Progesterone Receptor Membrane Component-1 is phosphorylated upon treatment with Progestins and binds to Estrogen Receptor α-coregulators PHB1 and PHB2 in breast cancer cells
Progesterone Receptor Membrane Component-1 is phosphorylated upon treatment with Progestins and binds to Estrogen Receptor α-coregulators PHB1 and PHB2 in breast cancer cells
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黄体酮受体膜成分 1 在孕激素治疗后被磷酸化，并与乳腺癌细胞中的雌激素受体 α 辅助调节因子 PHB1 和 PHB2 结合
DOI：
10.1055/s-0037-1602574
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发表时间：
2017
期刊：
影响因子：
--
通讯作者：
Neubauer H
中科院分区：
文献类型：
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作者：
Willibald M;Bayer G;Stahlhut V;Poschmann G;Stühler K;Seeger H;Mueck AO;Niederacher D;Fehm T;Neubauer H
使用雌激素和合成孕激素的更年期激素联合治疗与乳腺癌的风险增加有关。孕激素对乳腺细胞的影响是复杂的，目前还不完全清楚。在以往的体外和体内研究中，我们发现不同的孕激素可以促进孕激素受体膜组分-1(PGRMC1)高表达的MCF7乳腺癌细胞的增殖，揭示了PGRMC1在将膜启动的孕激素信号传递到细胞内的潜在作用。因此，本研究旨在进一步探讨孕激素作用下PGRMC1的激活机制及其下游信号转导机制。为了确定孕激素结合后受体的翻译后修饰和潜在的相互作用伙伴，用MCF-7/PGRMC1细胞进行了免疫共沉淀实验，然后进行了质谱分析。为了进一步验证结果，进行了邻近连接试验和共定位研究。在此基础上，进一步研究了孕激素对具有PGRMC1磷酸化位点突变的MCF-7/PGRMC1细胞和MCF7/PGRMC1细胞的增殖作用。在这里，我们表明，孕激素去甲孕酮(NET)处理的MCF7/PGRMC1细胞诱导酪蛋白激酶2(CK2)磷酸化位点Ser181的受体磷酸化。MCF7/PGRMC1细胞中Ser181磷酸化位点的点突变在净处理时会损害细胞的增殖。此外，雌激素受体α(ER-α)-协调制子Prohibitin 1(PHB1)和Prohibitin 2(PHB2)被确定为孕激素治疗后PgRMC1的相互作用伙伴。这项研究进一步揭示了孕激素结合时受体的激活机制。这进一步表明，在以孕激素为基础的激素替代治疗中，PGRMC1在乳腺癌的进展中起着重要作用。
Combined 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 fully understood yet. 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 breast cancer cells, revealing a potential role of PGRMC1 in forwarding membrane-initiated progestin signals into the cell. Therefore, the aim of this study was to further investigate the activation mechanism and downstream signaling of PGRMC1 upon progestin treatment. To identify posttranslational modifications and potential interaction partners of the receptor after progestin binding, co-immunoprecipitation experiments were performed using MCF-7/PGRMC1 cells, followed by mass spectrometry analysis. To further validate the results, proximity ligation assay and co-localization studies were conducted. Further, proliferation of MCF-7/PGRMC1 cells and MCF7/PGRMC1 cells, possessing point mutations at PGRMC1 phosphorylation sites, was investigated upon treatment with progestins. Here we show that treatment of MCF7/PGRMC1 cells with the progestin norethisterone (NET) induces phosphorylation of the receptor at Casein Kinase 2 (CK2) phosphorylation site Ser181. Point mutation of the Ser181 phosphorylation site in MCF7/PGRMC1 cells impairs proliferation upon NET treatment. Further, the Estrogen Receptor α (ERα)-coregulators Prohibitin 1 (PHB1) and Prohibitin 2 (PHB2) are identified as interaction partners of PGRMC1 after progestin treatment. This study gives further insight into the activation mechanism of the receptor upon progestin binding. It further suggests an important role of PGRMC1 in the progression of breast cancer in progestin-based hormone replacement therapy.