[RNA-binding protein ELAVL1 regulates SOX4 stability and promotes hormone-sensitive prostate cancer proliferation through m6A-dependent regulation]

Abstract

Objective: To investigate the expression of RNA binding protein ELAVL1 in prostate cancer (PCa), especially hormone-sensitive prostate cancer (HSPC), and its relationship with tumor proliferation. This study further aims to reveal the molecular mechanism by which ELAVL1 promotes HSPC proliferation by stabilizing SOX4 mRNA in an m6A-dependent manner.

Methods: The expression of ELAVL1 in PCa tissues and its relationship with prognosis were analyzed in the Cancer Genome Atlas (TCGA) database, and the differences in HSPC and hormone-resistant prostate cancer (HRPC) were compared. And its relationship with prognosis were analyzed in the Cancer Genome Atlas (TCGA) database, and the differences in HSPC and hormone-resistant prostate cancer (HRPC) were compared. Western blot was used to detect ELAVL1 protein expression in PCa cell lines. After ELAVL1 knockdown by siRNA, cell proliferation was evaluated using CCK-8 assays, and changes in downstream target genes were detected by RT-qPCR. Tumor xenograft experiments in nude mice were performed to further assess the impact of ELAVL1 on tumor growth. The interaction between ELAVL1 and SOX4 mRNA was verified by RIP-seq. And the mRNA and protein levels of SOX4 after knockdown of ELAVL1 were detected by RT-qPCR and Western blot, respectively. CCK-8 was used to evaluate the effect of SOX4 knockdown on cell proliferation. MeRIP-qPCR was used to detect the m6A modification level of SOX4 and the effect of knocking down METTL3. RNA pull-down experiments verified the interaction between SOX4 RNA fragments and ELAVL1 protein. RNA stability experiments evaluated the effect of ELAVL1 knockdown on SOX4 mRNA stability.

Results: The expression of ELAVL1 in PCa cells was higher than that in normal prostate epithelial cells. The prognosis of patients with high expression of ELAVL1 was significantly worse than that of patients with low expression. In the GSE32269 dataset, the expression level of ELAVL1 in HSPC was significantly higher than that in HRPC. After knocking down of ELAVL1 in LNCaP and VCaP cells, CCK-8 experiments showed that the cell proliferation ability was significantly affected after knocking down ELAVL1, and overexpressed ELAVL1 promoted the proliferation of HSPC cells. The results of in vivo studies showed that knockdown of ELAVL1 significantly inhibited the tumorigenic capacity of LNCaP cells and resulted in a marked reduction in xenograft tumor mass. The levels of SOX4 mRNA and protein in LNCaP and VCaP cells were significantly higher than those in normal prostate epithelial cells RWPE-1. RIP-qPCR confirmed the interaction between ELAVL1 protein and SOX4 mRNA. After knocking down of ELAVL1, the expression levels of SOX4 mRNA and protein were significantly decreased. After knocking down of SOX4, the proliferation ability of LNCaP and VCaP cells was significantly inhibited.

Conclusion: ELAVL1 is highly expressed in HSPC. High expression of ELAVL1 is associated with the proliferation of HSPC. SOX4 is a downstream molecule of ELAVL1 which promotes the proliferation of HSPC. ELAVL1 enhances the stability of SOX4 mRNA through an m6A-dependent mechanism.

Keywords: hormone-sensitive prostate cancer; ELAVL1; SOX4; prostate cancer; proliferation; m6A.