中文摘要：

目的研究FK506结合蛋白25（FKBP25）野生型及其突变体在真核细胞内的表达、定位及与胞内氯离子通道蛋白1（CLIC1）的共定位情况。方法以pc DNA3.1-FKBP25-FLAG为模板,分别构建以p CDGFP为载体的FKBP25野生型及其缺失突变体FKBP25（1～42 aa）、FKBP25（1～110aa）、FKBP25（43～224 aa）、FKBP25（43～110 aa）、FKBP25（111～224 aa）的真核表达质粒,并分别转染至HEK 293T细胞中,Western blot法检测重组蛋白表达情况;将上述质粒分别与带FLAG标签的CLIC1质粒共转染至COS7细胞,激光共聚焦扫描显微镜下观察、分析荧光共定位情况。结果成功构建了p CDGFP-FKBP25基因的一系列真核表达质粒;Western blot显示：p CDGFP-FKBP25及其突变体p CDGFPFKBP25（1～42 aa）、p CDGFP-FKBP25（1～110 aa）、p CDGFPFKBP25（43～110 aa）、p CDGFP-FKBP25（43～224 aa）、p CDGFP-FKBP25（111～224 aa）均能在HEK 293T细胞中有效表达;GFP-FKBP25野生型在COS7细胞中主要分布在细胞质中,细胞核中表达较少,而突变体在细胞质、细胞核均有表达,而且细胞核中的表达多于野生型。共定位实验结果表明：野生型FKBP25蛋白与CLIC1蛋白在细胞质内存在明显的共定位现象。其3个缺失突变体与CLIC1蛋白的共定位明显减少,而且共定位也存在区别：FKBP25-N与CLIC1共定位主要于细胞核中,部分分布于细胞质;FKBP25-PC和FKBP25-C与CLIC1只共定位于细胞质中。结论荧光共定位实验观察到FKBP25野生型及其3种突变体FKBP25（1～42aa）、FKBP25（43～224 aa）、FKBP25（111～224 aa）与CLIC1蛋白存在不同程度的共定位现象,为进一步揭示FKBP25的生物学功能提供了基础。

英文摘要：

Objective To investigate the expression and localization of FKBP25 including its deletion mutants FK- BP25 （ 1 - 42 aa） , FKBP25 （ 1 - 110 aa） , FKBP25 （43 - 224 aa） , FKBP25 （43 - 110 aa） , FKBP25 （ 111 - 224 aa） in eukaryotic cells, and the colocalization of FKBP25 and its mutants with CLIC1 in mammalian cells. Methods To construct some eukaryotic expression plasmids of pCDGFP-FKBP25, pCDGFP-FKBP25 （1 -42 aa） , pCDG- FP-FKBP25 （ 1 - 110 aa）, pCDGFP-FKBP25 （43 - 224 aa）, pCDGFP-FKBP25 （43 - 110 aa）, pCDGFP-FKBP25 （ 1 ! 1 - 224 aa）. These plasmids were transfected into HEK 293T cells respectively and the expression was checked by Western blot. Their localization and colocalization with FLAG-CLIC1 in COS7 cells were detected by fluores- cence microscopy. Results The expression plasmids of pCDGFP-FKBP25 and a series of its deletion mutants were constructed successfully, which could effectively express in HEK 293T and COS7 ceils. The localization of GFP- FKBP25 was mainly in cytoplasm and its mutants were in both nucleus and cytoplasm. The co-localization of its mutants with FLAG-CLLICI were much less than that of GFP-FKBP25 itself, and there were some differences in distribution. Conclusion FKBP25 and its deletion mutants can express efficiently in HEK 293T and COS7 cells. GFP-FKBP25 is mainly distributed in the cytoplasm, but the mutants are in both nucleus and cytoplasm. FKBP25 protein and its mutants appear to colocalize with CLIC1 to different degrees respectively, which implies that FK- BP25 and its mutants may interact with CLIC1 respectively in mammalian cells. It provides cellular basis to further study on biological functions of FKBP25 gene.