Fig. 4

CircRAD23B acted as a sponge to regulate YBX1 expression. (A) Volcano map displaying upregulated and downregulated genes after knock down circRAD23B. (B) Heat plot showing the differentially expressed genes between sh-NC and sh-circRAD23B groups. (C, D) qRT-PCR and western blotting assay were performed to detect expression of YBX1. (E) Venn diagram showing the overlap of downstream miRNAs of circRAD23B predicted by the CircInteractome database and miRNAs predicted by starBase to regulate YBX1. (F) The binding sequence of miR-1287-5p to YBX1 mRNA and circRAD23B. (G) Luciferase intensity in HO8910 and A2780 cells co-transfected with luciferase reporter containing with wild-type or mutated circRAD23B-miR-1287-5p binding sequences and the mimics of miR-1287-5p or control. (H) The expression levels of circRAD23B and YBX1 mRNA were tested by qRT-PCR after pull-down with biotin-labeled wild-type or mutant miR-1287-5p in HO8910 and A2780 cells. (I) Luciferase intensity in HO8910 and A2780 cells co-transfected with luciferase reporter containing with wild-type or mutated YBX1 3’UTR-miR-1287-5p binding sequences and the mimics of miR-1287-5p or control. (J) Immunofluorescence assay for elaborating the relationship between circRAD23B and miR - 1287-5p, scale = 25 μm. Graph represents mean ± SD; **p < 0.01, and ***p < 0.001