Okamoto Curtis T, Li Rui, Zhang Zhuo, Jeng Young Y, Chew C S
Department of Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles, CA 90089-9121, USA.
J Control Release. 2002 Jan 17;78(1-3):35-41. doi: 10.1016/s0168-3659(01)00479-5.
The characterization of endocytotic and post-endocytotic trafficking pathways at the apical membrane of epithelial cells presents a potential avenue for the identification of targets to modulate the initial stages of absorption and transepithelial transport of macromolecules. In addition, it is becoming increasingly clear that the activity of a number of apical membrane transporters is acutely regulated by vesicular trafficking. The gastric HCl-secreting parietal (oxyntic) cell is a model system to characterize an apical membrane vesicular trafficking pathway and its relationship to the regulation of the function of the gastric proton pump. The subapical tubulovesicular compartment of the parietal cell is highly enriched in the H,K-ATPase and is a key endosomal-like system in the apical membrane recycling pathway. In the process of cataloging the proteins that interact with the H,K-ATPase and tubulovesicles, we have identified novel components that may regulate protein sorting through this compartment and candidate linker proteins between the vesicular trafficking machinery and the cytoskeleton. One protein associated with H,K-ATPase-rich tubulovesicles is the nonreceptor tyrosine kinase c-src, identified by a screen for dynamin-binding proteins. The tyrosine kinase is active, as it can tyrosine-phosphorylate tubulovesicular proteins in vitro. One of the tyrosine-phosphorylated proteins of M(r) 100 kDa may be the H,K-ATPase itself, or a protein in a complex with the H,K-ATPase that is stable to dissociation by nonionic detergents. By virtue of its association with tubulovesicular membranes, c-src may regulate the trafficking and/or activity of the H,K-ATPase. A second protein identified by a screen for dynamin-binding proteins is the protein lasp-1. Lasp-1, through its modular protein structure, may bind to dynamin and to the actin cytoskeleton, thus linking the vesicular trafficking machinery with the cytoskeleton. These two examples illustrate the utility of the parietal cell in the biochemical characterization of components potentially involved in the regulation of apical membrane trafficking pathways.
上皮细胞顶端膜的内吞及内吞后运输途径的特征，为识别调控大分子吸收和跨上皮运输初始阶段的靶点提供了一条潜在途径。此外，越来越清楚的是，许多顶端膜转运蛋白的活性受到囊泡运输的严格调控。胃盐酸分泌壁细胞是一个用于表征顶端膜囊泡运输途径及其与胃质子泵功能调控关系的模型系统。壁细胞的顶端下微管泡区室富含H，K - ATP酶，是顶端膜再循环途径中的关键内体样系统。在对与H，K - ATP酶和微管泡相互作用的蛋白质进行分类的过程中，我们鉴定出了可能调控通过该区室的蛋白质分选的新成分，以及囊泡运输机制与细胞骨架之间的候选连接蛋白。一种与富含H，K - ATP酶的微管泡相关的蛋白质是非受体酪氨酸激酶c - src，它是通过筛选动力蛋白结合蛋白而鉴定出来的。该酪氨酸激酶具有活性，因为它能在体外使微管泡蛋白发生酪氨酸磷酸化。一种分子量为100 kDa的酪氨酸磷酸化蛋白可能是H，K - ATP酶本身，或者是与H，K - ATP酶形成复合物且对非离子去污剂解离稳定的一种蛋白质。由于其与微管泡膜的结合，c - src可能调控H，K - ATP酶的运输和/或活性。通过筛选动力蛋白结合蛋白鉴定出的第二种蛋白质是lasp - 1蛋白。Lasp - 1通过其模块化的蛋白质结构，可能与动力蛋白和肌动蛋白细胞骨架结合，从而将囊泡运输机制与细胞骨架连接起来。这两个例子说明了壁细胞在对可能参与顶端膜运输途径调控的成分进行生化表征方面的实用性。
