Huang Jing, Li Xiaodong, Maguire Casey A, Hilf Russell, Bambara Robert A, Muyan Mesut
University of Rochester School of Medicine and Dentistry, Department of Biochemistry and Biophysics, Rochester, New York 14642, USA.
Mol Endocrinol. 2005 Nov;19(11):2696-712. doi: 10.1210/me.2005-0120. Epub 2005 Jun 23.
The functions of 17beta-estradiol (E2) are mediated by estrogen receptor (ER) alpha and beta. ERs display similar DNA- and ligand-binding properties in vitro. However, ERbeta shows lower transcriptional activity than ERalpha from the estrogen response element (ERE)-dependent signaling. We predicted that distinct amino termini contribute to differences in transcription efficacies of ERs by affecting in situ ER-ERE interactions. We used chromatin immunoprecipitation and a novel in situ ERE competition assay, which is based on the ability of ER to compete for ERE binding with a designer activator that constitutively induces transcription from an ERE-driven reporter construct. Interference of activator-mediated transcription by unliganded or liganded ERs was taken as an indication of ER-ERE interaction. Results revealed that ERs interacted with ERE similarly in the absence of E2. However, E2 enhanced the ERE binding of ERalpha but not that of ERbeta. The removal of the amino terminus increased the ERbeta-ERE interaction independent of E2. The ERbeta amino terminus also prevented E2-mediated enhancement of the chimeric ERalpha-ERE interaction. Thus, the amino terminus of ERbeta impairs the binding of ERbeta to ERE. The abrogation of ligand-dependent activation function 2 of the amino-terminally truncated ERbeta resulted in the manifestation of E2 effect on ERbeta-ERE interaction. This implies that E2-mediated enhancement of ERbeta-ERE interaction is masked by the activation function 2, whereas the intact amino terminus is a dominant region that decreases the binding of ERbeta to ERE. Thus, ERbeta-ERE interaction is independent of E2 and is impaired by its amino terminus. These findings provide an additional explanation for differences between ERalpha and ERbeta functions that could differentially affect the physiology and pathophysiology of E2 signaling.
17β-雌二醇（E2）的功能由雌激素受体（ER）α和β介导。体外实验中，ERα和ERβ显示出相似的DNA结合和配体结合特性。然而，从雌激素反应元件（ERE）依赖性信号传导来看，ERβ的转录活性低于ERα。我们推测，不同的氨基末端通过影响原位ER-ERE相互作用，导致ERs转录效率存在差异。我们使用了染色质免疫沉淀和一种新型的原位ERE竞争试验，该试验基于ER与设计激活剂竞争ERE结合的能力，该激活剂可组成性地诱导ERE驱动的报告基因构建体转录。未结合或结合配体的ER对激活剂介导转录的干扰被视为ER-ERE相互作用的指标。结果显示，在没有E2的情况下，ERs与ERE的相互作用相似。然而，E2增强了ERα与ERE的结合，但未增强ERβ与ERE的结合。去除氨基末端可增加ERβ与ERE的相互作用，且与E2无关。ERβ的氨基末端还可阻止E2介导的嵌合ERα-ERE相互作用增强。因此，ERβ的氨基末端会损害ERβ与ERE的结合。氨基末端截短的ERβ的配体依赖性激活功能2的缺失，导致E2对ERβ-ERE相互作用的影响得以显现。这意味着E2介导的ERβ-ERE相互作用增强被激活功能2所掩盖，而完整的氨基末端是降低ERβ与ERE结合的主要区域。因此，ERβ-ERE相互作用独立于E2，且被其氨基末端所削弱。这些发现为ERα和ERβ功能差异提供了额外的解释，这些差异可能会对E2信号传导的生理和病理生理产生不同影响。
