Progesterone signals through membrane progesterone receptors (mPRs) in MDA-MB-468 and mPR-transfected MDA-MB-231 breast cancer cells which lack full-length and N-terminally truncated isoforms of the nuclear progesterone receptor.
Progesterone signals through membrane progesterone receptors (mPRs) in MDA-MB-468 and mPR-transfected MDA-MB-231 breast cancer cells which lack full-length and N-terminally truncated isoforms of the nuclear progesterone receptor.
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DOI：
10.1016/j.steroids.2011.01.008
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发表时间：
2011-08
期刊：
影响因子：
2.7
通讯作者：
Thomas P
中科院分区：
文献类型：
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作者：
Pang Y;Thomas P
利用在MDA-MB-231乳腺癌细胞中过表达的重组MPR蛋白，研究了膜孕酮受体(MPR)的功能特性。虽然这些细胞不表达全长孕激素受体(PR)，但目前尚不清楚它们是否表达N端截短的PR亚型，这可能解释了MPR的某些孕激素受体功能。在本研究中，研究了未转染和转导MPR的MDA-MB-231细胞和MDA-MB-468乳腺癌细胞中N端截短PR异构体的存在。在PR阳性的T47D Yb乳腺癌细胞中，用两组PR末端的PR引物均可检测到扩增产物，但在未转染组和转染组的MDA-MB-231细胞和MDA-MB-468细胞中均未检测到扩增产物。使用C端PR抗体2C11F1的Western印迹分析显示，PR在这些细胞系中的分布模式相同。另一种C端PR抗体C-19在所有细胞系中都检测到免疫反应条带，但也识别α-Actinin，表明该抗体不是PR的特异性抗体。Mda-MB-468细胞质膜上有高亲和力的孕激素受体结合，经mprα和mprβ的siRNAs处理后显著降低。细胞质膜对PR激动剂R5020、≤的亲和力很低，仅为孕酮的1%。孕酮处理引起MDA-MB-468细胞G蛋白活化和cAMP生成减少，这也是MPR的特征。结果表明，孕激素受体在这些细胞系中的功能是通过MPR介导的，不涉及任何N末端截短的PR亚型。
The functional characteristics of membrane progesterone receptors (mPRs) have been investigated using recombinant mPR proteins over-expressed in MDA-MB-231 breast cancer cells. Although these cells do not express the full-length progesterone receptor (PR), it is not known whether they express N-terminally truncated PR isoforms which could possibly account for some progesterone receptor functions attributed to mPRs. In the present study, the presence of N-terminally truncated PR isoforms was investigated in untransfected and mPR-transfected MDA-MB-231 cells, and in MDA-MB-468 breast cancer cells. PCR products were detected in PR-positive T47D Yb breast cancer cells using two sets of C-terminus PR primers, but not in untransfected and mPR-transfected MDA-MB-231 cells, nor in MDA-MB-468 cells. Western blot analysis using a C-terminal PR antibody, 2C11F1, showed the same distribution pattern for PR in these cell lines. Another C-terminal PR antibody, C-19, detected immunoreactive bands in all the cell lines, but also recognized α-actinin, indicating that the antibody is not specific for PR. High affinity progesterone receptor binding was identified on plasma membranes of MDA-MB-468 cells which was significantly decreased after treatment with siRNAs for mPRα and mPRβ. Plasma membranes of MDA-MB-468 cells showed very low binding affinity for the PR agonist, R5020, ≤1% that of progesterone, which is characteristic of mPRs. Progesterone treatment caused G protein activation and decreased production of cAMP in MDA-MB-468 cells, which is also characteristic of mPRs. The results indicate that the progestin receptor functions in these cell lines are mediated through mPRs and do not involve any N-terminally truncated PR isoforms.