Supplementary Figure 1: Landscape of AR-regulated transcriptome in prostate cancer.

(a) A schematic illustration of the procedure used to discover AR-regulated genes (ARGs) in LNCaP and VCaP prostate cancer cell lines. (b) Venn diagram indicating the overlap between AR-regulated genes in LNCaP and VCaP cells. (c) qPCR analysis of ARLNC1 expression and AR signaling gene (KLK3, TMPRSS2) expression in LNCaP cells (top panel) and VCaP cells (bottom panel), following DHT treatment of 6 hours or 24 hours. Mean ± s.e.m. are shown, n = 3. Significance was determined by two-tailed Student’s t-test. (d) Bar plot depicting the distribution of gene biotypes (protein, lncRNA, and other) of all overlapped ARGs identified in both LNCaP and VCaP cells. (e) Aggregate ChIP-Seq enrichment profile depicting AR ChIP-Seq signaling density on ARG promoters in LNCaP and VCaP cells. Representative of two biological replicates are shown. (f) Aggregate ChIP-Seq enrichment profile illustrating AR ChIP-Seq signaling density on ARG promoters in LNCaP cells following DHT stimulation, AR antagonist (enzalutamide) treatment, or BRD4 inhibitor (JQ1) treatment. Representative of two biologically replicated assays are shown. (g) Transcriptional response to DHT and enzalutamide treatment in VCaP cells, plotting AR regulated protein-coding genes (top panel), or AR regulated lncRNAs (bottom panel). (h) Motif discovery analysis of the top 250 AR ChIP-Seq peaks on AR promoters identifies a binding motif similar to the canonical AR response element. P value was calculated in MEME suite. n = 2 independent AR ChIP-Seq assays. (i) Aggregated ChIP-Seq enrichment profiles depicting ChIP-Seq signal density on direct ARG promoters for H3K27ac, H3K4me1, H3K4me3, H3K36me3, Pol II, and BRD4 in LNCaP cells. (j) Pie chart showing prostate cell line or tissue distribution of direct ARGs with AR binding at transcription start sites (TSS) in ChIP-Seq. (k,l) Cumulative distribution plots of distances between transcription start sites (TSS) of genes to nearest AR peak. (k) AR binding near ARGs in benign prostate, prostate cancer tissues (PCa), and prostate cell lines. Approximately 50% (75%) of ARGs have an AR peak within 1 kb (10 kb) from the TSS. (l) Comparison of distances between AR binding sites for ARGs and genes not regulated by AR. Globally, ARGs are positioned closer to AR binding sites.