2-HYBRID SYSTEM SCREEN WITH THE SMALL GTP-BINDING PROTEIN RAB6 - IDENTIFICATION OF A NOVEL MOUSE GDP DISSOCIATION INHIBITOR ISOFORM AND 2 OTHER POTENTIAL PARTNERS OF RAB6
2-HYBRID SYSTEM SCREEN WITH THE SMALL GTP-BINDING PROTEIN RAB6 - IDENTIFICATION OF A NOVEL MOUSE GDP DISSOCIATION INHIBITOR ISOFORM AND 2 OTHER POTENTIAL PARTNERS OF RAB6
复制标题
DOI：
10.1074/jbc.270.24.14801
复制
发表时间：
1995-06-16
影响因子：
4.8
通讯作者：
GOUD, B
中科院分区：
文献类型：
--
作者：
JANOUEIXLEROSEY, I;JOLLIVET, F;GOUD, B
Rab 6是一种小的GTP结合蛋白，属于Ras超家族，参与高尔基体内转运。使用双杂交系统筛选小鼠脑cDNA文库，我们已经分离出几个克隆编码与Rab 6相互作用的蛋白质。约60%的克隆鉴定编码新的小鼠Rab GDP解离抑制剂（GDI）亚型。该GDI同种型不同于先前已表征的小鼠mGDI-1和mGDI-2，并且最可能代表大鼠Rab GDI β同种型的小鼠对应物。在双杂交系统中，GDI β与野生型Rab 6和Rab 5相互作用，但不与GTP结合的Rab 6突变体或不能后处理的Rab 6突变体相互作用。我们进一步研究了小鼠GDI β是否具有功能;我们发现重组小鼠GDI β能够从膜上去除几种Rab蛋白，包括Rab 1，Rab 2，Rab 4和Rab 6。在小鼠中第三种GDI亚型的鉴定提出了一个问题，即GDI基因是否属于一个更大的多基因家族。我们已经表明，基因组DNA的Southern印迹分析，至少有五个GDI基因拷贝存在于小鼠和大鼠的基因组。在我们的双杂交筛选中，我们还鉴定了另一个与Rab 6特异性相互作用的克隆。对该克隆进行了部分测序，但与已知序列没有同源性。最后，第三个克隆，与Rab 5和Rab 6相互作用。似乎也编码一种新的蛋白质。
Rab6 is a small GTP-binding protein that belongs to the Ras superfamily and is involved in intra-Golgi transport. Using a two-hybrid system screen of a mouse brain cDNA library, we have isolated several clones encoding proteins that interact with Rab6. Approximately 60% of the clones identified encoded anew mouse Rab GDP dissociation inhibitor (GDI) isoform. This GDI isoform is distinct from mouse mGDI-1 and mGDI-2, which have been characterized previously, and most likely represents the mouse counterpart of the rat Rab GDI beta isoform. In the two-hybrid system, GDI beta interacts with wild-type Rab6 and Rab5, but not with a GTP-bound Rab6 mutant, or a Rab6 mutant that cannot be post-translationally processed. We further examined whether mouse GDI beta is functional; we show that recombinant mouse GDI beta is able to remove several Rab proteins, including Rab1, Rab2, Rab4, and Rab6, from membranes. The identification of a third GDI isoform in mouse raised the question whether GDI genes belong to a larger multigenic family. We have shown, by Southern blot analysis of genomic DNA, that at least five GDI gene copies exist in both the mouse and rat genomes. In our two-hybrid screen, we have also characterized another clone that specifically interacts with Rab6. This clone was partially sequenced but shows no homology to known sequences. Finally, a third clone, interacting with both Rab5 and Rab6. also appears to encode a novel protein.