RRP9 promotes prostate cancer metastasis and epithelial-mesenchymal transition through activation of the AKT/GSK3β/β-Catenin signaling pathway

Abstract

Ribosomal RNA Processing 9 (RRP9) is a gene associated with ribosomal function, and studies have demonstrated that its expression is aberrantly regulated in various tumor types, correlating with tumor progression. However, the specific role and underlying mechanism of RRP9 in prostate cancer (PCa) remain largely unexplored. In this study, bioinformatics analysis revealed that RRP9 is upregulated in PCa and is significantly associated with poor prognosis and lymph node metastasis. Further experimental data demonstrated that RRP9 knockdown notably inhibited the metastasis, invasion, and epithelial-mesenchymal transition (EMT) of PCa. Conversely, overexpression of RRP9 activated the AKT signaling pathway, resulting in the phosphorylation of GSK3β at Ser9, which in turn prevented β-catenin degradation and promoted cell metastasis, invasion, and EMT. Rescue experiments demonstrated that SC79 effectively reversed the inhibitory effects of RRP9 knockdown on PCa. These findings highlight the potential role of RRP9 in promoting the malignant biological behaviors of PCa, providing new insights and potential therapeutic strategies for the treatment of the disease.

Keywords: AKT; Epithelial-mesenchymal transition; GSK3β; Metastasis; Prostate cancer; RRP9; β-catenin.

Fig. 1

Bioinformatics analysis and expression validation of RRP9. A, B Differential expression of RRP9 between tumor and normal tissues in the TCGA database. C, D RRP9 expression was upregulated in tumor tissues in the GSE70768 and GSE46602 datasets. E, F The expression of RRP9 in PCa was analyzed with respect to different Gleason scores and lymphatic metastasis status. G, H Kaplan–Meier analysis of OS and DFS based on RRP9 expression in the TCGA database. I, J Immunohistochemical staining images of RRP9 in PCa and normal tissues. K Functional enrichment analysis. L RT-qPCR analysis demonstrated differential expression of RRP9 in PCa cell lines compared to prostate epithelial cell line. M Western blot analysis confirmed the differential expression of RRP9 in prostate cancer cell lines and prostate epithelial cell line

Fig. 2

RRP9 significantly promotes migration and invasion in PCa. A, B Western blot analysis confirms the stable knockdown and overexpression of RRP9 in PCa. C Wound healing assay demonstrates reduced migration in PCa with RRP9 knockdown. D Wound healing assay shows enhanced migration in PCa with RRP9 overexpression. E Migration and invasion capabilities of PCa with RRP9 knockdown were evaluated using Transwell assays. F Migration and invasion capabilities of PCa with RRP9 overexpression were evaluated using Transwell assays

Fig. 3

Effect of RRP9 on EMT and the AKT/GSK3β/β-catenin signaling pathway. A Western blot analysis showing the impact of RRP9 knockdown on the protein levels of EMT-related markers. B Western blot analysis showing the effect of RRP9 overexpression on the protein levels of EMT-related markers. C GSEA analysis indicates that genes co-expressed with RRP9 are significantly enriched in the WNT/β-catenin signaling pathway. D Western blot analysis demonstrating the effect of RRP9 knockdown on the protein levels of p-AKT/AKT, p-GSK3β(S9)/GSK3β, and β-catenin. E Western blot analysis demonstrating the effect of RRP9 overexpression on the protein levels of p-AKT/AKT, p-GSK3β(S9)/GSK3β, and β-catenin

Fig. 4

SC79 effectively reverses the impact of RRP9 knockdown on PCa. A Results of wound healing assays for the indicated subgroups. B Results of Transwell assays showing the migration and invasion capabilities of PCa cells in the indicated subgroups

Fig. 5

The effects of RRP9 on EMT through the AKT/GSK3β/β-catenin pathway. A SC79 reverses the effects of RRP9 knockdown on the protein levels of p-AKT/AKT, p-GSK3β(S9)/GSK3β, and β-catenin. B SC79 restores the expression levels of EMT-related proteins affected by RRP9 knockdown

Fig. 6

Schematic representation of RRP9-induced promotion of PCa metastasis and EMT via the AKT/GSK3β/β-catenin signaling axis. Upregulation of RRP9 activates AKT through phosphorylation, which subsequently phosphorylates the Ser9 residue of GSK3β. This modification inhibits GSK3β activity, leading to the stabilization and accumulation of β-catenin. The resultant increase in β-catenin levels promotes PCa metastasis and facilitates the EMT process