BACKGROUND & AIMS:
:The small GTPase Rab6 regulates retrograde membrane traffic from endosomes to the Golgi apparatus and from the Golgi to the endoplasmic reticulum (ER). We examined the role of a Rab6-binding protein, TMF/ARA160 (TATA element modulatory factor/androgen receptor-coactivator of 160 kDa), in this process. High-resolution immunofluorescence imaging revealed that TMF signal surrounded Rab6-positive Golgi structures and immunoelectron microscopy revealed that TMF is concentrated at the budding structures localized at the tips of cisternae. The knockdown of either TMF or Rab6 by RNA interference blocked retrograde transport of endocytosed Shiga toxin from early/recycling endosomes to the trans-Golgi network, causing missorting of the toxin to late endosomes/lysosomes. However, the TMF knockdown caused Rab6-dependent displacement of N-acetylgalactosaminyltransferase-2 (GalNAc-T2), but not beta1,4-galactosyltransferase (GalT), from the Golgi. Analyses using chimeric proteins, in which the cytoplasmic regions of GalNAc-T2 and GalT were exchanged, revealed that the cytoplasmic region of GalNAc-T2 plays a crucial role in its TMF-dependent Golgi retention. These observations suggest critical roles for TMF in two Rab6-dependent retrograde transport processes: one from endosomes to the Golgi and the other from the Golgi to the ER.
背景与目标:
: 小GTPase Rab6调节从内体到高尔基体以及从高尔基体到内质网 (ER) 的逆行膜运输。我们检查了Rab6-binding蛋白TMF/ARA160 (160 kDa的TATA元件调节因子/雄激素受体共激活因子) 在此过程中的作用。高分辨率免疫荧光成像显示TMF信号包围Rab6-positive高尔基体结构,免疫电子显微镜显示TMF集中在位于水箱尖端的萌芽结构上。RNA干扰对TMF或Rab6的敲除阻止了内吞志贺毒素从早期/循环内体到反式高尔基体网络的逆行运输,导致毒素向晚期内体/溶酶体的分类错误。然而,TMF敲低引起了N-acetylgalactosaminyltransferase-2 (GalNAc-T2) 的Rab6-dependent移位,但没有引起 β1,4-半乳糖基转移酶 (GalT) 的移位。使用嵌合蛋白进行的分析 (其中交换了GalNAc-T2和GalT的细胞质区域) 表明,GalNAc-T2的细胞质区域在其TMF依赖性高尔基体保留中起着至关重要的作用。这些观察结果表明,TMF在两个Rab6-dependent的逆行转运过程中起着关键作用: 一个从内体到高尔基体,另一个从高尔基体到ER。