Yamada M, Hashimoto K, Satoh T, Shibusawa N, Kohga H, Ozawa Y, Yamada S, Mori M
First Department of Internal Medicine, Gunma University School of Medicine, Japan.
J Clin Endocrinol Metab. 1997 Dec;82(12):4224-8. doi: 10.1210/jcem.82.12.4438.
We measured the amounts of TRH receptor (TRHR) messenger ribonucleic acid (mRNA) in human normal pituitary and pituitary tumors and found a novel transcript of the TRHR gene. Competitive PCR revealed expression of the TRHR mRNA in all pituitary adenomas examined, and its level was variable and similar to that in the normal pituitary. When the C-terminal region was amplified by PCR, an additional short product was observed. Cloning and sequence analysis of this short fragment revealed that the deleted sequence corresponded exactly to the 5'-sequence of exon 3, indicating alternative splicing of the TRHR mRNA. This alternative splicing resulted in a frame shift, yielding a C-terminal truncated protein (HTRHR2) on translation. Expression analysis of HTRHR2 in Chinese hamster ovary cells showed no significant binding to [3HIMeTRH or response of intracellular calcium to TRH administration. However, the mRNA ratio of HTRHR2 vs. the wild type (HTRHR1) was significantly different among pituitary tumors. The highest ratio was observed in prolactinomas (30%), and almost no detectable expression was found in GH-producing tumors. These findings indicate that this novel transcript of the human TRH receptor gene is produced in a tumor-specific manner and may be a useful parameter for evaluation of individual pituitary tumors.
我们检测了人正常垂体及垂体肿瘤中促甲状腺激素释放激素受体（TRHR）信使核糖核酸（mRNA）的含量，发现了TRHR基因的一种新转录本。竞争性聚合酶链反应（PCR）显示，在所检测的所有垂体腺瘤中均有TRHR mRNA表达，其水平存在差异且与正常垂体中的水平相似。当通过PCR扩增C末端区域时，观察到一个额外的短产物。对该短片段进行克隆和序列分析表明，缺失序列与外显子3的5'序列完全对应，这表明TRHR mRNA存在可变剪接。这种可变剪接导致了移码，在翻译时产生了一种C末端截短的蛋白（HTRHR2）。在中国仓鼠卵巢细胞中对HTRHR2进行表达分析，结果显示其与[3H]MeTRH无明显结合，且给予促甲状腺激素释放激素（TRH）后细胞内钙也无反应。然而，在垂体肿瘤中，HTRHR2与野生型（HTRHR1）的mRNA比例存在显著差异。在催乳素瘤中该比例最高（30%），而在生长激素分泌型肿瘤中几乎检测不到其表达。这些发现表明，人促甲状腺激素释放激素受体基因的这种新转录本是以肿瘤特异性方式产生的，可能是评估个体垂体肿瘤的一个有用参数。
