Expression and characterization of Rab38, a new member of the Rab small G protein family
Expression and characterization of Rab38, a new member of the Rab small G protein family
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DOI：
10.1515/bc.2005.018
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发表时间：
2005-02-01
影响因子：
3.7
通讯作者：
Voelker, DR
中科院分区：
文献类型：
--
作者：
Osanai, K;Takahashi, K;Voelker, DR
Rab38是rabb小G蛋白家族的新成员，调控细胞内囊泡运输。Rab38在黑素细胞中表达，并且已经明确Rab38的gtp结合区域的点突变是巧克力小鼠皮肤白化病的基因。然而，关于重组蛋白生产、细胞内定位和组织特异性表达模式的基本信息尚未报道。我们利用杆状病毒/昆虫细胞蛋白表达系统制备了重组Rab38。Triton X-114相分离和镍亲和层析相结合，得到了结合[α - p -32]-GTP的纯烯丙基化Rab38。mRNA和天然蛋白以组织特异性的方式表达，例如在肺、皮肤、胃、肝和肾中。新分离的大鼠肺泡II型细胞mRNA信号高度阳性，但随着时间的推移，信号迅速消失。免疫荧光染色表明，表达的gst标记的Rab38主要与内质网驻留蛋白共定位，也部分与内质网和高尔基复合体之间的间歇性囊泡共定位。这些结果表明，Rab38在特定组织中非普遍表达，并调节与内质网相关的早期囊泡运输，因此Rab38异常可能导致多器官疾病以及眼皮肤白化病。
Rab38 is a new member of the Rab small G protein family that regulates intracellular vesicle trafficking. Rab38 is expressed in melanocytes and it has been clarified that a point mutation in the postulated GTP-binding domain of Rab38 is the gene responsible for oculocutaneous albinism in chocolate mice. However, basic information regarding recombinant protein production, intracellular location, and tissue-specific expression pattern has not yet been reported. We produced recombinant Rab38 using a baculovirus/insect cell-protein expression system. A combination of Triton X-114 phase separation and nickel-affinity chromatography yielded exclusively prenylated Rab38 that bound [alpha-P-32]-GTP. The mRNA and the native protein were expressed in a tissue-specific manner, e.g., in the lung, skin, stomach, liver, and kidney. Freshly isolated rat alveolar type II cells were highly positive for the mRNA signal, but the signal was rapidly lost over time. Immunofluorescence staining demonstrated that expressed GST-tagged Rab38 was mainly colocalized with endoplasmic reticulum-resident protein and also partly with intermittent vesicles between the endoplasmic reticulum and the Golgi complex. These results indicate that Rab38 is expressed non-ubiquitously in specific tissues and regulates early vesicle transport relating to the endoplasmic reticulum, and hence suggest that Rab38 abnormality may cause multiple organ diseases as well as oculocutaneous albinism.