Hierarchy of mechanisms involved in generating Na/K-ATPase polarity in MDCK epithelial cells.
Hierarchy of mechanisms involved in generating Na/K-ATPase polarity in MDCK epithelial cells.
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DOI：
10.1083/jcb.130.5.1105
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发表时间：
1995-09
期刊：
影响因子：
--
通讯作者：
Nelson WJ
中科院分区：
文献类型：
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作者：
Mays RW;Siemers KA;Fritz BA;Lowe AW;van Meer G;Nelson WJ
我们研究了Na/ k - atp酶在两个MDCK II细胞克隆的基底-外侧膜中产生极化分布的机制。两个克隆在稳定状态下均表现出尖膜和基侧膜标记蛋白的极化分布，包括Na/K- atp酶。然而，新合成的Na/ k - atp酶从高尔基复合体传递到一个无性系（II/J）的顶膜和基侧膜，以及另一个无性系（II/G）的基侧膜；Na/ k - atp酶选择性地保留在基侧膜上，从而在两个克隆中产生完整的细胞表面极性。另一种基底-外侧膜蛋白E-钙粘蛋白在两个MDCK克隆中都被分选到基底-外侧膜上，这表明克隆II/J细胞中基底-外侧膜蛋白没有普遍的分选缺陷。在克隆II/G细胞中，糖基磷脂酰肌醇（GPI）锚定蛋白（GP-2）和鞘糖脂（葡萄糖神经酰胺，glcer）优先被转运到根尖膜，但在克隆II/J细胞中，GP-2和glcer同样被转运到根尖膜和基侧膜，类似于Na/ k - atp酶。为了检验glcer、GP-2和Na/K- atp酶的分选之间的明显相互关系，真菌代谢物伏马菌素B1 （FB1）抑制了克隆II/G细胞鞘脂合成。在FB1存在的情况下，GP-2和Na/ k - atp酶被传递到顶膜和基侧膜，类似于克隆II/J细胞；FB1对II/G细胞基底-外侧膜上e -钙粘蛋白的分选无影响。添加外源性神经酰胺，以绕过FB1阻滞，分别恢复GP-2和Na/K- atp酶对顶膜和基侧膜的分选。这些结果表明，Na/ k - atp酶的完全细胞表面极性的产生涉及高尔基复体和质膜中的分层分选机制，并且MDCK细胞高尔基复体中的Na/ k - atp酶分选可能通过排除顶端途径来调节。这些结果也为其他顶端和基底外侧膜蛋白的分选途径提供了新的见解。
We have studied mechanisms involved in generating a polarized distribution of Na/K-ATPase in the basal-lateral membrane of two clones of MDCK II cells. Both clones exhibit polarized distributions of marker proteins of the apical and basal-lateral membranes, including Na/K- ATPase, at steady state. Newly synthesized Na/K-ATPase, however, is delivered from the Golgi complex to both apical and basal-lateral membranes of one clone (II/J), and to the basal-lateral membrane of the other clone (II/G); Na/K-ATPase is selectively retained in the basal- lateral membrane resulting in the generation of complete cell surface polarity in both clones. Another basal-lateral membrane protein, E- cadherin, is sorted to the basal-lateral membrane in both MDCK clones, demonstrating that there is not a general sorting defect for basal- lateral membrane proteins in clone II/J cells. A glycosyl- phosphatidylinositol (GPI)-anchored protein (GP-2) and a glycosphingolipid (glucosylceramide, GlcCer) are preferentially transported to the apical membrane in clone II/G cells, but, in clone II/J cells, GP-2 and GlcCer are delivered equally to both apical and basal-lateral membranes, similar to Na/K-ATPase. To examine this apparent inter-relationship between sorting of GlcCer, GP-2 and Na/K- ATPase, sphingolipid synthesis was inhibited in clone II/G cells with the fungal metabolite, Fumonisin B1 (FB1). In the presence of FB1, GP-2 and Na/K-ATPase are delivered to both apical and basal-lateral membranes, similar to clone II/J cells; FB1 had no effect on sorting of E-cadherin to the basal-lateral membrane of II/G cells. Addition of exogenous ceramide, to circumvent the FB1 block, restored GP-2 and Na/K- ATPase sorting to the apical and basal-lateral membranes, respectively. These results show that the generation of complete cell surface polarity of Na/K-ATPase involves a hierarchy of sorting mechanisms in the Golgi complex and plasma membrane, and that Na/K-ATPase sorting in the Golgi complex of MDCK cells may be regulated by exclusion from an apical pathway(s). These results also provide new insights into sorting pathways for other apical and basal-lateral membrane proteins.