Recombinant MUC1 probe authentically reflects cell-specific O-glycosylation profiles of endogenous breast cancer mucin -: High density and prevalent core 2-based glycosylation
Recombinant MUC1 probe authentically reflects cell-specific O-glycosylation profiles of endogenous breast cancer mucin -: High density and prevalent core 2-based glycosylation
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DOI：
10.1074/jbc.m202921200
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发表时间：
2002-07-19
影响因子：
4.8
通讯作者：
Hanisch, FG
中科院分区：
文献类型：
--
作者：
M端ller, S;Hanisch, FG
有关肿瘤相关MUC1的O-连接糖链的知识主要基于酶和免疫化学证据。为了获得结构信息并克服内源性粘蛋白缺乏的局限性，我们在四个乳腺癌细胞系中表达了对应于六个MUC1串联重复序列的重组糖基化探针。2-氨基苯甲酰胺标记的葡聚糖经肼裂解和正相层析后，对O-葡聚糖谱进行了比较分析。除了O-糖链长度普遍缩短和高密度糖基化外，没有发现共同的结构模式。T47D融合蛋白的表达几乎完全从核心2转移到核心1，并以唾液酸化的糖链为主。相比之下，MCF-7、MDA-MB231和ZR75-1细胞优先糖基化MUC1重复序列，核心2-糖链主要以α3连接的唾液酸(MDA-MB231，ZR75-1)或α2/3连接的岩藻糖(MCF-7)终止。T47D和MCF-7细胞上清液中的内源MUC1O-糖基化图谱与相应的重组探针几乎相同，表明融合蛋白反映了细胞中真实的O-糖链图谱。大多数被研究的细胞的结构模式与乳腺癌MUC1O-糖基化的生物合成模型相矛盾，该模型声称，正常的基于核心2的聚乳糖胺结构被截断为基于短唾液酸化的核心1的糖链是由于形成核心2的β6-N-乙酰氨基葡萄糖转移酶的活性降低和/或竞争性α-唾液酸基转移酶的过度表达。
Knowledge about the O-linked glycan chains of tumor-associated MUC1 is primarily based on enzymatic and immunochemical evidence. To obtain structural information and to overcome limitations by the scarcity of endogenous mucin, we expressed a recombinant glycosylation probe corresponding to six MUC1 tandem repeats in four breast cancer cell lines. Comparative analyses of the O-glycan profiles were performed after hydrazinolysis and normal phase chromatography of 2-aminobenzamide-labeled glycans. Except for a general reduction in the O-glycan chain lengths and a high density glycosylation, no common structural pattern was revealed. T47D fusion protein exhibits an almost complete shift from core 2 to core 1 expression with a preponderance of sialylated glycans. By contrast, MCF-7, MDA-MB231, and ZR75-1 cells glycosylate the MUC1 repeat peptide preferentially with core 2-based glycans terminating mostly with alpha3-linked sialic acid (MDA-MB231, ZR75-1) or alpha2/3-linked fucose (MCF-7). Endogenous MUC1 from T47D and MCF-7 cell supernatants revealed almost identical O-glycosylation profiles compared with the respective recombinant probes, indicating that the fusion proteins reflected the authentic O-glycan profiles of the cells. The structural patterns in the majority of cells under study are in conflict with biosynthetic models of MUC1 O-glycosylation in breast cancer, which claim that the truncation of normal core 2-based polylactosamine structures to short sialylated core 1-based glycans is due to the reduced activity of core 2-forming beta6-N-acetylglucosaminyltransferases and/or to overexpression of competitive alpha3-sialyltransferase.