The miR-96 and RAR? signaling axis governs androgen signaling and prostate cancer progression.
Singh PK, Rosario S, Campbell MJ, Russell JR, Llimos G, Sucheston-Campbell LE, Smiraglia DJ, Rizvi A, Long MD, van den Berg PR, Kirk J
Oncogene in press 08/17/2018
Expression levels of retinoic acid receptor gamma (NR1B3/RARG, encodes RAR?) are commonly reduced in prostate cancer (PCa). Therefore, we sought to establish the cellular and gene regulatory consequences of reduced RAR? expression, and determine RAR? regulatory mechanisms. RARG shRNA approaches in non-malignant (RWPE-1 and HPr1-AR) and malignant (LNCaP) prostate models revealed that reducing RAR? levels, rather than adding exogenous retinoid ligand, had the greatest impact on prostate cell viability and gene expression. ChIP-Seq defined the RAR? cistrome, which was significantly enriched at active enhancers associated with AR binding sites. Reflecting a significant genomic role for RAR? to regulate androgen signaling, RAR? knockdown in HPr1-AR cells significantly regulated the magnitude of the AR transcriptome. RAR? downregulation was explained by increased miR-96 in PCa cell and mouse models, and TCGA PCa cohorts. Biochemical approaches confirmed that miR-96 directly regulated RAR? expression and function. Capture of the miR-96 targetome by biotin-miR-96 identified that RAR? and a number of RAR? interacting co-factors including TACC1 were all targeted by miR-96, and expression of these genes were prominently altered, positively and negatively, in the TCGA-PRAD cohort. Differential gene expression analyses between tumors in the TCGA-PRAD cohort with lower quartile expression levels of RARG and TACC1 and upper quartile miR-96, compared to the reverse, identified a gene network including several RAR? target genes (e.g., SOX15) that significantly associated with worse disease-free survival (hazard ratio 2.23, 95% CI 1.58 to 2.88, p?=?0.015). In summary, miR-96 targets a RAR? network to govern AR signaling, PCa progression and disease outcome.