Molecular cloning of a novel thyroid hormone-responsive gene, ZAKI-4, in human skin fibroblasts
Molecular cloning of a novel thyroid hormone-responsive gene, ZAKI-4, in human skin fibroblasts
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DOI：
10.1074/jbc.271.24.14567
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发表时间：
1996-06-14
影响因子：
4.8
通讯作者：
Seo, H
中科院分区：
文献类型：
--
作者：
Miyazaki, T;Kanou, Y;Seo, H
利用称为“通过聚合酶链反应的mRNA差异显示”的方法，从培养的人皮肤成纤维细胞中克隆了甲状腺激素应答基因ZAKI-4的cDNA片段，北方印迹分析显示，有两种ZAKI-4 mRNA种类（3.4和1.4个酶（kb）），并且它们被生理浓度的三碘甲腺原氨酸（T-3）上调。T-3的这种作用被放线菌酮所消除，表明基因ZAKI-4可能是由T-3以间接的方式调节的，通过T-3的中间产物，而不是由T-3本身直接调节。T-3对ZAKI-4 mRNA稳定性没有影响，表明T-3在转录水平上诱导mRNA。cDNA末端的快速扩增证实了两种mRNA的存在。在心脏、脑、肝脏和骨骼肌中检测到ZAKI-4 mRNA，但在胎盘、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是主要种类，而1.4-kb mRNA在心脏、脑和肝脏中占主导地位。序列分析表明，这两种mRNA来源于交替的多聚腺苷酸化，编码一个192个氨基酸的蛋白质。在数据库中没有发现同源蛋白质序列。阐明ZAKI-4基因产物的功能将为T-3在各种器官中的重要作用提供新的见解。
Utilizing a method called ''differential display of mRNAs by means of polymerase chain reaction'', the cDNA fragment of a thyroid hormone-responsive gene ZAKI-4 was cloned from cultured human skin fibroblasts, Northern blot analysis revealed that there were two ZAKI-4 mRNA species (3.4 and 1.4 kilobases (kb)), and they were up-regulated by a physiological concentration of triiodothyronine (T-3). This T-3 effect was abolished by the treatment with cycloheximide, indicating the possibility that gene ZAKI-4 is regulated by T-3 in an indirect fashion, through an intermediate product of T-3, rather directly by T-3 itself, No effect of T-3 on ZAKI-4 mRNA stability suggested that T-3 induces the mRNA at the transcriptional level. Rapid amplification of cDNA ends confirmed the presence of two mRNA species. 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. The sequence analysis suggested that the two mRNA species arise from alternative polyadenylation and code a single protein of 192 amino acids. No homologous protein sequence was found in a data base. Elucidation of the function of ZAKI-4 gene product will provide new insights into an important role of T-3 in various organs.