Studies on the gene expression involved in the placetal development.

胎盘发育相关基因表达的研究。

• 批准号: 08671324
• 负责人: YAMADA Kazuyo
• 金额: $ 1.34万
• 依托单位: Fujita Health University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08671324/
• 关键词: transcription factor; aromatase; trophoblast; placenta; 栄養胚芽細胞; hCG; プロモーター; シスエレメント

We found a new cis-element neccessary for the placenta-specific enhancer in the aromatase gene. Co-presence of the element and the trophoblast specific element (TSE) in the aromatase enhancer region is sufficient to direct trophoblast specific expression driven by a heterologous tk promoter. The binding activity to this site was found in the nuclear extracts prepared from JEG-3 cells and the human placenta but not in the cell extract from HeLa or Hep G2 cells. This binding activity is distinct in electrophoreic mobility from the protein that binds to the TSE-like elements in aromatase enhancer. The core DNA sequence for the binding of the new trans-factor is also distinct from that of the TSE binding protein. The similar sequence is however found in the 24-base pair Trophoblast Specific Element (TSE) derived from the a-hCG promoter in reverse direction thus this DNA fragment also binds to the new factor with low affinity. This new trans-factor is tentatively called TSEBP2.A2.65 Kilobase cDNA clone that encodes a 436 amino acid was isolated from a human placental cDNA library using a yeast one-hybrid system with the TSEBP2 recognition sequence (GATCCATAAGACCCTCATTCCAGAG). The transfection of this clone with baculovirus system in sf21 cells resulted the production of a nuclear protein that showed the same binding specificity with the TSEBP2 in the nuclear extract prepared from Jeg 3. Northern-blot analysis showed that the mRNA coresponding to this clone was found exclusively in the placenta. This protein is likely to be a trophoblast specific transcription factor that confer placenta-specific expression of aromatase gene. The primary structure of the tentative transactivation domain however, has no similarity with other known transcription factors. The proof of transcriptional activity is on the way.

我们在芳香化酶基因中发现了一个新的胎盘特异性增强子所必需的顺式元件。该元件和滋养细胞特异性元件（TSE）在芳香化酶增强子区域的共同存在足以指导由异源tk启动子驱动的滋养细胞特异性表达。在JEG-3细胞和人胎盘的核提取物中发现了与该位点的结合活性，但在HeLa或Hep G2细胞的细胞提取物中没有发现。这种结合活性在电泳迁移率上不同于芳香化酶增强剂中与tse样元素结合的蛋白质。结合新反式因子的核心DNA序列也不同于TSE结合蛋白的核心DNA序列。然而，在从a-hCG启动子衍生的24碱基对滋养细胞特异性元件（Trophoblast Specific Element， TSE）中发现了类似的序列，但方向相反，因此该DNA片段也以低亲和力与新因子结合。该反式因子暂称为TSEBP2. a2.65千碱基cDNA克隆，编码436个氨基酸，利用酵母单杂交系统与TSEBP2识别序列（GATCCATAAGACCCTCATTCCAGAG）从人胎盘cDNA文库中分离得到。用杆状病毒系统转染sf21细胞后，在Jeg 3的核提取物中产生了与TSEBP2具有相同结合特异性的核蛋白。Northern-blot分析表明，该克隆对应的mRNA只存在于胎盘中。该蛋白可能是滋养细胞特异性转录因子，赋予芳香酶基因胎盘特异性表达。然而，暂定转激活结构域的主要结构与其他已知的转录因子没有相似性。转录活性的证据已经在路上了。