Monlauzeur L, Breuza L, Le Bivic A
Laboratoire de Génétique et Physiologie du Développement, UMR6545, IBDM, Faculté des Sciences de Luminy, Case 907, Université de la Méditerranée, 13288 Marseille Cedex 09, France.
J Biol Chem. 1998 Nov 13;273(46):30263-70. doi: 10.1074/jbc.273.46.30263.
We have expressed the human neurotrophin receptor p75 (p75(NTR)) in the intestinal epithelial cell line Caco-2 as a model to study intracellular transport and subcellular sorting signals in intestinal cells. p75(NTR) was localized at the apical membrane of Caco-2 cells and reached this membrane mainly via an indirect pathway. Apical localization, intracellular routing, and basolateral to apical transcytosis were not affected by truncation of the cytoplasmic domain or replacement of the transmembrane domain by a glycosyl phosphatidylinositol anchor. Removal of membrane anchoring resulted in basolateral secretion of the ectodomain of p75(NTR) in Caco-2 cells but in apical secretion in Madin-Darby canine kidney (MDCK) cells. Substitution of potential O-glycosylation sites present in the stalk of p75(NTR) led to intracellular cleavage and secretion of the ectodomain into the basolateral medium both in Caco-2 and MDCK cells. These results suggest that the stalk of p75(NTR) carries an apical sorting information that is recognized efficiently by Caco-2 cells only when attached to the membrane. This apical sorting information is linked to the presence of predicted O-glycosylation sites in that region. These putative O-glycosylation sites also play a role in the regulation of p75(NTR) transport to the cell surface and in the prevention of rapid degradation by cleavage of the stalk domain.
我们已在肠上皮细胞系Caco-2中表达人神经营养因子受体p75（p75(NTR)），以此作为研究肠细胞内细胞运输和亚细胞分选信号的模型。p75(NTR)定位于Caco-2细胞的顶端膜，主要通过间接途径到达该膜。细胞质结构域的截断或用糖基磷脂酰肌醇锚替换跨膜结构域，均不影响顶端定位、细胞内运输途径以及从基底外侧到顶端的转胞吞作用。去除膜锚定导致p75(NTR)胞外结构域在Caco-2细胞中向基底外侧分泌，但在Madin-Darby犬肾（MDCK）细胞中向顶端分泌。替换p75(NTR)柄部存在的潜在O-糖基化位点，导致胞外结构域在Caco-2和MDCK细胞中均在细胞内裂解并分泌到基底外侧培养基中。这些结果表明，p75(NTR)的柄部携带一种顶端分选信息，该信息仅在与膜相连时才能被Caco-2细胞有效识别。这种顶端分选信息与该区域预测的O-糖基化位点的存在有关。这些假定的O-糖基化位点在调节p75(NTR)向细胞表面的运输以及防止柄部结构域裂解导致的快速降解中也起作用。
