VIRMA-Dependent N6-Methyladenosine Modifications Regulate the Expression of Long Non-Coding RNAs CCAT1 and CCAT2 in Prostate Cancer

Abstract

RNA methylation at position N6 in adenosine (m⁶A) and its associated methyltransferase complex (MTC) are involved in tumorigenesis. We aimed to explore m⁶A biological function for long non-coding RNAs (lncRNAs) in prostate cancer (PCa) and its clinical significance. m⁶A and MTC levels in PCa cells were characterized by ELISA and western blot. Putative m⁶A-regulated lncRNAs were identified and validated by lncRNA profiler qPCR array and bioinformatics analysis, followed by m⁶A/RNA co-immunoprecipitation. Impact of m⁶A depletion on RNA stability was assessed by Actinomycin D assay. The association of m⁶A-levels with PCa prognosis was examined in clinical samples. Higher m⁶A-levels and VIRMA overexpression were detected in metastatic castration-resistant PCa (mCRPC) cells (<i>p</i> < 0.05). VIRMA knockdown in PC-3 cells significantly decreased m⁶A-levels (<i>p</i> = 0.0317), attenuated malignant phenotype and suppressed the expression of oncogenic lncRNAs CCAT1 and CCAT2 (<i>p</i> < 0.00001). VIRMA depletion and m⁶A reduction decreased the stability and abundance of CCAT1/2 transcripts. Higher expression of VIRMA, CCAT1, and CCAT2 as a group variable was an independent predictor of poor prognosis (HR = 9.083, CI95% 1.911-43.183, <i>p</i> = 0.006). VIRMA is a critical factor sustaining m⁶A-levels in PCa cells. VIRMA downregulation attenuates the aggressive phenotype of PCa by overall reduction of m⁶A-levels decreasing stability and abundance of oncogenic lncRNAs.