Fig. 5

YY1 activates circFIRRE transcription. A Wayne diagram showed the overlap between transcription factors of circFIRRE predicted by three different algorithms. B Relative mRNA expression of YY1 in 35 paired OS and adjacent normal samples were examined by RT-qPCR. C Pearson correlation analysis indicated that YY1 was positively correlated with circFIRRE in 35 paired samples. D The relative expression of YY1 was detected in chosen OS cell lines (MG63, U2OS) and control osteoblasts (n=3 in each group). E Schematic illustration of the binding motif of YY1 and the two possible YY1 binding sites in circFIRRE promoter. F Dual luciferase assay was performed to confirm the binding between YY1 and circFIRRE promoter, as well as two specific binding sites in HEK-293 T cells (n=3 in each group). G Three YY1-targeted siRNAs were transfected into MG63 and U2OS cells, and relative mRNA of YY1 were estimated by RT-qPCR (n=3 in each group). H-I The relative circFIRRE and linear FIRRE expression were detected in MG63 and U2OS cells after YY1 silencing by RT-qPCR (n=3 in each group). J-L The relative YY1, circFIRRE and linear FIRRE expression were detected in MG63 and U2OS cells after YY1 overexpression by RT-qPCR (n=3 in each group). Values are presented as mean ± SD; the bar charts, error bars and dots represent the quantitative analysis of 3 independent experiments; (B, F, 2-tailed Student t test; D, one-way ANOVA; G-L, two-way ANOVA); *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001; ns = not significant