a target for anti-cancer therapy.

Project description:IntroductionBasal phenotype breast cancers (BPBC) are often associated with apparent epithelial to mesenchymal transition (EMT). The role of progesterone (P4) in regulating EMT of BPBC has not been reported.MethodsThe EMT relevant biology was investigated in vitro using human BPBC cell models (MDA-MB468 and MDA-MB231) with P4, PR agonist (RU486), and PR antagonist (R5020) treatments. The essential role of membrane progesterone receptor alpha (mPRalpha) in the P4-regulated EMT was demonstrated by knocking down the endogenous gene and/or stably transfecting exogenous mPRalpha gene in the BPBC cell models.ResultsThe expression of snail and down-stream EMT proteins such as occludin, fibronectin, and E-cadherin was significantly regulated by P4 incubation, which was accompanied by cell morphological reversion from mesenchymal to epithelial phenotypes. In searching for the cell mediator of P4' action in the MDA-MB468 (MB468) cells, it was found that mPRalpha but not the nuclear PR has an essential role in the P4 mediated EMT inhibition. Knocking down the expression of mPRalpha with specific siRNA blocked the P4's effects on expression of the EMT proteins. In another BPBC cell line--MDA-MB231 (MB231), which is mPRalpha negative by Western blotting--P4 treatment did not alter cell proliferation and EMT protein expressions. Introduction of the exogenous mPRalpha cDNA into these cells caused cell proliferation, but not EMT, to become responsive to P4 treatment. In further studies, it was found that activation of the PI3K/Akt pathway is necessary for the P4-induced EMT reversion. To define the potential inter-mediate steps between mPRalpha and PI3K, we demonstrated that mPRalpha, caveolin-1 (Cav-1), and epidermal growth factor receptor (EGFR) are colocalized in the membrane of caveolar vesicle and the P4-repressed EMT in MB468 cells can be blocked by EGFR inhibitor (AG1478) and PI3K inhibitor (wortmannin).ConclusionsOur data suggest that the signaling cascade of P4 induced mesenchymal repression is mediated through mPRalpha and other caveolae bound signaling molecules namely Cav-1, EGFR, and PI3K. This novel finding may have great impact on fully understanding the pathogenesis of BPBC and provide an essential clue for developing a targeted therapeutic strategy for treatment of BPBC.

Dataset Information

Progesterone receptor membrane component 1 is phosphorylated upon progestin treatment in breast cancer cells.

Publications

Progesterone receptor membrane component 1 is phosphorylated upon progestin treatment in breast cancer cells.

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