Glycosylation status of endogenous CFTR does not affect cAMP-stimulated Cl- secretion in epithelial cells.
Glycosylation status of endogenous CFTR does not affect cAMP-stimulated Cl- secretion in epithelial cells.
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内源性CFTR的糖基化状态不影响上皮细胞中cAMP刺激的Cl-分泌。
DOI：
10.1152/ajpcell.1993.265.3.c688
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发表时间：
1993
期刊：
影响因子：
--
通讯作者：
Frizzell,RA
中科院分区：
文献类型：
--
作者：
Morris,AP;Cunningham,SA;Benos,DJ;Frizzell,RA
囊性纤维化 (CF) 损害上皮组织的 Cl- 分泌，由 N 连接糖蛋白（囊性纤维化跨膜电导调节因子 (CFTR)）突变引起。我们使用寡糖加工抑制剂修改了 CFTR 的糖基化模式，并确定了它们对人结肠 (HT-29) 细胞系中激动剂诱导的 Cl 分泌和 CFTR 定位的影响。在极化和非极化 HT-29 细胞中，使用针对 CFTR 的单克隆抗体对细胞提取物进行免疫沉淀，得到 170 kDa 处的单条带。 N-连接糖基化抑制剂降低了该条带的分子量：苦马豆素降低了 10 kDa，脱氧甘露尻霉素降低了 30 kDa，脱氧野尻霉素降低了 10-20 kDa。然而，腺苷3',5'-环单磷酸(cAMP) 或Ca(2+) 依赖性促分泌剂刺激的跨上皮Cl- 电流和电导不受影响。在极化细胞中，CFTR 位于顶膜域。用糖加工抑制剂处理单层细胞并不影响 CFTR 的位置。因此，在内源性表达CFTR的人结肠细胞中，CFTR糖基化的程度不影响CFTR对顶膜结构域的靶向或其作为激动剂刺激的Cl-通道的功能。
Cystic fibrosis (CF) impairs Cl- secretion across epithelial tissues and is caused by mutations in an N-linked glycoprotein, the cystic fibrosis transmembrane conductance regulator (CFTR). We modified the glycosylation pattern of CFTR using inhibitors of oligosaccharide processing and determined their effects on both agonist-induced Cl- secretion and CFTR location in human colon (HT-29) cell lines. In both polarized and unpolarized HT-29 cells, immunoprecipitation of cell extracts using a monoclonal antibody against CFTR gave a single band at 170 kDa. Inhibitors of N-linked glycosylation reduced the molecular mass of this band: swainsonine by 10 kDa, deoxymannojirimycin by 30 kDa, and deoxynojirimycin by 10-20 kDa. However, the transepithelial Cl- current and conductance stimulated by adenosine 3',5'-cyclic monophosphate (cAMP)- or Ca(2+)-dependent secretagogues was not affected. In the polarized cells, CFTR was localized in the apical membrane domain. Treatment of the monolayers with glycoprocessing inhibitors did not affect CFTR's location. Thus, in human colonocytes that endogenously express CFTR, the extent of CFTR glycosylation does not influence the targeting of CFTR to the apical membrane domain or its function as an agonist-stimulated Cl- channel.