FOXO3A Antibody - #AF7624

Figure 3 Human telomerase reverse transcriptase (hTERT) promotes integrin β1 (ITGB1) expression by modulating the binding of both forkhead box M1 (FOXM1) and forkhead box O3 (FOXO3a) to the ITGB1 gene. (A) Chromatin immunoprecipitation (ChIP)-qPCR analysis of the binding sites of hTERT in the regulatory region of the ITGB1 gene. ChIP assays were performed using chromatin isolated from SGC-7901 and HGC-27 cells, and final DNA extractions were qPCR amplified by 49 pairs of primers covering the regulatory region of ITGB1 (Chr10: 33 253 528∼Chr10: 33 226 885). (B) Influence of hTERT on the luciferase activity of the pGL3-promoter constructs. The recombinant constructs (named pGL3-p1-6 (Left) and pGL3-I1-6 (Right)) were separately cotransfected with pIRES2-hTERT plasmid (or control plasmid) into U2OS cells, and cells were harvested 24 h later for dual luciferase activity assays. (C) ChIP analysis of the binding of transcription factors to the (−1166∼−1074) or (+9913∼+10 010) region of ITGB1 in both SGC-7901 and HGC-27 cells. (D) ITGB1 expression after FOXM1 or FOXO3a overexpression in SGC-7901 cells. Forty-eight hours after the transient transfection of pEGFP-FOXM1 or pcDNA-FOXO3a plasmids, together with the corresponding controls, SGC-7901 cells were harvested for qPCR and western blot analysis. (E) Effect of FOXM1 or FOXO3a on the activity of pGL3-P5 or pGL3-I5, respectively. SGC-7901 cells in 24-well plates were transiently transfected with the luciferase reporters, together with pEGFP-FOXM1 or pcDNA-FOXO3a plasmids. Twenty-four hours later, the luciferase activity was measured using luciferase assay substrate. (F) ChIP assays were performed using chromatin isolated from gastric cancer cells infected with lenti-hTERT or lenti-Control. Final DNA extractions were PCR amplified using primers that cover FOXM1 binding element (F1BE) (−1104∼−1109) or FOXO3a binding element (F3BE) (+9972∼+9978) within the ITGB1 gene. *p<0.05, **p<0.01.