Transcriptional synergism between the vitamin D3 receptor and other nonreceptor transcription factors.
Transcriptional synergism between the vitamin D3 receptor and other nonreceptor transcription factors.
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DOI：
10.1210/mend.8.12.7708050
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发表时间：
1994-12
影响因子：
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通讯作者：
Min Liu;Leonard P. Freedman
中科院分区：
文献类型：
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作者：
Min Liu;Leonard P. Freedman
反式作用因子浓度和/或组合的微小变化可能导致基因表达的深刻改变。与启动子/增强子区域内的不同位点结合的不同类别的转录因子之间的协同相互作用是发生这种情况的一种机制。反映这一点的是，激素反应元件（类固醇/核受体的 DNA 识别位点）通常存在于组织为多个拷贝的启动子区域中，或者聚集在其他反式作用因子的结合位点中。为了系统地检查此类受体维生素 D3 受体 (VDR) 与其他非受体转录因子之间的潜在相互作用，我们构建了一系列报告质粒，其中含有骨桥蛋白 (Spp1) 维生素 D 反应元件 (VDRE) 的一个副本，由两个间隔 3 个碱基对的同向重复序列和一个转录因子 SP1、NF-1、Oct-1 或 AP-1 的结合位点组成。我们还生成了受两个 Spp1 VDRE 拷贝控制的报告基因，或一个与人骨钙素基因启动子不同的 VDRE 控制下的报告基因。在存在和不存在 1,25-二羟基维生素 D3 的情况下，使用各种报告基因瞬时转染 HeLa 或 CV-1 细胞。我们的结果表明，两个 Spp1-VDRE 的 VDR 反式激活强度比一个 Spp1-VDRE 强 12-20 倍，表明 VDR 与其自身产生协同作用。 VDR 还与其他非受体因子具有协同作用，因为我们在添加配体后观察到 6 至 12 倍的协同诱导程度，具体取决于特定因素。转录协同作用的功能基础似乎是在协同 DNA 结合的水平上，至少对于单独的 VDR 和 VDR-Oct-1 而言，如使用纯化因子的凝胶迁移率变化测定在体外所证明的那样。与此一致的是，我们表明 VDR 体内转录协同作用的最低要求是其 DNA 结合域。
Small changes in the concentrations and/or combinations of trans-acting factors can result in profound alterations in gene expression. Synergistic interaction between different classes of transcription factors bound to distinct sites within a promoter/enhancer region is one mechanism by which this can occur. Reflecting this, hormone response elements, DNA recognition sites for steroid/nuclear receptors, are often found in promoter regions organized as multiple copies or are clustered among binding sites for other trans-acting factors. To systematically examine the potential interactions between one such receptor, the vitamin D3 receptor (VDR), and other nonreceptor transcription factors, we constructed a series of reporter plasmids containing one copy of the osteopontin (Spp1) vitamin D response element (VDRE), consisting of two direct repeats spaced by 3 base pairs, and one binding site for the transcription factors SP1, NF-1, Oct-1, or AP-1. We also generated reporters either under the control of two copies of Spp1 VDRE, or a distinct VDRE from the human osteocalcin gene promoter. The various reporters were used to transiently transfect HeLa or CV-1 cells in the presence and absence of 1,25-dihydroxyvitamin D3. Our results show that VDR transactivates 12-20 times more strongly from two Spp1-VDREs than from one, indicating that VDR synergizes with itself. VDR also synergizes with the other nonreceptor factors, since we observe a 6- to 12-fold degree of synergistic induction after ligand addition, depending on the particular factor. The functional basis for the transcriptional synergism appears to be at the level of cooperative DNA binding, at least for VDR alone and VDR-Oct-1, as demonstrated in vitro by gel mobility shift assays using purified factors. Consistent with this, we show that the minimal requirement for transcriptional synergism in vivo by VDR is its DNA-binding domain.