Park Seong-Eun, Xu Jianming, Frolova Antonina, Liao Lan, O'Malley Bert W, Katzenellenbogen Benita S
University of Illinois, Department of Molecular and Integrative Physiology, 524 Burrill Hall, 407 South Goodwin Ave., Urbana, IL 61801-3704, USA.
Mol Cell Biol. 2005 Mar;25(5):1989-99. doi: 10.1128/MCB.25.5.1989-1999.2005.
We previously identified a coregulator, repressor of estrogen receptor activity (REA), that directly interacts with estrogen receptor (ER) and represses ER transcriptional activity. Decreasing the intracellular level of REA by using small interfering RNA knockdown or antisense RNA approaches in cells in culture resulted in a significant increase in the level of up-regulation of estrogen-stimulated genes. To elucidate the functional activities of REA in vivo, we have used targeted disruption to delete the REA gene in mice. The targeting vector eliminated, by homologous recombination, the REA exon sequences encoding amino acids 12 to 201, which are required for REA repressive activity and for interaction with ER. The viability of heterozygous animals was similar to that of the wild type, whereas homozygous animals did not develop, suggesting a crucial role for REA in early development. Female, but not male, heterozygous animals had an increased body weight relative to age-matched wild-type animals beginning after puberty. REA mRNA and protein levels in uteri of heterozygous animals were half that of the wild type, and studies with heterozygous animals revealed a greater uterine weight gain and epithelial hyperproliferation in response to estradiol (E2) and a substantially greater stimulation by E2 of a number of estrogen up-regulated genes in the uterus. Even more dramatic in REA heterozygous animals was the loss of down regulation by E2 of genes in the uterus that are normally repressed by estrogen in wild-type animals. Mouse embryo fibroblasts derived from heterozygous embryos also displayed a greater transcriptional response to E2. These studies demonstrate that REA is a significant modulator of estrogen responsiveness in vivo: it normally restrains estrogen actions, moderating ER stimulation and enhancing ER repression of E2-regulated genes.
我们之前鉴定出一种共调节因子，雌激素受体活性抑制因子（REA），它可直接与雌激素受体（ER）相互作用并抑制ER转录活性。在培养细胞中使用小分子干扰RNA敲低或反义RNA方法降低细胞内REA水平，会导致雌激素刺激基因的上调水平显著增加。为了阐明REA在体内的功能活性，我们利用靶向破坏技术在小鼠中删除了REA基因。靶向载体通过同源重组消除了编码氨基酸12至201的REA外显子序列，这些序列是REA抑制活性以及与ER相互作用所必需的。杂合动物的活力与野生型相似，而纯合动物无法发育，这表明REA在早期发育中起关键作用。从青春期开始，雌性而非雄性杂合动物相对于年龄匹配的野生型动物体重增加。杂合动物子宫中的REA mRNA和蛋白质水平是野生型的一半，对杂合动物的研究表明，对雌二醇（E2）的反应中子宫重量增加和上皮细胞过度增殖更为明显，并且E2对子宫中一些雌激素上调基因的刺激也显著增强。在REA杂合动物中更显著的是，子宫中那些在野生型动物中通常被雌激素抑制的基因失去了E2介导的下调作用。源自杂合胚胎的小鼠胚胎成纤维细胞对E2也表现出更大的转录反应。这些研究表明，REA是体内雌激素反应性的重要调节因子：它通常抑制雌激素作用，调节ER刺激并增强ER对E2调节基因的抑制作用。
