Identification of transforming activity of free fatty acid receptor 2 by retroviral expression screening
Identification of transforming activity of free fatty acid receptor 2 by retroviral expression screening
复制标题
DOI：
10.1111/j.1349-7006.2009.01348.x
复制
发表时间：
2010-01-01
期刊：
影响因子：
5.7
通讯作者：
Mano, Hiroyuki
中科院分区：
文献类型：
--
作者：
Hatanaka, Hisashi;Tsukui, Mamiko;Mano, Hiroyuki
胆囊癌（GBC）是一种高度致命的恶性肿瘤。KRAS或TP 53的遗传改变以及ERBB 2的过表达已被证明有助于某些类型GBC的发展。然而，许多GBC病例并没有这种遗传变化，其他转化事件有待发现。我们在这里试图确定新的促癌基因在GBC中，使用逆转录病毒cDNA表达文库。从手术切除的GBC临床标本中构建逆转录病毒cDNA表达文库，并在病灶形成试验中用于感染3 T3成纤维细胞。通过PCR拯救掺入转化灶中的cDNA。其中一种cDNA编码游离脂肪酸受体2（FFAR 2），一种短链脂肪酸的G蛋白偶联受体。FFAR 2的致癌潜力在体外用病灶形成试验和通过评价软琼脂中的细胞生长以及在裸鼠体内用致瘤性试验证实。分离的FFAR 2 cDNA没有序列改变，表明FFAR 2表达的上调可能有助于恶性转化。事实上，所有的定量RT-PCR，原位杂交和免疫组化分析表明，FFAR 2 mRNA及其蛋白产物的量在消化道癌标本中增加。此外，短链脂肪酸增强FFAR 2在3 T3细胞中的促有丝分裂作用。因此，我们的数据首次表明FFAR 2与消化道癌的发生有关。（Cancer Sci 2009）。
Gallbladder cancer (GBC) is a highly fatal malignancy in humans. Genetic alterations in KRAS or TP53 as well as overexpression of ERBB2 have been shown to contribute to the development of certain types of GBC. However, many cases of GBC do not harbor such genetic changes, with other transforming events awaiting discovery. We here tried to identify novel cancer-promoting genes in GBC, with the use of a retroviral cDNA expression library. A retroviral cDNA expression library was constructed from a surgically resected clinical specimen of GBC, and was used to infect 3T3 fibroblasts in a focus formation assay. cDNA incorporated into the transformed foci was rescued by PCR. One such cDNA was found to encode free fatty acid receptor 2 (FFAR2), a G protein-coupled receptor for short-chain fatty acids. The oncogenic potential of FFAR2 was confirmed both in vitro with the focus formation assay and by evaluation of cell growth in soft agar as well as in vivo with a tumorigenicity assay in nude mice. The isolated FFAR2 cDNA had no sequence alterations, suggesting that upregulation of FFAR2 expression may contribute to malignant transformation. Indeed, all of quantitative RT-PCR, in situ hybridization, and immunohistochemical analyses showed that the amount of FFAR2 mRNA and its protein product was increased in digestive tract cancer specimens. Furthermore, short-chain fatty acids potentiated the mitogenic action of FFAR2 in 3T3 cells. Our data thus, for the first time, implicate FFAR2 in carcinogenesis of the digestive tract. (Cancer Sci 2009).