Department of Cell Biology, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands.
Mol Biol Cell. 2010 Apr 1;21(7):1293-304. doi: 10.1091/mbc.e09-09-0767. Epub 2010 Feb 3.
Polarized epithelial cells develop and maintain distinct apical and basolateral surface domains despite a continuous flux of membranes between these domains. The Na(+)/H(+)exchanger NHE6 localizes to endosomes but its function is unknown. Here, we demonstrate that polarized hepatoma HepG2 cells express an NHE6.1 variant that localizes to recycling endosomes and colocalizes with transcytosing bulk membrane lipids. NHE6.1 knockdown or overexpression decreases or increases recycling endosome pH, respectively, and inhibits the maintenance of apical, bile canalicular plasma membranes and, concomitantly, apical lumens. NHE6.1 knockdown or overexpression has little effect on the de novo biogenesis of apical surface domains. NHE6.1 knockdown does not inhibit basolateral-to-apical transcytosis of bulk membrane lipids, but it does promote their progressive loss from the apical surface, leaving cells unable to efficiently retain bulk membrane and bile canalicular proteins at the apical surface. The data suggest that a limited range of endosome pH mediated by NHE6.1 is important for securing the polarized distribution of membrane lipids at the apical surface and maintenance of apical bile canaliculi in HepG2 cells and hence cell polarity. This study underscores the emerging role of the endosomal recycling system in apical surface development and identifies NHE6 as a novel regulatory protein in this process.
极化的上皮细胞在这些区域之间持续不断的膜通量的情况下，仍然会发展和维持明显的顶端和基底外侧表面结构域。Na(+)/H(+)交换器 NHE6 定位于内体，但它的功能尚不清楚。在这里，我们证明极化的肝癌 HepG2 细胞表达一种定位于再循环内体并与转胞吞作用的大膜脂质共定位的 NHE6.1 变体。NHE6.1 敲低或过表达分别降低或增加再循环内体 pH 值，并抑制顶端、胆小管质膜的维持，同时也抑制顶端腔。NHE6.1 敲低或过表达对顶端表面结构域的从头生物发生几乎没有影响。NHE6.1 敲低不抑制大膜脂质的基底外侧到顶端的转胞吞作用，但它确实促进它们从顶端表面逐渐丢失，使细胞无法有效地在顶端表面保留大膜和胆小管蛋白。数据表明，由 NHE6.1 介导的内体 pH 的有限范围对于确保膜脂质在顶端表面的极化分布以及维持 HepG2 细胞中的顶端胆小管和因此细胞极性是重要的。本研究强调了内体再循环系统在顶端表面发育中的新兴作用，并确定 NHE6 为该过程中的一种新型调节蛋白。
