Abstract：Abstract: To observe the subcellular localization of full-length and splicing variants of human SMAD4 and analyze their transcriptional activity, pEGFP-C1 recombinant plasmids containing SMAD4 full-length gene and splicing variants were transfected separately into HEK-293T cells, and the subcellular localization of the GFP-tagged SMAD4 and its splicing variants was observed. Activation of SMAD binding element (SBE) by full-length SMAD4 and splicing variants in HEK-293T cells was compared using dual-luciferase reporter experiment. The fluorescence results showed that the full length SMAD4 and five kinds of SMAD4 splicing variants, including SMAD4△4, SMAD4△6, SMAD4△5-6, SMAD4△4-6 and SMAD4△4-7, were mainly distributed in the cytoplasm, while the GFP-tagged SMAD4△3 was mainly localized in the nucleus, compa-red with the uniform distribution of GFP across both nucleus and cytoplasm in pEGFP-C1 empty vectortrans-fected group. The dual-luciferase reporter experiment showed that, among the six variants, only SMAD4△3 could up-regulate the luciferase activity of the SBE reporter gene when compared with the vector group; however, the full-length SMAD4 can significantly up-regulate the luciferase activity of the SBE reporter gene when compared with SMAD4△3. In summary, observation of subcellular localization and analysis of the transcriptional activity of full-length human SMAD4 and its splicing variants could provide the basis for further functional study of SMAD4 splicing variants.
引用本文:
方晋仁, 尹小慧, 周 涛, 李国庆, 秦 辉, 杨南扬. 人SMAD4全长及剪接体在HEK-293T细胞中的亚细胞定位及转录活性分析[J]. 生命科学研究, 2021, 25(2): 95-101.
FANG Jin-ren, YIN Xiao-hui, ZHOU Tao, LI Guo-qing, QIN Hui, YANG Nan-yang. Subcellular Localization and Transcriptional Activity Analysis of Full-length Human SMAD4 and Its Splicing Variants in HEK-293T Cells. Life Science Research, 2021, 25(2): 95-101.
