Zeiger M A, Norton J A
Metabolism Section, National Cancer Institute, National Institutes of Health, Bethesda, Md. 20892.
Surgery. 1993 Aug;114(2):458-62; discussion 462-3.
The isolation of mRNA molecules that are either uniquely or more highly expressed by tumor and not normal tissue is a powerful tool in the study of cell regulation and growth. To this end we constructed a complementary DNA (cDNA) library from messenger RNA (mRNA) isolated from a human insulinoma and, by differential hybridization with cDNA from both normal pancreas and insulinoma, isolated clones more highly expressed by insulinoma.
Total RNA was isolated from human insulinoma and normal pancreas and purified to mRNA by oligo (dT) column. An insulinoma cDNA library was constructed and screened with 32P-labeled cDNA from pancreas and insulinoma. Northern blots from insulinoma, pancreas, carcinomas, normal endocrine tissues, and endocrine tumors were then probed with the 32P-labeled inserts.
Two clones that consistently hybridized with 32P cDNA from insulinoma and not pancreas proved to represent mRNAs for insulin and the alpha subunit of the Gs protein. There was a markedly higher expression (30-fold) of the gene for Gs alpha in mRNA from insulinoma compared with normal pancreas by Northern blot analysis. We found Gs alpha to be more highly expressed by a pheochromocytoma, a corticotropin-producing islet cell tumor of the pancreas, and a corticotropin-producing thymic carcinoid (up to 35-fold) compared with normal pancreas, whereas normal endocrine tissues, a parathyroid adenoma, thyroid follicular adenoma, gastrinoma, and several carcinomas showed no expression.
This study showed that Gs alpha is highly expressed in insulinoma and certain endocrine tumors. It is not expressed in several cancers or normal endocrine tissues. Others have implicated mutated Gs proteins in the tumorigenesis of pituitary and thyroid tumors. G proteins are also known to mediate hormonal transmembrane signaling. Its overexpression in four of seven endocrine tumors tested suggests that it may have a role in the unregulated hormone secretion and/or a role in the tumorigenesis of differentiated endocrine tumors.
分离肿瘤组织而非正常组织中独特表达或高表达的mRNA分子是研究细胞调控与生长的有力工具。为此，我们从人胰岛素瘤中分离出的信使RNA（mRNA）构建了一个互补DNA（cDNA）文库，并通过与正常胰腺和胰岛素瘤的cDNA进行差异杂交，分离出胰岛素瘤中高表达的克隆。
从人胰岛素瘤和正常胰腺中分离总RNA，并通过寡聚（dT）柱纯化至mRNA。构建胰岛素瘤cDNA文库，并用来自胰腺和胰岛素瘤的32P标记的cDNA进行筛选。然后用32P标记的插入片段对胰岛素瘤、胰腺、癌、正常内分泌组织和内分泌肿瘤的Northern印迹进行杂交。
两个始终与胰岛素瘤而非胰腺的32P cDNA杂交的克隆被证明代表胰岛素和Gs蛋白α亚基的mRNA。通过Northern印迹分析，胰岛素瘤mRNA中Gsα基因的表达明显高于正常胰腺（30倍）。我们发现，与正常胰腺相比，嗜铬细胞瘤、胰腺促肾上腺皮质激素分泌胰岛细胞瘤和促肾上腺皮质激素分泌胸腺类癌中Gsα的表达更高（高达35倍），而正常内分泌组织、甲状旁腺腺瘤、甲状腺滤泡性腺瘤、胃泌素瘤和几种癌均无表达。
本研究表明Gsα在胰岛素瘤和某些内分泌肿瘤中高表达。在几种癌症或正常内分泌组织中不表达。其他人认为突变的Gs蛋白与垂体和甲状腺肿瘤的发生有关。已知G蛋白介导激素跨膜信号传导。在七个测试的内分泌肿瘤中有四个中其过表达表明它可能在激素分泌失控和/或分化型内分泌肿瘤的发生中起作用。
