TY - JOUR T1 - The miR-96 and RARγ signaling axis governs androgen signaling and prostate cancer progression JF - bioRxiv DO - 10.1101/198465 SP - 198465 AU - Mark D Long AU - Prashant K Singh AU - James R Russell AU - Gerard Llimos AU - Spencer Rosario AU - Abbas Rizvi AU - Patrick R. van den Berg AU - Jason Kirk AU - Lara E Sucheston-Campbell AU - Dominic J Smiraglia AU - Moray J Campbell Y1 - 2018/01/01 UR - http://biorxiv.org/content/early/2018/04/26/198465.abstract N2 - Expression levels of retinoic acid receptor gamma (NR1B3/RARG, encodes RARγ), are commonly reduced in prostate cancer (PCa). Therefore we sought to establish the cellular and gene regulatory consequences of reduced RARγ expression, and determine RARγ regulatory mechanisms. RARG shRNA approaches in non-malignant (RWPE-1 and HPr1-AR) and malignant (LNCaP) prostate models revealed that reducing RARγ levels, rather than adding exogenous retinoid ligand, had the greatest impact on prostate cell viability and gene expression. ChIP-Seq defined the RARγ cistrome which was significantly enriched at active enhancers associated with AR binding sites. Reflecting a significant genomic role for RARγ to regulate androgen signaling, RARγ knockdown in HPr1-AR cells significantly regulated the magnitude of the AR transcriptome. RARγ down-regulation was explained by increased miR-96 in PCa cell and mouse models, and TCGA PCa cohorts. Biochemical approaches confirmed that miR-96 directly regulated RARγ expression and function Capture of the miR-96 targetome by biotin-miR96 identified that RARγ and a number of RARγ 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 RARγ 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 RARγ network to govern AR signaling, PCa progression and disease outcome.Conflict of interest: The authors certify that they have NO affiliations with or involvement in any organization or entity with any financial interest (such as honoraria; educational grants; participation in speakers’ bureaus; membership, employment, consultancies, stock ownership, or other equity interest; and expert testimony or patent-licensing arrangements), or non-financial interest (such as personal or professional relationships, affiliations, knowledge or beliefs) in the subject matter or materials discussed in this manuscript.FUNDING LESC acknowledges support, in part, of Roswell Park Comprehensive Cancer Center-University of Pittsburg Cancer Institute Ovarian Cancer Specialized Program of Research Excellence National Institutes of Health [P50CA159981-01A1].MDL acknowledges support of Molecular Pharmacology and Experimental Therapeutics NRSA T32 program [T32CA009072] held at Roswell Park Comprehensive Cancer Center.MJC and DJS acknowledges support in part from the Prostate program of the Department of Defense Congressionally Directed Medical Research Programs [W81XWH-14-1-0608, W81XWH-11-2-0033] and the National Cancer Institute (NCI) grant P30CA016056 involving the use of Roswell Park Comprehensive Cancer Center’s Genomic Shared Resource.MJC, GL, AR, HW and PvdB acknowledges support from the European Union-United States Atlantis Program [P116J090011].MJC and LESC acknowledge support from the National Cancer Institute (NCI) grant P30CA016056 involving the use of OSUCCC The James, CCSG P30CA016058ARAndrogen receptorCEBPBCAAT/enhancer binding protein (C/EBP), beta,ChIPChromatin ImmunoprecipitationCNVCopy number variationCTCFCCCTC-binding factor (zinc finger protein)DEGsDifferentially expressed genesENCODEEncyclopedia of DNA elementsERαEstrogen receptor alphaETSE26 transformation-specificFAIREFormaldehyde-Assisted Isolation of Regulatory ElementsFDRFalse discovery rateGATAGATA-BINDING PROTEIN 1GEOGene Expression OmnibusGSEAGene set enrichment analysesHHistoneHDACHistone deacetylaseKLysineKDM1A/LSD1Lysine (K)-Specific Demethylase 1AmiRNAmicroRNANCOR1Nuclear receptor corepressor 1NCOR2/SMRTNuclear receptor corepressor 1/Silencing mediator for retinoid or thyroid-hormone receptorsNESNormalized enrichment scoreNRNuclear hormone receptorONEC2Onecut2PCaProstate cancerPPARPeroxisome proliferator-activated receptor gammaPPARGC1APeroxisome Proliferator-Activated Receptor Gamma, Coactivator 1 AlphaPRADProstate cancer cohort in TCGARARγRetinoic acid receptor gammaRPKMreads per kilobase per million mapped readsSeqSequencingshRNAshort hairpin RNASOX15SRY (Sex Determining Region Y)-Box 15STAT5ASignal Transducer And Activator Of Transcription 5ATACC1Transforming, Acidic Coiled-Coil Containing Protein 1TCGAThe cancer genome atlasTNFTumor necrosis factorTRIM28Tripartite motif containing 28 ER -