Multiple Rab GTPase Binding Sites in GCC185 Suggest a Model for Vesicle Tethering at the Trans-Golgi
Multiple Rab GTPase Binding Sites in GCC185 Suggest a Model for Vesicle Tethering at the Trans-Golgi
复制标题
DOI：
10.1091/mbc.e08-07-0740
复制
发表时间：
2009-01-01
影响因子：
3.3
通讯作者：
Pfeffer, Suzanne R.
中科院分区：
文献类型：
--
作者：
Hayes, Garret L.;Brown, Frank C.;Pfeffer, Suzanne R.
GCC185是一种反式高尔基体网络定位蛋白，被认为具有长螺旋结构，是依赖Rab9将甘露糖6-磷酸受体(MPR)循环到高尔基体和通过CLAP蛋白在高尔基体形成微管所必需的。GCC185通过在其C末端附近与Rab6和ARL1 GTP酶协同作用而定位于高尔基体。我们在这里通过酵母双杂交和直接生化测试表明，GCC185包含至少四个额外的结合位点，在其整个长度上多达14种不同的Rab GTP酶。一个中心螺旋线圈结构域包含一个特定的Rab9结合位点，功能分析表明该结构域对于MPR循环到高尔基复合体是重要的。在培养的细胞中使用小干扰RNA耗尽GCC185后，通过质粒挽救确定GCC185的功能也需要N-末端螺旋线圈。高尔基体-Rab结合位点可能允许GCC185促进高尔基体池的堆积和侧向相互作用，并帮助其发挥小泡系绳的作用。
GCC185, a trans-Golgi network-localized protein predicted to assume a long, coiled-coil structure, is required for Rab9-dependent recycling of mannose 6-phosphate receptors (MPRs) to the Golgi and for microtubule nucleation at the Golgi via CLASP proteins. GCC185 localizes to the Golgi by cooperative interaction with Rab6 and Arl1 GTPases at adjacent sites near its C terminus. We show here by yeast two-hybrid and direct biochemical tests that GCC185 contains at least four additional binding sites for as many as 14 different Rab GTPases across its entire length. A central coiled-coil domain contains a specific Rab9 binding site, and functional assays indicate that this domain is important for MPR recycling to the Golgi complex. N-Terminal coiled-coils are also required for GCC185 function as determined by plasmid rescue after GCC185 depletion by using small interfering RNA in cultured cells. Golgi-Rab binding sites may permit GCC185 to contribute to stacking and lateral interactions of Golgi cisternae as well as help it function as a vesicle tether.