Nanog influences the proliferative ability of HepG2 cells through the modulation of the cell cycle related proteins / 实用肿瘤学杂志

ABSTRACT

Objective To study the effect of pluripotency factor Nanog on the expression of the cell cy-cle related proteins,and then to explore its effect on the proliferative ability of HepG 2 cells.Methods TALENs gene editing tool was employed to induce mutation and downregulation expression of Nanog .T7 endonuclease 1 and genomic sequencing was used to analyze the mutation efficiency of Nanog .RT-PCR and western blot were used to determine the expression of mRNA and protein of Nanog ,respectively .Real-time cell based assay system was used to measure the proliferative ability of wild -type HepG2 cells and monoclonal HepG 2 cells with Nanog mutation.Results TALENs successfully induced mutation of Nanog gene .The targeting efficiency of mixed cells was analyzed by T7 endonuclease 1 approached 40%after two transfection with plasmid of Nanog -TALENs.Ad-ditionally,the Nanog mRNA expression level of monoclonal HepG 2 with Nanog mutation was downregulated by 3.4 times compared to the wild type HepG 2 cells,and the Nanog protein expression level was downregulated by 3.6 times.The cell cycle related proteins CyclinD1/D3,CyclinE1 and CDK2 expression were downregulated in monoclonal HepG2 with Nanog mutation in comparison to the wild type HepG 2 cells.Conclusion Nanog plays a role in influencing the proliferative ability of HepG 2 cells through modulating the expression of the cell cycle re-lated proteins CyclinD1/D3,CyclinE1 and CDK2.The downregulation expression of Nanog can inhibit the prolif-erative capacity of HepG 2 cells via the regulation of the cell cycle related proteins .