Fig. 5

CCL20 is a direct target of miR-1322 and activates PI3K-Akt pathway through circSMARCC1/miR-1322/CCL20 axis. A, B The heatmap and volcano plot of differentially expressed mRNAs in DU145 cells transfected with vector or circSMARCCC1. Each sample was mixed with three replicates. C Venn diagram illustrating the overlapping of miR-1322 targeting mRNAs and up-regulated mRNAs in DU145 cells. D qRT-PCR was performed to detect the mRNA expression of 9 candidate molecules (MS4A1, CBFB, ARPP21, PHF6, FTO, CCL20, MED7, CD40 and SLC25A15). E, F The expression of CCL20 in PCa cells transfected with miR-1322 mimics or miR-1322 inhibitors was detected by qRT-PCR and Western blotting. G Schematic illustration of CCL20 wide type (Wt) and mutant (Mut) luciferase reporter vectors were shown. H The relative luciferase activities measured in 293 T cells co-transfected with CCL20-Wt or CCL20-Mut and miR-1322 mimics or miR-nc by luciferase reporter assay. I IF analysis detected the protein expression of CCL20 in PCa cells transfected or co-transfected with lv-circSMARCC1, sh-circSMARCC1, miR-1322 mimics or inhibitors (scale bar, 50 μm). J The relative protein levels of CCL20, EMT biomarkers and cell cycle-related molecules detected by Western blotting in miR-1322 rescue experiments. K IHC detection of CCL20 expression in PCa tissues and para-cancerous tissues (scale bar, 100 μm and 20 μm). L the Secretion of CCL20 in the supernatants of circSMARCC1-overexpressing DU145 and PC-3 cells cultured in RPMI-1640 medium or 10% FBS medium was measured by ELISA. M, N The CCL20 recombinant and CCL20 neutralizing antibodies were applied to transwell assays and CCK8 assays to explore the effects of CCL20 on PCa cell proliferation, migration and invasion. O, P KEGG and GSEA analyses of DEGs in DU145 cells. Q The relative protein levels of CCL20 and AKT pathway-related molecules measured by Western blotting in miR-1322 rescue experiments. The data are presented as the mean ± SD, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001