Supplementary Figure 1 Supplementary Figure 1. Representative micrographs of migration and invasion assays. miR-452 or si-WWP1 transfectants inhibited.

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Supplementary Figure 1 Supplementary Figure 1. Representative micrographs of migration and invasion assays. miR-452 or si-WWP1 transfectants inhibited cancer cell migration and invasion in PCa cells. PC3DU145PC3DU145 Migration assayInvasion assay mock control miR-452 mock control si-WWP1-1 si-WWP1-2 mock control si-WWP1-1 si-WWP1-2 PC3DU145 mock control miR-452 PC3DU145 Migration assayInvasion assay

Supplementary Figure 2 Supplementary Figure 2. Effects of co-transfection of miR-224 and miR-452 on migration assay Co-transfection of miR-224 and miR-452 did not show synergistic effects on cancer cell migration compared with miR-224 or miR-452 transfection in PC3 cells Cell migration (relative to mock) PC3 (%) mock control miR-452 miR miR-452 miR-224 mock control miR-452 miR miR-452

TargetScan Database (release 6.2) 3,161 putative miR-452 target genes GEO Database PCa expression data (GSE 29079) Fold change (log 2 ratio) > candidate genes Gene expression analysis miR-452 transfection into PC3, DU145 (GSE 56243) Fold change (log 2 ratio) < genes Supplementary Figure 3 Supplementary Figure 3. Strategy for selecting putative target genes of miR-452. The TargetScan program showed that 3,161 genes had putative target sites for miR-452 in their 3′ UTRs. A total of 704 genes were significantly upregulated in PCa specimens (log 2 ratio > 0.1) in GSE Finally, we performed genome-wide gene expression analysis using PC3 and DU145 cells (GSE56243). Genes downregulated (log 2 ratio < -0.2) by transfection with miR-452 were selected as putative target genes.

Supplementary Figure Expression of mRNA WWP1 (relative to mock) PC3 * (%) * mock control si-WWP1-1si-WWP DU145 (%) * * mock control si-WWP1-1si-WWP1-2 WWP1 GAPDH mock controlsi-WWP1-1si-WWP1-2 mock controlsi-WWP1-1si-WWP1-2 A B Supplementary Figure 4. Effects of silencing WWP1 mRNA and protein expression by si-WWP1 transfection in PCa cells. (A) WWP1 mRNA expression was determined at 72 h after transfection with si-WWP1. GUSB was used as an internal control. (B) WWP1 protein expression was evaluated by western blotting at 72 h after transfection with si-WWP1. GAPDH was used as a loading control. *P < The bars indicate SDs.

Supplementary Figure Expression of WWP1 Primary PCa (n = 65) Metastatic PCa (n = 25) P = Supplementary Figure 5. WWP1 expression in metastatic PCa and primary PCa. Expression levels of WWP1 were significantly higher in metastatic PCa compared with primary PCa (GEO accession number: GDS2545).