新型人雌激素受体(hERα)变异体的克隆及在乳腺癌细胞MDA-MB-231中的核定位

Cloning of a New Human Estrogen Receptor α(hERα) Splice Variant and Its Localization in the Nuclear of Breast Cancer Cell MDA-MB-231

本研究通过提取人雌激素受体(hERα)阴性的人乳腺癌组织RNA,半巢式RT-PCR扩增获得一种新的hERα变异体ERα30的完整编码序列,并构建pEGFP-N1-ERα30融合表达载体转染人乳腺癌细胞MDA-MB-231,荧光显微镜观察ERα30的表达及亚细胞定位。结果表明,ERα30的开放阅读框编码271个氨基酸的蛋白质,预测分子量为30 kD,该变体缺乏LBD/AF2结构域,在其C端具有一段特殊的10个氨基酸的结构域;ERα30-EGFP融合蛋白在MDA-MB-231细胞的表达主要分布于细胞核;对ERα30的深入研究有望为乳腺癌的基础与临床研究中有关雌激素效应和ERα功能的问题提供更多的解释。

In this research, the complete coding sequence of a new human estrogen receptor α （hERα） splice variant named ERα30 was cloned by half-nested RT-PCR by using the extraction of hERa negative breast cancer tissue RNA, afterwards, the pEGFP-N1-ERα30 eukaryotic expression vector was constructed to transfect breast cancer cell MDA-MB-231, and the expression of ERα30 and subcellular localization were observed by fluorescent microscope. The results showed that the open-reading frame （ORF） of ERα30 encoded a protein which included 271 amino acids with a predicted molecular weight of 30 kD. This variant lacked LBD/AF2 domain and there was a special domain of 10 amino acids at its C terminal. The expression of ERα30-EGFP fusion protein was mainly in the nuclear of MDA-MB-231. Further researches about ERα30 might provide much more explanations for the issues of ERα＇s function in basic and clinical studies about breast cancer.