Cloning of a G-protein-coupled receptor that shows an activity to transform NIH3T3 cells and is expressed in gastric cancer cells
Cloning of a G-protein-coupled receptor that shows an activity to transform NIH3T3 cells and is expressed in gastric cancer cells
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DOI：
10.1111/j.1349-7006.2004.tb03193.x
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发表时间：
2004-02-01
期刊：
影响因子：
5.7
通讯作者：
Ikenaka, K
中科院分区：
文献类型：
--
作者：
Okumura, S;Baba, H;Ikenaka, K
本研究旨在确定导致胃癌形成的转化过程中涉及的新因素。利用逆转录病毒载体构建了人胃癌细胞的cDNA文库。通过筛选转导的NIH3T3细胞的转化活性进行功能克隆。共分离到6个克隆，其中1个编码已知在肿瘤发生中起作用的延伸因子1α亚基。其中一个在筛选过程中反复分离的cDNA(克隆56-2)编码一个与G蛋白偶联受体蛋白GPR35相同的蛋白质。此外，另一个克隆(72.3)是GPR35基因的选择性剪接产物，在GPR35的N-端增加了31个氨基酸。因此，克隆56.2和72.3编码的蛋白质分别命名为GPR35a和GPR35b。RT-PCR实验显示，GPR35基因在周围非癌区低表达或不表达，而在所有被检测的胃癌中都存在这两种mRNAs。72.3编码的信使核糖核酸水平始终显著高于56.2编码信使核糖核酸。在正常肠粘膜中检测到与胃癌相似的表达模式。根据这两个GPR35克隆在NIH3T3细胞中的明显转化活性，以及它们在癌组织中的表达水平显著上调，推测这两个新的GPR35亚型参与了胃癌的形成过程。
The present study was directed towards the identification of novel factors involved in the transformation process leading to the formation of gastric cancer. A cDNA library from human gastric cancer cells was constructed using a retroviral vector. Functional cloning was performed by screening for transformation activity in transduced NIH3T3 cells. Six cDNA clones were isolated, including one encoding the elongation factor 1alpha subunit, which was already known to play a role in tumorigenesis. One cDNA (clone 56-2), which was repeatedly isolated during the course of screening, encoded a protein identical to a G-protein-coupled receptor protein, GPR35. In addition, another cDNA clone (72.3) was found to be an alternatively spliced product of the GPR35 gene, whereby 31 amino acids were added to the N-terminus of GPR35. Hence, the proteins encoded by clones 56.2 and 72.3 were designated GPR35a and GPR35b, respectively. RT-PCR experiments revealed that GPR35 gene expression is low or absent in surrounding non-cancerous regions, while both mRNAs were present in all of the gastric cancers examined. The level of 72.3-encoded mRNA was consistently significantly higher than that of 56.2 encoded mRNA. An expression pattern similar to that observed in gastric cancers was detected in normal intestinal mucosa. Based on the apparent transformation activities of the two GPR35 clones in NIH3T3 cells, and the marked up-regulation of their expression levels in cancer tissues, it is speculated that these two novel isoforms of GPR35 are involved in the course of gastric cancer formation.