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首页> 外文期刊>Molecular and Cellular Biology >Cloning, characterization, and expression of a novel GDP dissociation inhibitor isoform from skeletal muscle.
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Cloning, characterization, and expression of a novel GDP dissociation inhibitor isoform from skeletal muscle.

机译：从骨骼肌中克隆，表征和表达新型GDP离解抑制剂同工型。

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Cellular mechanisms for controlling membrane trafficking appear to involve small GTP-binding proteins such as the Rab proteins. Rab function is regulated by GDP dissociation inhibitor (GDI), which releases Rab proteins from membranes and inhibits GDP dissociation. Here we report the isolation of a full-length cDNA encoding a novel GDI isoform of 445 amino acids (GDI-2) with a deduced molecular weight of 50,649 from mouse skeletal muscle. Full-length and partial cDNA clones encoding a previously reported GDI protein (GDI-1) were also isolated from cDNA libraries prepared from rat brain and mouse skeletal muscle, respectively. The degree of deduced amino acid sequence identity between mouse GDI-2 and our mouse GDI-1 cDNA clone is 86%. Northern (RNA blot) analysis revealed that in human tissues, both GDI-1 and GDI-2 transcripts were abundant in brain, skeletal muscle, and pancreas but were weakly expressed in heart and liver. GDI-1 mRNA was expressed in kidney, whereas GDI-2 was almost absent, while in lung the relative amounts of these mRNA species were reversed. Specific antibodies against mouse GDI-1 and GDI-2 based on unique peptide sequences in the proteins were raised. Differentiation of 3T3-L1 fibroblasts into highly insulin-responsive adipocytes was accompanied by large increases in both mRNA and protein levels of GDI-1 and GDI-2. GDI-1 and GDI-2 expressed as glutathione S-transferase fusion proteins were both able to solubilize the membrane-bound forms of Rab4 and Rab5 in a GDP/GTP-dependent manner. Taken together, these data demonstrate that the protein products of at least two genes regulate the membrane dynamics of Rab proteins in mice.
机译：控制膜运输的细胞机制似乎涉及小的GTP结合蛋白，例如Rab蛋白。 Rab功能由GDP解离抑制剂（GDI）调节，GDI可从膜上释放Rab蛋白并抑制GDP解离。在这里，我们报告从小鼠骨骼肌中分离出全长cDNA，全长cDNA编码445个氨基酸的新型GDI同工型（GDI-2），推导分子量为50,649。还分别从大鼠脑和小鼠骨骼肌制备的cDNA文库中分离了编码先前报道的GDI蛋白（GDI-1）的全长和部分cDNA克隆。小鼠GDI-2和我们的小鼠GDI-1 cDNA克隆之间推导的氨基酸序列同一性的程度为86％。 Northern（RNA印迹）分析显示，在人体组织中，GDI-1和GDI-2转录本在大脑，骨骼肌和胰腺中均含量很高，但在心脏和肝脏中却很少表达。 GDI-1 mRNA在肾脏中表达，而GDI-2几乎不存在，而在肺中这些mRNA种类的相对数量却相反。基于蛋白质中独特的肽序列，产生了针对小鼠GDI-1和GDI-2的特异性抗体。 3T3-L1成纤维细胞向高度胰岛素反应性脂肪细胞的分化伴随着GDI-1和GDI-2的mRNA和蛋白水平的大幅增加。表示为谷胱甘肽S-转移酶融合蛋白的GDI-1和GDI-2都能够以GDP / GTP依赖性的方式溶解Rab4和Rab5的膜结合形式。综上所述，这些数据证明至少两个基因的蛋白质产物调节小鼠中Rab蛋白的膜动力学。

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