The POU domain transcription factor Oct4 is a master regulator in maintaining self-renewal and pluripotency of embryonic stem （ES） cells. To further explore the functional network of Oct4, the yeast two-hybrid syste...The POU domain transcription factor Oct4 is a master regulator in maintaining self-renewal and pluripotency of embryonic stem （ES） cells. To further explore the functional network of Oct4, the yeast two-hybrid system was used to search for Oct4 interacting proteins. PH domain （containing POU domain and homeodomain） of human OCT4 was used as a bait. From the human testis cDNA library, we identified a strong interaction between OCT4 and karyopberin-alpha 2 （KPNA-2）. KPNA2 is involved in active nuclear import of proteins. This finding was confirmed by glutathione S-transferase pull-down and co-immunoprecipitation assays. The interaction between OCT4 and KPNA-2 was further mapped to multiple regions of the two proteins. In addition, we studied nuclear localization signal （NLS） of mouse Oct4 and demonstrated that it is essential for Oct4 nuclear localization. Thus, our data suggest that Oct4 nuclear localization may be mediated by its interaction with KPNA-2.展开更多
基金supported by the Shanghai Science & Technology Developmental Foundations (No.06dj14001)the National High Technology Research and Development Program of China (No. 2006AA02Z197, 2006CB943901, and 2007CB947904)
文摘The POU domain transcription factor Oct4 is a master regulator in maintaining self-renewal and pluripotency of embryonic stem （ES） cells. To further explore the functional network of Oct4, the yeast two-hybrid system was used to search for Oct4 interacting proteins. PH domain （containing POU domain and homeodomain） of human OCT4 was used as a bait. From the human testis cDNA library, we identified a strong interaction between OCT4 and karyopberin-alpha 2 （KPNA-2）. KPNA2 is involved in active nuclear import of proteins. This finding was confirmed by glutathione S-transferase pull-down and co-immunoprecipitation assays. The interaction between OCT4 and KPNA-2 was further mapped to multiple regions of the two proteins. In addition, we studied nuclear localization signal （NLS） of mouse Oct4 and demonstrated that it is essential for Oct4 nuclear localization. Thus, our data suggest that Oct4 nuclear localization may be mediated by its interaction with KPNA-2.
文摘目的探讨沉默KPNA2基因的表达对舌鳞癌细胞CAL-27迁移侵袭能力的影响。方法实验分为正常组、对照组、转染组、阴性对照组。转染组通过脂质体Lipofectamine誖2000将KPNA2-si RNA转染到舌鳞癌细胞CAL-27中,Western blot检测转染成功后,通过细胞划痕实验、侵袭实验检测KPNA2基因沉默后舌cal-27细胞系迁移侵袭能力的变化。结果 Western blot检测结果显示:转染KPNA2 si RNA 48h后,与正常对照组相比,KPNA2蛋白表达显著下降(P<0.05),细胞迁移及侵袭能力明显降低(P<0.05)。结论 KPNA2-si RNA能够下调KPNA2在舌鳞癌细胞CAL-27中的蛋白表达,沉默KPNA2的表达将降低舌鳞癌细胞的迁移侵袭能力。KPNA2作为有望成为口腔鳞癌治疗的一个新的靶向因子。
