The analysis of the transport vesicle tethering mechanism in the Golgi apparatus.

蛋白质在细胞器之间的转运是由转运囊泡介导的。转运囊泡定位到正确的膜室是一个由系留、对接和融合组成的多层次过程。高保真的囊泡转移对于细胞器结构的建立和维持以及生物合成运输至关重要。系缚是囊泡与其靶膜在融合前接触的第一步。最近已经确定了这一步骤中的许多因素。作为候选系留因子的蛋白质可分为两大类；一种是长螺旋蛋白，另一种是多亚基复合物。在高尔基膜内转运过程中，前者是p115 /GM 130/巨肽（称为高尔金斯）复合物，后者是保守的低聚高尔基（COG）复合物。p115通过与另外两种高尔基膜上的高尔基巨球蛋白和高尔基膜上的GM130相互作用将COPI囊泡拴在高尔基膜上。COG复合体包含8个亚基（COG 1-8）。表达突变COG1或COG2的细胞表现出异常的糖基化。这两组复合体被并行提出作为候选系留因子；因此，它们是单独行动还是合作行动还有待解决。在这项研究中，我展示了两组复杂的相互作用。通过酵母双杂交筛选，鉴定出COG2为p115结合蛋白。p115与COG2在体内和体外均有相互作用。在p115缺失的细胞中，缺乏COG2结合域的p115的表达导致核周区域形成不规则的高尔基结构，包括管状和囊状结构的融合。这些结果表明p115和COG复合物之间的相互作用对于正常高尔基带的形成是必不可少的。

Protein transport between organelle is mediated by transport vesicles. Targeting of transport vesicles to the correct membrane compartment is multi-layered processes consist of tethering, docking and fusion. High fidelity vesicle transfer is essential for the establishment and maintenance of organelle structure and biosynthetic transport.Tethering is the first step of contact between a vesicle and its target membrane prior to fusion. Recently many factors in this step have been identified. Proteins proposed as candidate tethering factors fall into two general classes; one belongs to a class of long coiled-coil proteins, and the other is multi-subunit complexes. In the intra-Golgi membrane transport process, the former is the p 1 15/GM 130/giantin (called golgins) complex and the latter is the conserved oligomeric Golgi (COG) complex. p115 tethers COPI vesicles to the Golgi membrane by interaction with two other golgins giantin on the vesicles and GM130 on the Golgi membrane. The COG complex contains eight subunits (COG 1-8). Cells expressing mutant COG1 or COG2 show abnormal glycosylation. These two sets of complex are proposed in parallel as candidate tethering factors; therefore it is remain to resolve whether they act separately or cooperatively.In this study I have demonstrated two sets of complex interact each other. COG2 was identified as a p 115 binding protein by the yeast two-hybrid screening. The interaction of p 115 and COG2 was showed both in vitro and in vivo.The expression of p115 lacking COG2 binding domain-in p115-depleted cells resulted in the formation of an irregular Golgi structure consist of converged tubular and vesicular structures in the perinuclear area. These results suggest that the interaction between p 115 and the COG complex is Essential for normal Golgi ribbon formation.