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. 2019 Apr 13;16:543–553. doi: 10.1016/j.omtn.2019.04.004

Figure 4.

Figure 4

SBF2-AS1 Binds with miR-338-3p and miR-362-3p

SBF2-AS1 is located in both cytoplasm and nucleus in LUAD cells (A). PAR-CLIP-seq data suggested that SBF2-AS1 could bind with miR-338-3p and miR-362-3p (B). RIP assay using antibody specifically targeting Ago2 showed that SBF2-AS1 could bind with Ago2 protein in A549 cells (C). Biotin-coupled miRNA pull-down assay showed that SBF2-AS1 was significantly enriched by miR-338-3p and miR-362-3p (D). Compared with control RNA, overexpression of miR-338-3p (E) or miR-362-3p (F) did not alter SBF2-AS1 expression. E2F1 is the target of miR-338-3p and miR-362-3p. Venn plot showed that 8 cell-cycle-related genes were overlapped with predicted targets of miR-338-3p and miR-362-3p (G). Blue circle: cell-cycle-related genes that were selected from downregulated genes upon SBF2-AS1 knockdown; yellow circle: predicted targets of miR-338-3p; green circle: predicted targets of miR-362-3p. E2F1 was mostly downregulated upon SBF2-AS1 knockdown (H). Biotin-labeled miRNA pull-down assay showed that miR-338-3p and miR-362-3p could bind with E2F1 (I). In a dual-luciferase reporter assay, luciferase activity was inhibited by miR-338-3p (J) and miR-362-3p (K), but the inhibition was abolished when the binding sites of miR-338-3p (J) and miR-362-3p (K) were mutated, respectively. E2F1 mRNA expression level decreased upon ectopic expression of miR-338-3p (L) or miR-362-3p (M). *p < 0.05; **p < 0.01; N.S, no statistical significance. Error bars stand for mean and SE.