Guo Hua-Bei, Lee Intaek, Kamar Maria, Akiyama Steven K, Pierce Michael
Department of Biochemistry and Molecular Biology and Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia 30602, USA.
Cancer Res. 2002 Dec 1;62(23):6837-45.
Altered expression of cell surface N-linked oligosaccharides is associatedwith the oncogenic transformation of many types of animal cells. One of the most common forms of glycosylation in transformed cells and human tumors is the highly elevated beta1,6 branching of N-linked oligosaccharides caused by increased transcription of N-acetylglucosaminyltransferase V (GnT-V). To characterize the effects of increased beta1,6 branching on cell-matrix adhesion-mediated phenotypes, human fibrosarcoma HT1080 cells were transfected with retroviral systems encoding GnT-V that used both noninducible and tetracycline-inducible promoters. Increased GnT-V expression resulted in a >25% inhibition of cell attachment to and a >50% inhibition of cell spreading on fibronectin. Both cell adhesion and spreading were suppressed by function-blocking antibodies specific for the alpha(5) and beta(1) integrin subunits of the fibronectin receptor. Cell migration toward fibronectin and invasion through Matrigel were both substantially stimulated in cells with induced expression of GnT-V. Induction of GnT-V had no effect on the level of cell surface expression of alpha(5) and beta(1) integrin subunits but did result in a more diffuse staining of the alpha(5) and beta(1) integrin subunits on the cell surface, suggesting that inhibition of integrin clustering may be causing these cells to be less adhesive and more motile. Surprisingly, there was no detectable expression of N-linked beta1,6 branching on the alpha(5) subunit purified from HT1080 cells before and after induction of GnT-V; by contrast, however, the beta(1) subunit showed a basal level of beta1,6 branching that was greatly increased after induction of GnT-V. These results suggest that changes in N-linked beta1,6 branching that occur during oncogenesis alter cell-matrix adhesion and migration by modulating integrin clustering and subsequent signal transduction pathways. These effects most likely result from altered N-linked carbohydrate expression on the beta(1) integrin subunit.
细胞表面N - 连接寡糖表达的改变与多种动物细胞的致癌转化有关。转化细胞和人类肿瘤中最常见的糖基化形式之一是N - 乙酰葡糖胺转移酶V（GnT - V）转录增加导致N - 连接寡糖的β1,6分支高度升高。为了表征β1,6分支增加对细胞 - 基质粘附介导的表型的影响，用编码GnT - V的逆转录病毒系统转染人纤维肉瘤HT1080细胞，该系统使用非诱导型和四环素诱导型启动子。GnT - V表达增加导致细胞对纤连蛋白的附着抑制>25%，在纤连蛋白上的细胞铺展抑制>50%。细胞粘附和铺展均被纤连蛋白受体的α(5)和β(1)整合素亚基的功能阻断抗体所抑制。在诱导表达GnT - V的细胞中，细胞向纤连蛋白的迁移和通过基质胶的侵袭均受到显著刺激。GnT - V的诱导对α(5)和β(1)整合素亚基的细胞表面表达水平没有影响，但确实导致细胞表面α(5)和β(1)整合素亚基的染色更加弥散，这表明整合素聚集的抑制可能导致这些细胞粘附性降低且运动性增强。令人惊讶的是，在诱导GnT - V前后从HT1080细胞纯化的α(5)亚基上未检测到N - 连接β1,6分支的表达；然而，相比之下，β(1)亚基显示出基础水平的β1,6分支，在诱导GnT - V后大大增加。这些结果表明，肿瘤发生过程中发生的N - 连接β1,6分支变化通过调节整合素聚集和随后的信号转导途径改变细胞 - 基质粘附和迁移。这些效应很可能是由于β(1)整合素亚基上N - 连接碳水化合物表达的改变所致。
