Fig. 4

CircSMYD4 as a miRNA sponge could target miR-584-5p. a Cytoplasmic and nuclear RNA was isolated from SNU-387 or SK-Hep1 cells. The relative expression level of circSMYD4 in the cytoplasm or nucleus was detected by RT-qPCR. GAPDH was used as the cytoplasmic control and U6 was used as the nuclear control. b CircSMYD4 in the SNU-387 or SK-Hep1 lysate was extracted and enriched with a 3′ UTR biotinylated circSMYD4 specific probe, and then detected by RT-qPCR. c The relative levels of five candidate miRNAs in SNU-387 or SK-Hep1 lysates were detected by RT-qPCR. d CircInteractome and e dual luciferase reporter gene assay were employed to predict and validate the binding site for circSMYD4 on miR-584-5p, respectively. f, g The effect of silenced or overexpressed circSMYD4 on miR-584-5p expression was detected by RT-qPCR. h The expression of miR-584-5p in adjacent tissues (ANT) and liver cancer tissues was analyzed by RT-qPCR. i The correlation between miR-584-5p and circSMYD4 expression was analyzed using Spearman correlation coefficients. The experiment was repeated for 3 times. ^***p < 0.001 vs. nuclear; ^###p < 0.001 vs. Oligo probe; ^ΔΔΔp < 0.001 vs. MC; ^{^^^}p < 0.001 vs. NC; ^†††p < 0.001 vs. ANT. RT-qPCR: reverse transcription real time quantitative polymerase chain reaction