【作者】 贾爽；

【导师】 燕秋；

【作者基本信息】 大连医科大学 ， 生物化学与分子生物学， 2008， 硕士

【摘要】 α1,3-岩藻糖基转移酶(α1,3 FUT)参与Lewis糖链合成最后一步岩藻糖化反应,包括FUT3、FUT4、FUT5、FUT6、FUT7、FUT9六种。其中FUT4是催化LeY寡糖合成的关键酶,它可将GDP-岩藻糖转移到N-乙酰氨基葡萄糖(GlcNAc)的末端形成α1,3糖苷键。正常生理条件下,FUT4主要在白细胞和上皮细胞中表达;而在肠癌、克隆癌、胰腺癌中表达异常升高。因此,调控特异岩藻糖基转移酶表达可改变LeY寡糖的合成,进而影响LeY寡糖参与的生物学功能。LeY寡糖[Fucα1→2Galβ1→4(Fucα1→3)GlcNAcβ1→R]是细胞表面的糖蛋白和糖脂分子上的Ⅱ型岩藻化寡糖。研究显示:LeY寡糖参与个体发育、器官形成、造血细胞分化及肿瘤发生与转移等。约70-90%的人类上皮细胞来源的肿瘤高表达LeY,并与肿瘤细胞的增殖、浸润和转移潜能密切相关,因此被认为是一种肿瘤相关的标志物。本课题组前期工作表明:应用RNAi技术抑制FUT4表达及利用LeY寡糖抗原的特异抗体封闭LeY,可抑制人上皮癌细胞A431的生长;而FUT4的过表达可促进LeY的合成,并抑制细胞凋亡。关于FUT4对人上皮癌细胞增殖和凋亡的影响机制还不完全清楚。信号传导分子蛋白激酶C在肿瘤的发生发展中有重要作用,并且蛋白激酶Cα亚型(PKCα)作为靶向药物治疗肿瘤已经在临床上得到初步应用。本文旨在通过细胞凋亡相关检测及Western blot等方法,研究转染FUT4干扰序列和反义PKCα对A431细胞凋亡作用的影响及初步探讨其作用机制。实验结果表明:FUT4 RNAi及PKCαAS均可以可以诱导A431细胞的凋亡,而且二者有一定的协同作用;FUT4 RNAi可能通过调控PKCα的表达诱导细胞凋亡。更多还原

【Abstract】 α1,3-fucosyltransferases(α1,3 FUT)participate in the fucosylation synthesis of Lewis oligosaccharide chains by transferring GDP-fucose to the terminal N-acetyllactosamine(GlcNAc)with theα1,3 orα1,4 linkage.α1,3 FUTs include FUT3, 4, 5, 6, 7 and 9 genes. Fucosyltransferase IV (FUT4) is an essential enzyme that catalyzes the synthesis of Lewis Y (LeY) by transferring GDP-fucose to the terminal N-acetylglucosamine(GlcNAc)with theα1,3-linkage. FUT4 is mainly expressed in leukocyte and some epithelial cells. FUT4 was found abnormally expressed, eg, gastric carcinoma, colorectal cancerous, pancreatic cancer, et al. Therefore, regulating the expression of fucosyltransferases may change the synthesis of LeY and affect the biological function of LeY.LeY antigen carried by the glycoproteins and glycolipids on cell surface, is a difucosylated oligosaccharide with the chemical structure [Fucα1→2Galβ1→4(Fucα1→3)GlcNAcβ1→R] . LeY is able to facilitate henogenesis, organ formation, cell differentiation, tumorigenesis et al. LeY has been recognized as a tumor-associated antigen in a variety of cancer process particularly in cell proliferation, invasion and metastasis, highly expressed LeY is found in the majority (70-90%) of human cancers of epithelial orogin.The previous work in our lab showed that the suppression of FUT4 by RNAi and LeY antibody could inhibit the A431 cell proliferation, the overpression of FUT4 could increase the synthesis of LeY and inhibit cell apoptosis. The effect of FUT4 on cell proliferation and cell apoptosis in A431 cells is not clearly elucidated. It is found that protien kinase C has an important role in the development and progress of cancers. PKCαas a targeted anti-cancer drugs has made significant progress in clinic trial. This work mainly explored the effect of FUT4 RNAi and PKCαas on cell apoptosis in A431 cells.更多还原