Sp6 regulation of Rock1 promoter activity in dental epithelial cells.
Sp6 regulation of Rock1 promoter activity in dental epithelial cells.
复制标题
DOI：
10.2152/jmi.61.306
复制
发表时间：
2014
期刊：
影响因子：
--
通讯作者：
Ryna Dwi Yanuaryska;Keiko Miyoshi;Arya Adiningrat;Taigo Horiguchi;Ayako Tanimura;Hiroko Hagita;T. Noma
中科院分区：
文献类型：
--
作者：
Ryna Dwi Yanuaryska;Keiko Miyoshi;Arya Adiningrat;Taigo Horiguchi;Ayako Tanimura;Hiroko Hagita;T. Noma
Sp 6是SP/KLF家族的转录因子，是牙齿发育过程中成釉细胞分化的形态学动力学的不可或缺的调节因子。然而，潜在的分子机制仍不清楚。我们以前已经确定了一个SP 6的下游基因，Rock 1，这是参与成釉细胞极化。在这项研究中，我们研究了Rock 1的转录调控机制的Sp 6。首先，我们确定了转录起始位点（TSS），并克隆了Rock 1的5 '侧翼区。系列缺失分析确定了一个关键区域内的249 bp的TSS上游区域的Rock 1启动子活性，染色质免疫沉淀试验显示Sp 6结合到这个区域。随后的瞬时转染实验表明，Rock 1启动子活性增强的Sp 6，但减少的Sp1。用GC选择性DNA结合抑制剂，光辉霉素A，治疗牙上皮细胞，影响Rock 1启动子活性的增强Sp 6的损失，但不抑制Sp1。进一步的定点突变表明，从-206到-150的区域含有Sp 6的响应元件。综上所述，我们得出结论，Sp 6正调控Rock 1的转录直接结合到Rock 1启动子区域从-206到-150，这在功能上不同于Sp1。
Sp6 is a transcription factor of the SP/KLF family and an indispensable regulator of the morphological dynamics of ameloblast differentiation during tooth development. However, the underlying molecular mechanisms remain unclear. We have previously identified one of the Sp6 downstream genes, Rock1, which is involved in ameloblast polarization. In this study, we investigated the transcriptional regulatory mechanisms of Rock1 by Sp6. First, we identified the transcription start sites (TSS) and cloned the 5'-flanking region of Rock1. Serial deletion analyses identified a critical region for Rock1 promoter activity within the 249-bp upstream region of TSS, and chromatin immunoprecipitation assays revealed Sp6-binding to this region. Subsequent transient transfection experiments showed that Rock1 promoter activity is enhanced by Sp6, but reduced by Sp1. Treatment of dental epithelial cells with the GC-selective DNA binding inhibitor, mithramycin A, affected Rock1 promoter activity in loss of enhancement by Sp6, but not repression by Sp1. Further site-directed mutagenesis indicated that the region from -206 to -150 contains responsive elements for Sp6. Taken together, we conclude that Sp6 positively regulates Rock1 transcription by direct binding to the Rock1 promoter region from -206 to -150, which functionally distinct from Sp1.