Research Paper Volume 14, Issue 22 pp 9149—9166

Figure 6. KLF10 directly bound to the promoter of LINC00629. (A) Schematic illustration of the pGL3-based reporter constructs (P0-P3) used in luciferase assays to examine the transcriptional activity of LINC00629. (B) The promoters of LINC00629, named P0-P3, were individually transfected into UM-SCC6 cells, and the cells were treated with or without 40 μM apigenin. Luciferase activity was measured. (C, D) The promoter of P1 was transfected into UM-SCC6 and Cal-27 cells with or without KLF10 knockdown, and the cells were then treated with or without 40 μM apigenin for 36 h. Luciferase activity was measured. (E) Schematic illustration of the KLF10 wild-type binding site (BS1 WT) and the matching mutant (BS1 MUT) that were used in luciferase assays. (F, G) BS1 WT and MUT were transfected into 293T cells with or without KLF10 overexpression. KLF10 expression was determined by Western blotting (F). Luciferase activity was detected (G). (H) BS1 WT and BS1 MUT were transfected into UM-SCC6 cells treated with or without apigenin. The luciferase activity was measured. (I–K) ChIP assay showing the binding of KLF10 to the promoter of LINC00629 in UM-SCC6 cells with or without apigenin treatment or KLF10 knockdown. Isotype-matched IgG was used as the negative control. In (B–D, G, H) the results represent three independent experiments; *p<0.05, **p<0.01, ***p<0.001.