Cloning and characterization of the 5'-flanking region of the Ehox gene.
Cloning and characterization of the 5'-flanking region of the Ehox gene.
复制标题
Ehox 基因 5 侧翼区域的克隆和表征。
DOI：
10.1016/j.bbrc.2005.12.176
复制
发表时间：
2006
影响因子：
3.1
通讯作者：
Westphal,Heiner
中科院分区：
文献类型：
--
作者：
Lee,WoonKyu;Kim,Yong-Man;Malik,Nasir;Ma,Chang;Westphal,Heiner
含有配对同源盒的基因Ehox在胚胎干细胞分化中起作用，并在发育中的胎盘和胸腺中高度表达。为了了解Ehox基因表达的调控机制，我们从小鼠BAC文库中分离出Ehox基因的5 ' -侧翼区域。5 ' -RACE分析显示在翻译起始密码子上游130个核苷酸处有一个转录起始位点。用荧光素酶报告基因在序列缺失的5 '侧序列控制下瞬时转染，发现nt - 84至- 68区域含有一个阳性的顺式作用元件，可有效表达Ehox基因。对该区域的突变分析和电泳迁移率转移试验中的寡核苷酸竞争显示存在CCAAT盒子，这是转录核因子Y （NFY）的靶标。NFY是正向基因调控所必需的。在Ehox基因的1.9 kb 5 '侧区未发现组织特异性增强子。Ehox在胚胎发育的早期阶段，特别是在9.5dpc的大脑中表达，以及在胚胎发育的后期表达。这些结果表明，NFY是Ehox转录活性的重要调控因子，在胚胎着床后发育阶段起重要作用。
The paired-like homeobox-containing gene Ehox plays a role in embryonic stem cell differentiation and is highly expressed in the developing placenta and thymus. To understand the mechanisms of regulation of Ehox gene expression, the 5′-flanking region of the Ehox gene was isolated from a mouse BAC library. 5′-RACE analysis revealed a single transcriptional start site 130 nucleotides upstream of the translation initiation codon. Transient transfection with a luciferase reporter gene under the control of serially deleted 5′-flanking sequences revealed that the nt −84 to −68 region contained a positive cis-acting element for efficient expression of the Ehox gene. Mutational analysis of this region and oligonucleotide competition in the electrophoretic mobility shift assay revealed the presence of a CCAAT box, which is a target for transcription nuclear factor Y (NFY). NFY is essential for positive gene regulation. No tissue-specific enhancer was identified in the 1.9-kb 5′-flanking region of the Ehox gene. Ehox is expressed during the early stages of embryo development, specifically in the brain at 9.5dpc, as well as during the late stages of embryo development. These results suggest that NFY is an essential regulatory factor for Ehox transcriptional activity, which is important for the post-implantation stage of the developing embryo.