Department of Reproductive Biology, Instituto Nacional de Ciencias Médicas y Nutrición Salvador Zubirán, Vasco de Quiroga No. 15, Mexico City 14000, Mexico.
Gene. 2010 Jun 15;458(1-2):37-44. doi: 10.1016/j.gene.2010.03.007. Epub 2010 Mar 24.
Estrogen receptors (ER), members of the nuclear steroid receptor superfamily, act to activate transcription through ligand-dependent recruitment of coregulators and chromatin modifications. A series of synthetic A-ring reduced 19-nortestosterone-derived progestins has the capacity to selectively bind ERalpha for activated transcription, and to recruit coregulatory factors. In this study, we have analyzed the ability of synthetic 19-nortestosterone derivatives to visibly alter the configuration of ER-target gene chromatin using a novel mammalian promoter transcriptional biosensor (PRL-array) stably transfected into the genome of HeLa cells (PRL-HeLa cells). Results from synthetic steroid-treated cells expressing functional GFP-ERalpha or YFP-ERbeta chimeras were compared to those obtained with estradiol (E(2)) and the antiestrogen tamoxifen. In the presence of synthetic ligands or E(2) a concentration-dependent increase in area of the biosensor array was observed in GFP-ERalpha-expressing PRL-HeLa cells. No significant differences were found between the effects obtained with natural and synthetic steroids. Similarly, E(2) or synthetic steroids-treated PRL-HeLa cells also resulted in similar colocalization of SRC-1- and RNAPII-immunofluorescence at the array. YFP-ERbeta-expressing PRL-HeLa cells treated with E(2) showed increases in array area that were similar to ERalpha; however, treatment of YFP-ERbeta-expressing cells with synthetic ligands was indistinguishable from vehicle controls. These data indicate that A-ring reduced 19-nortestosterone derivatives have an estrogen-like effect on chromatin, including recruitment of transcription factors through selective interactions with ERalpha.
雌激素受体（ER）是核甾体受体超家族的成员，通过配体依赖性募集共激活因子和染色质修饰来激活转录。一系列合成的 A 环还原 19-去甲睾酮衍生孕激素具有选择性结合 ERalpha 以激活转录和募集共激活因子的能力。在这项研究中，我们使用一种新型哺乳动物启动子转录生物传感器（PRL-array）分析了合成 19-去甲睾酮衍生物改变 ER 靶基因染色质构象的能力，该生物传感器稳定转染到 HeLa 细胞的基因组中（PRL-HeLa 细胞）。用功能性 GFP-ERalpha 或 YFP-ERbeta 嵌合体表达的合成类固醇处理的细胞的结果与用雌二醇（E(2)）和抗雌激素他莫昔芬获得的结果进行了比较。在合成配体或 E(2)存在的情况下，GFP-ERalpha 表达的 PRL-HeLa 细胞中的生物传感器阵列面积呈浓度依赖性增加。天然和合成类固醇获得的效果之间没有发现显著差异。同样，用 E(2)或合成类固醇处理的 PRL-HeLa 细胞也导致在阵列处 SRC-1-和 RNAPII-免疫荧光的相似共定位。用 E(2)处理的表达 YFP-ERbeta 的 PRL-HeLa 细胞显示出与 ERalpha 相似的阵列面积增加；然而，用合成配体处理 YFP-ERbeta 表达细胞与载体对照没有区别。这些数据表明，A 环还原的 19-去甲睾酮衍生物对染色质具有雌激素样作用，包括通过与 ERalpha 的选择性相互作用募集转录因子。
