两种存活蛋白变异剪接体在HeLa细胞中的表达定位及相互作用

存活蛋白(survivin)是重要的肿瘤相关抗原基因，在肿瘤的发生发展中 起着重要的作用. 除了标准的剪接形式外，它至少还编码2种变异剪接产物—存活蛋白-2B 和存活蛋白-ΔEx3，这2个变异剪接体所编码的蛋白具有不同的生物学功能.为研究这2个变 异剪接体在肿瘤细胞中的相互作用情况，本实验利用增强型青色荧光蛋白（enhanced cyan fluorescent protein, ECFP）和增强型黄色荧光蛋白（enhanced yellow fluorescent protein, EYFP）分别标记存活蛋白-2B 和存活蛋白-ΔEx3.首先通过激光共聚焦扫描显微镜观 察它们的细胞定位；同时利用荧光共振能量转移（fluorescence resonance energy transf er, FRET）技术研究两者在细胞内的相互作用情况.研究结果表明, 存活蛋白-2B主要分布 在细胞质中，而存活蛋白-ΔEx3则主要分布在细胞核内，少量分布在细胞质中；FRET分析结 果显示,两者仅在细胞质中存在着很弱的相互作用，表明两者很可能是通过某种间接的方式发 挥功能上的相互调节作用.本研究为进一步探讨存活蛋白变异剪接体的生物学功能及相互作用 机制奠定了基础.

Expression and Location of Two Survivin Splice Variants and Their Interaction in HeLa Cells

Survivin is one of tumor associated antigen genes that plays an important role in cancer development. Survivin-2B and survivin-ΔEx3 are the two normal splice variants (isoforms) encoding two different proteins. To investigate the interaction between the two isoforms, survivin-2B was labeled with enhanced cyan fluorescent protein (ECFP) and survivin-ΔEx3 was labeled with enhanced yellow fluorescent protein (EYFP). The expression and subcellular localization of survivin-2B and survivin-ΔEx3 were observed with a confocal laser scanning microscope (CLSM), and their intermolecular interaction was quantified by fluorescence resonance energy transfer (FRET) method. It was shown that survivin-2B expressed in the cytoplasm, but most of the survivin-ΔEx3 localized in the nucleus. Very weak intermolecular interaction between the two isoforms was observed in the cytoplasm. These results might provide useful materials for further clarifying the biological function and the interactions of survivin-2B and survivin-ΔEx3.