Analysis of the thyroid transcription factor-1(TTF-1) gene expression in thyroid neoplastic tissues.

甲状腺肿瘤组织中甲状腺转录因子1(TTF-1)基因表达分析

• 批准号: 07671121
• 负责人: ENDO Toyoshi
• 金额: $ 1.47万
• 依托单位: Yamanashi Medical University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1995
• 资助国家: 日本
• 起止时间: 1995 至 1996
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07671121/
• 关键词: thyroid; transcription; TTF-1; promoter; 遺伝子発現; 転写開始点; 組織特異性

Regulation of the thyroid transcription factor-1(TTF-1) gene expression in the thyroid was investigated. We identified a new transcription start site as nucleotide (ut)-1917, -1700 bp upstream of previously described site, and the region encom passing nt -1242 to -14 as the first intron. Northern blot analysis with FRTL-5 cell mRNA revealed that a probe targeted to exon 2 recognized 3.7 and 2.7 kb-transcripts, whereas both probes targeted to the untranslated exon 1 and the second intron deteced only the 3.7 kb-transcript, suggesting that the 3.7 kb-mRNA is transcribed from nt -1917 and that it contains the sacond intron. Chloramphenicol acetyltransferase (CAT) reporter gene assays demonstrated that the 5'-flanking region exhibited promoter activity in FRTL-5 cells but not in rat liver cells, indicating that this region confers the property of thyroid-specific exprression. Two consensus TTF-1 binding motifs were detected in this promoter region, and electrophoretic mobility-shift assays showed that oligonucleotide probes, each containing one of these motifs, formed a complex with the recombinant TTF-1 homeodomain. Moreover, recombinant TTF-1 increased the transcriptional activity in cells lacking TTF-1. These results suggest that transcriptional from the newly identified start site in the TTF-1 gene is positively regulated by TTF-1 in the thyroid.

研究了甲状腺中甲状腺转录因子-1（TTF-1）基因表达的调节。我们确定了一个新的转录起始位点为核苷酸（ut）-1917，位于先前描述的位点上游-1700 bp，并且包含nt -1242至-14的区域作为第一个内含子。对 FRTL-5 细胞 mRNA 的 Northern 印迹分析显示，靶向外显子 2 的探针识别 3.7 和 2.7 kb 转录本，而靶向非翻译外显子 1 和第二个内含子的两个探针仅检测到 3.7 kb 转录本，表明 3.7 kb-mRNA 是从 nt -1917 转录的，并且它包含第二个内含子。氯霉素乙酰转移酶 (CAT) 报告基因检测表明，5'-侧翼区域在 FRTL-5 细胞中表现出启动子活性，但在大鼠肝细胞中则没有，表明该区域具有甲状腺特异性表达的特性。在该启动子区域检测到两个共有的 TTF-1 结合基序，并且电泳迁移率变动测定表明，每个含有这些基序之一的寡核苷酸探针与重组 TTF-1 同源域形成复合物。此外，重组 TTF-1 增加了缺乏 TTF-1 的细胞的转录活性。这些结果表明，TTF-1 基因中新确定的起始位点的转录受到甲状腺中 TTF-1 的正向调节。