Cloning of a human thyroid hormone responsive gene

人甲状腺激素反应基因的克隆

• 批准号: 07671125
• 负责人: MURATA Yoshiharu
• 金额: $ 1.6万
• 依托单位: Nagoya 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-07671125/
• 关键词: thyroid hormone; fibroblast; human; cloning; gene

3,3', 5-Triodothyronine (T3), an active form of the thyroid hormone, plays an important role in fetal development and throughout life in humans. T3 exerts its effect through binding to its nuclear receptor and regulating expression of target genes. Thus, identification of T3-responsive genes in various tissue is important to elucidate T3 action at a molecular level in human. Utilizing a method called "differential display of mRNAs by means of polymerase chain reaction", the cDNA fragment of a T3-responsive gene named ZAKI-4 was cloned from cultured human skin fibroblasts. Northern blot analysis revealed that there were tow ZAKI-4mRNA species (3.4 and 1.4kb) and both mRNAs were increased by a physiological concentration of T3 within 12hr. This increase was due to an increased transcription of ZAKI-4 gene rather than increased stability of ZAKI-4 mRNA by T3. The T3 effect on ZAKI-4 expression was abolished by the treatment with cycloheximide, indicating the possibility that the regulatio … More n of ZAKI-4 mRNA by T3 is indirect, through an intermediate product induced by T3. ZAKI-4 mRNA was detected in heart, brain, liver, and skeletal muscle but not in placenta, lung, kidney and pancreas. In skin fibroblasts and skeletal muscle, 3.4-kb mRNA was the major species, whereas 1.4-kb mRNA was dominant in heart, brain, and liver. By using Rapid Amplification of cDNA Ends (RACE), the full-length cDNA was cloned. The sequence data confirmed the presence of two mRNA species which are possibly generated by an altermative polyadenylation and showed that these mRNAs code a single protein of 192 amino acids. Transcription coupled in vitro translation of the two mRNA species revealed both the two mRNA species code a protein with a molecular mass of 26 kDa, that roughly corresponds to the data of sequence analysis od ZAKI-4 cDNA.In fibroblasts from a patient with thyroid hormone resistance due to deletion of beta isoform of T3 receptor (TR) gene, T3 was able to induce ZAKI-4 mRNA like in normal fibroblasts. It is thus suggested that the other isoform, TR alpha can mediate the up-regulation of ZAKI-4 mRNA expression. Less

3,3'，5-三刺蛋白（T3）是一种甲状腺马酮的活性形式，在胎儿发育和整个生命中都起着重要作用。 T3通过与其核接收器结合并调节靶基因的表达来发挥其作用。这是，在各种组织中鉴定T3响应基因对于阐明人类分子水平的T3作用很重要。利用一种称为“通过聚合酶链反应的mRNA差异显示的方法”，从培养的人类皮肤成纤维细胞中克隆了一个T3反应基因的cDNA片段。 Northern印迹分析表明，有Tow Zaki-4 mRNA物种（3.4和1.4KB），两个mRNA在12小时以内的T3的物理浓度增加。这种增加是由于Zaki-4基因的转录增加而不是T3的Zaki-4 mRNA稳定性提高。用环己酰亚胺处理消除了T3对Zaki-4表达的影响，表明通过T3诱导的中间产物，T3的调节……更多的Zaki-4 mRNA是间接的。在心脏，大脑，肝脏和骨骼肌中检测到Zaki-4 mRNA，但在Plopeta，肺，肾脏和胰腺中未检测到。在皮肤成纤维细胞和骨骼肌中，3.4-kb mRNA是主要物种，而1.4-kb mRNA在心脏，大脑和肝脏中占主导地位。通过使用cDNA末端的快速扩增（种族），将全长cDNA克隆。序列数据证实存在两个mRNA物种，这些mRNA可能是由替代聚腺苷酸化产生的，并表明这些mRNA代码是192个氨基酸的单个蛋白质。 Transcription coupled in vitro translation of the two mRNA species revealed both the two mRNA species code a protein With a molecular mass of 26 kDa, that roughly corresponds to the data of sequence analysis od ZAKI-4 cDNA.In fibroblasts from a patient with thyroid homone resistance due to deletion of beta isoform of T3 receptor (TR) gene, T3 was able to induce ZAKI-4 mRNA like in normal成纤维细胞。因此，建议其他同工型TRα可以介导Zaki-4 mRNA表达的上调。较少的

项目成果

RE.Weiss, Y.Hayashi, T.Nagaya, KJ.Petty, Y.Murata, H.Tunka H.Seo, S.Refetoff: "Dominant inheritance of resistance to thyroid hormone not linked to defects in the thyroid hormone receptors alpha or beta genes may be due to a defective co-factor." J.Clin.En

RE.Weiss、Y.Hayashi、T.Nagaya、KJ.Petty、Y.Murata、H.Tunka H.Seo、S.Refetoff：“甲状腺激素抵抗的显性遗传与甲状腺激素受体 α 或 α 或 α 的缺陷无关。

S.Yamaguchi: "Hepatocytes cultured in monolayers have a lower responsiveness to thyroid hormone and retinoic acid than do hepatocytes cultured in spheroids : Possible role of retinoid X receptor." Environmental Medicine. 40(1). 47-50 (1996)

S.Yamaguchi：“单层培养的肝细胞对甲状腺激素和视黄酸的反应性比球状体培养的肝细胞低：类视黄醇 X 受体的可能作用。”

T. Miyazaki: "Molecular cloning of a novel thyroid hormone-responsive gene, ZAKI-4, in human skin fibroblasts." Journal of Biological Chemistry. 271. 14567-14571 (1996)

T. Miyazaki：“在人类皮肤成纤维细胞中分子克隆一种新型甲状腺激素反应基因 ZAKI-4。”

RE. Weiss: "Dominant inheritance of resistance to thyroid hormone not linked to defects in the thyroid hormone receptors a or β genes maby be due to a defective co-factor." Journal of Clinical Endocrinology and Metabolism. 81. 1496-4203 (1996)

RE. Weiss：“甲状腺激素抵抗的显性遗传与甲状腺激素受体 a 或 β 基因的缺陷无关，可能是由于辅助因子缺陷所致。” 81. 1496-4203 (1996) ）

S. Yamaguchi: "Hepatocytes cultured in monolayers have a lower responsiveness to thyroid hormone and retinoic acid than do hepatocytes cultured in spheroids : Possible role of retinoid X receptor." Environmental Medicine. 40. 47-50 (1996)

S. Yamaguchi：“单层培养的肝细胞对甲状腺激素和视黄酸的反应性比球状体培养的肝细胞低：类视黄醇 X 受体的可能作用。”