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Analysis of cellular functions of cells transfected with sense or antisense β-1,4-galactosyltransferase cDNA

正义或反义β-1,4-半乳糖基转移酶cDNA转染细胞的细胞功能分析

基本信息

批准号：
10680696
负责人：
FURUKAWA Kiyoshi
金额：
$ 2.11万
依托单位：
Tokyo Metropolitan Institute of Gerontology
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 1999
项目状态：
已结题

项目摘要

Mammalian cells are now shown to contain several β-1、4-galactosyltransferases (β-1,4-GalTs) which are involved in the biosynthesis of glycoprotein. When cells are transformed, they express the highly branched sugar chains with tumor-associated carbohydrate antigens on proteins which are critical to tumorigenicity and metastasis of the cell. During transformation, the gene expression levels of β-1,4-GalTs I, III, IV and VI were not changed while that of β-1,4-GalT II decreased and that of β-1,4-GalT V increased. In order to investigate biological significance of the carbohydrates formed by β-1,4-GalT II and V, their genes were introduced into several cell lines. When β-1,4-GalT II-sense cDNA or β-1,4-GalT V-antisense cDNA was transfected into mouse melanoma F10 cells, altered glycosylation of the membrane glycoproteins was detected by lectin blot analysis. The cells were inoculated into C57B mouse and the ability to form tumors were examined. Mouse melanoma F10 cells formed tumors in the animals while those transfected with β-1,4-Galt II-sense cDNA or β-1,4-GalT V-antisense cDNA failed or developed tumors very slowly. These results indicated that the sugar chains synthesized by β-1,4-GalTs II and V are involved in the growth control mechanism of cells. Further studies are necessary to identify which carbohydrate structures are really engaged in this biological event.

现在显示哺乳动物细胞含有几种参与糖蛋白生物合成的β-1、4-半乳糖基转移酶(β-1,4-GalTs)。当细胞转化时，它们在蛋白质上表达具有肿瘤相关碳水化合物抗原的高度支化糖链，这对细胞的致瘤性和转移至关重要。转化过程中，β-1,4-GalTs I、III、IV、VI基因表达量没有变化，β-1,4-GalT II基因表达量下降，β-1,4-GalT V基因表达量增加。为了研究β-1,4-GalT II和V形成的碳水化合物的生物学意义，将它们的基因引入到几种细胞系中。当 β-1,4-GalT II 有义 cDNA 或 β-1,4-GalT V 反义 cDNA 转染小鼠黑色素瘤 F10 细胞时，通过凝集素印迹分析检测到膜糖蛋白糖基化的改变。将细胞接种到C57B小鼠体内并检查形成肿瘤的能力。小鼠黑色素瘤F10细胞在动物体内形成肿瘤，而用β-1,4-Galt II-sense cDNA或β-1,4-GalT V-反义cDNA转染的细胞则失败或形成肿瘤非常缓慢。这些结果表明β-1,4-GalTs II和V合成的糖链参与细胞的生长控制机制。需要进一步的研究来确定哪些碳水化合物结构真正参与了这一生物事件。