Nordeen S K, Ogden C A, Taraseviciene L, Lieberman B A
Department of Pathology, University of Colorado Health Sciences Center, Denver 80262, USA.
Mol Endocrinol. 1998 Jun;12(6):891-8. doi: 10.1210/mend.12.6.0118.
Hormone response elements (HREs) are considered enhancers, activating transcription in a relatively position- and orientation-independent fashion. Upon binding to an HRE, steroid receptors presumably contact coactivators and/or proteins associated with the transcription initiation complex. As a receptor target site is moved further from a fixed position such as the TATA box, not only will the spatial separation of the receptor with respect to its interaction partners change, so will the orientation due to the rotation of the DNA helix. Additional constraints may be imposed by the assembly of DNA into chromatin. Therefore, we have endeavored to test rigorously the assertion that HRE action is position independent. We have constructed a series of 42 chloramphenicol acetyltransferase expression vectors that contain a single progesterone/glucocorticoid receptor-binding site separated from a TATA box by 4 to 286 bp. The enhancer activity of the HRE was assessed after transient transfection of progesterone receptor-expressing fibroblasts. We find that the position of the HRE has a dramatic influence on induction by progestins. When closely juxtaposed to the TATA box, the HRE was unable to support a hormone response, perhaps due to direct steric hindrance with the transcription initiation complex. Full activity was gained by moving the HRE 10 bp further from the TATA sequence. As the HRE was moved incrementally further, activity remained near maximal over the next 26 bp. HRE activity then declined over the subsequent 26 bp and remained low for another 2.5 helical turns. Surprisingly, a narrow window of HRE activity occurred at an HRE-TATA box separation of 90-100 bp. Little or no hormone-induced transcriptional activity was observed when the HRE was positioned further from the TATA box. The addition of a second HRE or a basal (nuclear factor-1) element failed to relieve this constraint. A similar series of experiments was carried out in a mammary carcinoma cell line that expressed high levels of both glucocorticoid and progestin receptors. Data in these cells indicate that glucocorticoids and progestins supported a similar HRE position-activity profile, but this pattern of HRE activity was quite distinct from that seen in fibroblasts. This may be indicative of cell type-specific interactions between steroid receptors and adapter/coactivator proteins or cell type-specific activities such as acetylases or deacetylases participating in the steroid response.
激素反应元件（HREs）被认为是增强子，以相对位置和方向独立的方式激活转录。与HRE结合后，类固醇受体大概会与共激活因子和/或与转录起始复合物相关的蛋白质相互作用。当受体靶位点远离诸如TATA框这样的固定位置时，不仅受体与其相互作用伙伴之间的空间距离会改变，由于DNA螺旋的旋转，方向也会改变。DNA组装成染色质可能会带来额外的限制。因此，我们努力严格测试HRE作用与位置无关这一论断。我们构建了一系列42个氯霉素乙酰转移酶表达载体，这些载体包含一个单一的孕酮/糖皮质激素受体结合位点，与TATA框相隔4至286个碱基对。在瞬时转染表达孕酮受体的成纤维细胞后，评估HRE的增强子活性。我们发现HRE的位置对孕激素诱导有显著影响。当与TATA框紧密相邻时，HRE无法支持激素反应，这可能是由于与转录起始复合物存在直接的空间位阻。将HRE从TATA序列再移开10个碱基对可获得完全活性。随着HRE逐渐移得更远，在接下来的26个碱基对中活性保持在接近最大值的水平。然后HRE活性在随后的26个碱基对中下降，并在另外2.5个螺旋圈中保持较低水平。令人惊讶地是，在HRE与TATA框相隔90 - 100个碱基对时出现了一个狭窄的HRE活性窗口。当HRE离TATA框更远时，几乎没有观察到激素诱导的转录活性。添加第二个HRE或一个基础（核因子-1）元件未能缓解这种限制。在一个同时高表达糖皮质激素和孕激素受体的乳腺癌细胞系中进行了一系列类似的实验。这些细胞中的数据表明，糖皮质激素和孕激素支持类似但与成纤维细胞中所见不同的HRE位置 - 活性图谱。这可能表明类固醇受体与衔接蛋白/共激活因子之间存在细胞类型特异性相互作用，或者存在参与类固醇反应的细胞类型特异性活性，如乙酰化酶或去乙酰化酶。
