Rab18 and Rab43 have key roles in ER-Golgi trafficking
Rab18 and Rab43 have key roles in ER-Golgi trafficking
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DOI：
10.1242/jcs.021808
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发表时间：
2008-08-15
影响因子：
4
通讯作者：
Presley, John F.
中科院分区：
文献类型：
--
作者：
Dejgaard, Selma Y.;Murshid, Ayesha;Presley, John F.
Rabs和Arfs/Arls是Ras相关的小GTP酶，与膜运输特别相关。据认为，这些蛋白质通过与外壳蛋白、马达蛋白、系链蛋白和SNARE蛋白的相互作用来调节特定的通路。我们筛选了一个全面的列表的Arf/Arl/Rab蛋白，以前确定的纯化高尔基体膜上的蛋白质组学方法（共37），高尔基体或内高尔基体定位，显性阴性和过表达表型。进一步分析这些蛋白质，Rab 18和Rab 43，强烈表明ER-高尔基体运输的作用。Rab 43-T32 N将高尔基体元件重新分配到ER出口位点，而不阻断分泌标记物VSVG-GFP从ER到细胞表面的运输。野生型Rab 43重新分配p150（Glued）的dynactin亚基，与调节前高尔基体中间体与微管的特定作用一致。野生型GFP-Rab 18的过表达或与针对Rab 18的三种siRNA中的任何一种一起孵育严重破坏高尔基复合体并减少VSVG的分泌。在光漂白试验中，Rab 18突变体特异性地增强非COPI依赖性货物β-1，4-半乳糖基转移酶（半乳糖酶）-YFP而不是COPI依赖性货物p58-YFP从高尔基体到ER的逆行高尔基体-ER转运。Rab 18-S22 N还增强了布雷菲德菌素A诱导的ER-高尔基体融合。这项研究是第一次全面应用大规模蛋白质组学的细胞生物学的小GTP酶的分泌途径。
Rabs and Arfs/Arls are Ras-related small GTPases of particular relevance to membrane trafficking. It is thought that these proteins regulate specific pathways through interactions with coat, motor, tether and SNARE proteins. We screened a comprehensive list of Arf/Arl/Rab proteins, previously identified on purified Golgi membranes by a proteomics approach (37 in total), for Golgi or intra-Golgi localization, dominant-negative and overexpression phenotypes. Further analysis of two of these proteins, Rab18 and Rab43, strongly indicated roles in ER-Golgi trafficking. Rab43-T32N redistributed Golgi elements to ER exit sites without blocking trafficking of the secretory marker VSVG-GFP from ER to cell surface. Wild-type Rab43 redistributes the p150(Glued) subunit of dynactin, consistent with a specific role in regulating association of pre-Golgi intermediates with microtubules. Overexpression of wild-type GFP-Rab18 or incubation with any of three siRNAs directed against Rab18 severely disrupts the Golgi complex and reduces secretion of VSVG. Rab18 mutants specifically enhance retrograde Golgi-ER transport of the COPI-independent cargo beta-1,4-galactosyltransferase (Galtase)-YFP but not the COPI-dependent cargo p58-YFP from the Golgi to ER in a photobleach assay. Rab18-S22N also potentiated brefeldin-A-induced ER-Golgi fusion. This study is the first comprehensive application of large-scale proteomics to the cell biology of small GTPases of the secretory pathway.