Bennett M K, Wandinger-Ness A, Simons K
European Molecular Biology Laboratory, Heidelberg, FRG.
EMBO J. 1988 Dec 20;7(13):4075-85. doi: 10.1002/j.1460-2075.1988.tb03301.x.
Mechanically perforated MDCK cells were used to study membrane transport between the trans-Golgi network and the apical and basolateral plasma membrane domains in vitro. Three membrane transport markers--an apical protein (fowl plague virus haemagglutinin), a basolateral protein (vesicular stomatitis virus G protein), and a lipid marker destined for both domains (C6-NBD-sphingomyelin)--were each accumulated in the trans-Golgi by a 20 degrees C block of transport and their behaviour monitored following cell perforation and incubation at 37 degrees C. In the presence of ATP and in the absence of calcium ions a considerable fraction of the transport markers were released from the perforated cells in sealed membrane vesicles. Control experiments showed that the vesicles were not generated by non-specific vesiculation of the Golgi complex or the plasma membrane. The vesicles had well defined sedimentation properties and the orientation expected of transport vesicles derived from the trans-Golgi network.
机械穿孔的MDCK细胞用于体外研究反式高尔基体网络与顶端和基底外侧质膜结构域之间的膜转运。三种膜转运标记物——一种顶端蛋白（禽瘟病毒血凝素）、一种基底外侧蛋白（水泡性口炎病毒G蛋白）和一种定位于两个结构域的脂质标记物（C6-NBD-鞘磷脂）——通过20℃的转运阻断分别在反式高尔基体中积累，并在细胞穿孔和37℃孵育后监测它们的行为。在有ATP且无钙离子的情况下，相当一部分转运标记物从穿孔细胞中以密封膜泡的形式释放出来。对照实验表明，这些膜泡不是由高尔基体复合体或质膜的非特异性囊泡化产生的。这些膜泡具有明确的沉降特性以及来自反式高尔基体网络的转运膜泡所预期的方向。
