Figure 5.

Hypoxia-induced POU5F1 promotes miR-1246 transcription

(A) Levels of pri-miR-1246 in normoxia- and hypoxia-conditioned glioma cells. (B) TransmiR (http://www.cuilab.cn/transmir) prediction of miR-1246 transcription factors. (C) Levels of TP73, TP63, AR, POU5F1, and FOXA1 were measured in normoxia- and hypoxia-conditioned U87MG cells. (D) The expression of miR-1246 was measured in POU5F1 and AR knockdown hypoxia-stimulated glioma cells. (E) The expression of miR-1246 was examined in control and POU5F1 knockdown hypoxia-conditioned GDEs. (F) The expression of pri-miR-1246 was examined in control and POU5F1 knockdown hypoxia-conditioned glioma cells. (G) Computational prediction of POU5F1 binding sites (S1–S9) in the miR-1246 regulatory region and confirmation of POU5F1 binding to candidate miR-1246 sites via ChIP-qPCR analysis in hypoxia-conditioned U87MG cells. (H) Dual-luciferase reporter assay demonstrating the luciferase activity of the wild-type, S1 deletion, S7 deletion, and S1+S7 deletion miR-1246 promoter in U87MG cells transfected with si-NC or si-POU5F1 (n = 4 for each group). (I) POU5F1 protein levels were measured in normoxia- and hypoxia-conditioned U87MG and P3 cells. HIF1A knockdown inhibited the hypoxia-induced POU5F1 expression. The data are presented as the mean ± SD; ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001.