Figures & data
Table 1. Primer sequences used in this study
Figure 1. Circ_0000423 expression is significantly up-regulated in GC tissues and cell lines
(a) Venn diagram was used to show the differentially expressed circRNAs in GSE78092, GSE89143 and GSE141977. (b) The schematic diagram of circ_0000423, which was generated from PPP1R12A transcript. (c) Circ_0000423 expression in 43 pairs of GC tissues and para-tumorous tissues was detected by qRT-PCR. (d) Circ_0000423 expression in GES-1 cells and 4 GC cell lines (AGS, MGC-803, HGC-27 and MKN-45) was detected by qRT-PCR. (e) The effect of RNase R on circ_0000423 was evaluated by qRT-PCR, to show the stability of circ_0000423. **P < 0.01 and ***P < 0.001.
Figure 2. Circ_0000423 knockdown suppresses the proliferation, migration and invasion of MGC-803 and AGS cells
(a) AGS and MGC-803 cells were transfected with si-circ_0000423#1, si-circ_0000423#2 or si-NC, and qRT-PCR was conducted for detecting circ_0000423 expression. (b) The impacts that knocking down circ_0000423 had on the growth of GC cells were detected by CCK-8 assay. (c) The impacts of knocking down circ_0000423 had on the migration of GC cells were detected by Transwell assay. (d) The impacts of knocking down circ_0000423 had on the invasion of GC cells were detected by Transwell assay. *P < 0.05, **P < 0.01 and ***P < 0.001.
Figure 3. Circ_0000423 directly targets miR-582-3p
(a) Venn diagram of the downstream miRNA targets predicted by CircInteractome (https://circinteractome.nia.nih.gov/) and StarBase (http://starbase.sysu.edu.cn/). (b) The binding site of circ_0000423 and miR-582-3p was predicted by StarBase. (c) The efficiency of miR-582-3p overexpression in MGC-803 and AGS cells was detected by qRT-PCR. (d) The luciferase activity of the cells co-transfected with miR-582-3p mimics and circ_0000423-MUT reporter or circ_0000423-WT reporter was detected. (e) MiR-582-3p expression in GC tissues and para-cancerous tissues was detected by qRT-PCR. (f) Pearson correlation analysis was used to evaluate the correlation between miR-582-3p expression and circ_0000423 expression in the 43 cases of GC tissues. (g) The expression of miR-582-3p in AGS cells and MGC-803 cells after knocking down circ_0000423 was detected by qRT-PCR. ***P < 0.001.
Figure 4. MiR-582-3p inhibition weakens the inhibitory effects of silencing circ_0000423 expression on the malignant biological behaviors of GC cells
(a) AGS and MGC-803 cells were transfected with si-NC, si-circ_0000423 or si-circ_0000423 + miR-582-3p inhibitor, and qRT-PCR was conducted for detecting miR-582-3p expression. (b) The growth of MGC-803 cells and AGS cells was detected by CCK-8 assay. (c) The migration of MGC-803 cells and AGS cells was detected by Transwell assay. (d) The invasion of MGC-803 cells and AGS cells was detected performing Transwell assay.*P < 0.05, **P < 0.01 and ***P < 0.001.
Figure 5. MiR-582-3p targets DIXDC1
(a) Venn diagram of the downstream targets of miR-582-3p predicted by StarBase, mirDIP, miRTarbase and TargetScan. (b) The expressions of DIXDC1, RREB1 and INO80D mRNA in GC tissues and matched adjacent tissues were detected through qRT-PCR. (c) The binding sites between DIXDC1 mRNA 3ʹUTR and miR-582-3p were predicted by StarBase. (d) The luciferase activity of cells co-transfected with miR-582-3p mimics and DIXDC1-WT reporter or DIXDC1-MUT1/2/1&2 reporter was measured by dual-luciferase reporter gene assay. (e) Western blot was applied to detect the effects of circ_0000423 and miR-582-3p on DIXDC1 expression.NS: differences were not statistically significant; *P < 0.05, **P < 0.01 and ***P < 0.001.
Supplemental material
Supplemental Material
Download MS Word (1 MB)Data availability statement
The data used to support the findings of this study are available from the corresponding author upon request.