HIV结构蛋白p24与三基序蛋白22胞内定位及出胞研究

Intracellular localization and exocytosis of HIV structural protein p24 conjugated with TRIM22

比较p24-DsRed1、p24-DsRed2、p24-DsRed-Monomer三种荧光蛋白的表达强弱,观察p24-DsRed1与TRIM22-EGFP结合物在细胞中的分布特点。构建pDsRed1-N1-p24及pDsRed2-N1-p24真核表达载体,转染HEK293T细胞,经荧光显微镜比较三种p24-DsRed荧光偶联蛋白的表达强度。选择荧光强度最强的pDsRed1-N1-p24与pEGFP-N3-TRIM22共转染HEK293T细胞,荧光显微镜观察两者结合物在细胞中的分布特点。结果显示,经荧光显微镜检测,转染48h后红色荧光蛋白强度由大到小依次为p24-DsRed1、p24-DsRed2、p24-DsRedm。在HEK293T细胞中共转染p24-DsRed1和TRIM22-EGFP,发现两者存在共定位关系,有意思的是两者的结合体能以出胞形式脱离细胞。结果表明,p24-DsRed1荧光蛋白强度显著高于p24-DsRed2和p24-DsRedm,TRIM22-p24结合物能以出芽形式脱离细胞。

This article aimed to compare the fluorescence intensity of three kinds of HIV capsid p24-DsRed fusion protein and to observe the budding of conjugates of TRIM22 with p24. Plamids pDsRed1-N1-p24 and pDsRed2-N1-p24 were constructed and transfected into HEK293T cells respectively to compare the fluorescence intensity with pDsRedl-N1-Monomer-p24 by a fluo- rescence microscope. Subsequently, plamids pEGFP-N3-TRIM22 and pDsRedl-N1-p24 were co-transfected into HEK293T cells to find out whether TRIM22 could colocalize with p24. Forty-eight hours after transfection, it was showed that the inten- sity of red fluorescence fusion protein,in descending order, was p24-DsRedl, p24-DsRed2, p24-DsRedm. When pEGFP-N3- TRIM22 and pDsRed1-N1-p24 were cotransfected, fluorescence microscopy revealed that TRIM22-EGFP was colicalized with p24-DsRed1 in HEK293T cells. Interestingly, the conjugates of TRIM22 with p24 were released by budding. In conclusion, the fluorescence intensity of p24-pDsRedl was significantly higher than the other two p24-pDsRed proteins and the conjugates of TRIM22-p24 could be released by budding.