中文摘要：

目的构建稳定表达谷胱甘肽S转移酶mu5（GSTM5）的人支气管上皮16HBE细胞株，探讨诱导GSTM5核转位的关键因素。方法构建表达GSTM5基因的重组慢病毒载体（PLVX-puro-GSTM5-SBP-3＊flag-N、PLVX-puro-3＊flag-SBP-GSTM5-C）并制备出相应的慢病毒，感染人支气管上皮16HBE细胞后，经嘌呤霉素筛选获得细胞克隆；实时荧光定量PCR和蛋白印迹法（Westernblot）分别检测细胞株中GSTM5的mRNA、蛋白表达水平；以肿瘤坏死因子-α（TNF-α；10ng/ml）刺激稳定转染细胞株（稳转株）0.5h，共聚焦荧光显微镜检测GSTM5核转位。结果成功构建稳定表达GSTM5人支气管上皮细胞株（16HBE-GSTM5-SBP-3＊flag-N、16HBE-3＊flag-SBP-GSTM5-C）；荧光共聚焦显微镜显示稳转株在TNF-α刺激诱导后，GSTM5由胞浆转入胞核，以16HBE-GSTM5-SBP-3＊flag-N稳转株更显著。结论TNF-α刺激可诱导GSTM5人支气管上皮细胞株中GSTM5发生核转位，可能与其N端含有非经典核定位信号密切相关。

英文摘要：

ObjectiveTo establish 16HBE cell lines stably expressing glutathione S-transferase mu 5 （GSTM5） gene, and explore the mechanism of GSTM5 nuclear translocation. MethodsRecombinant lentiviral expression vector containing GSTM5 gene was constructed and lentivirus was produced. After lentivirus infection of 16HBE cells, 16HBE-GSTM5 cell lines were obtained by screening with puromycin. Expression of GSTM5 in different cells was examined by RT-qPCR and Western blot. The nuclear translocation of GSTM5 was observed by confocal laser scanning microscope, after the 16HBE-GSTM5 cell lines were treated with tumor necrosis factor-α （TNF-α; 10 ng/ml） for 0.5 hour. ResultsLentiviral expression plasmids, PLVX-puro-3＊flag-SBP-GSTM5-C and PLVX-puro-GSTM5-SBP-3＊flag-N, were constructed and lentiviral particles were successfully packed. After infected with lentivirus and screened by puromycin, two cell lines, 16HBE-GSTM5-SBP-3＊flag-N and 16HBE-3＊flag-SBP-GSTM5-C, were obtained. GSTM5 expression in these two cell lines was significantly higher compared with the control group and parental cells. After treated with TNF-α for 0.5 hour, the nuclear translocation of GSTM5 in 16HBE-GSTM5-SBP-3＊flag-N was much more obviously than that in 16HBE-3＊flag-SBP-GSTM5-C. ConclusionThe N-terminal region of GSTM5 is critical for nuclear translocation induced by TNF-α, which is mediated by a novel and non-classical nuclear localization signal.