p(O)estrogen receptor-α (ERα), a hormone-dependent transcription factor belonging to the steroid/thyroid-hormone-receptor superfamily, plays an essential role in the development and maintenance of the skeleton. Here we report the analysis of an unexplored sequence inside the bone-specific distal promoter F (PF) with respect to the regulation of ERα gene expression in bone. This sequence, 785 bp in size, is localized upstream of the assigned transcription start site of exon F, at ?117140 bp from the originally described transcription start site +1. It contains a TA reach box, a conventional CAAT box and potential regulatory elements for many transcription factors, including Cbfa1 [OSE2 (osteoblast-specific element) core binding factor], GATA-1 [(A/T)GATA(A/G) binding protein], Sox5 [sex-determining region Y (SRY)-type HMG bOX protein, belonging to a subfamily of DNA-binding proteins with an HMG domain], Sry, AP1 (activator protein 1) and CP2 (activator of γ-globin). It is able to strongly activate the luciferase reporter gene in MG-63 osteoblastic-like cells, but not in MCF7 breast-cancer cells. This is in agreement with different transcripts that we found in the two cell types. The footprinting and electrophoretic mobility-shift assays (EMSAs) showed that, inside the region analysed, there were some sequences that specifically reacted to nuclear proteins isolated from MG-63 cells. In particular, we identified two regions, named PFia/i and PFib/i, that do not present binding sites for known transcription factors and that are involved in a strong DNA–protein interaction in MG-63, but not in MCF7, cells. The analysis of three transcription factors (GATA-1, Sry and Sox) that might bind the identified footprinted areas suggested a possible indirect role of these proteins in the regulation of ERα gene expression in bone. These data provide evidence for different promoter usage of the ERα gene through the recruitment of tissue-specific transcription activators and co-regulators./p
展开▼
机译：（o）雌激素受体-α（ERα）是一种激素依赖性转录因子，属于类固醇/甲状腺激素受体超家族，在骨骼的发育和维持中起着至关重要的作用。在这里，我们报告了骨特异性远端启动子F（PF）内部未开发序列的分析，该序列涉及骨中ERα基因表达的调节。该序列的大小为785 bp，位于外显子F的指定转录起始位点的上游，距最初描述的转录起始位点+1约117140 bp。它包含一个TA到达盒，一个常规的CAAT盒和许多转录因子的潜在调控元件，包括Cbfa1 [OSE2（成骨细胞特异性元件）核心结合因子]，GATA-1 [（A / T）GATA（A / G）结合蛋白]，Sox5 [性决定区Y（SRY）型HMG bOX蛋白，属于具有HMG结构域的DNA结合蛋白的亚家族]，Sry，AP1（激活蛋白1）和CP2（γ-激活蛋白）珠蛋白）。它能够在MG-63成骨细胞样细胞中强烈激活萤光素酶报告基因，但在MCF7乳腺癌细胞中却不能。这与我们在两种细胞类型中发现的不同转录本是一致的。足迹和电泳迁移率迁移分析（EMSA）显示，在所分析的区域内，有一些序列与从MG-63细胞分离的核蛋白发生了特异性反应。特别是，我们确定了两个区域，分别称为PF a 和PF b ，这两个区域不存在已知转录因子的结合位点，并且参与了强烈的DNA-蛋白质相互作用在MG-63中，但不在MCF7中。对可能结合已确定的足迹区域的三种转录因子（GATA-1，Sry和Sox）的分析表明，这些蛋白可能在调节骨骼中ERα基因表达中起间接作用。这些数据通过募集组织特异性转录激活因子和共调节因子，为ERα基因的不同启动子用法提供了证据。
展开▼
