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. Author manuscript; available in PMC: 2016 Jan 29.
Published in final edited form as: Cell. 2015 Jan 22;160(3):367–380. doi: 10.1016/j.cell.2014.12.023

Figure 1. TOP1 Occupies AR-enhancers and Affects the Transcriptional Program of the Prostate Cancer Cell Line LNCaP.

Figure 1

(A) Recruitment of AR and TOP1 to the KLK3 and KLK2 enhancers. The highest TOP1 binding is detected at 15 min DHT treatment. Data points show mean ± s.d.; (n=3), *P<0.05, **P<0.01. (B) The UCSC genome browser screenshot of the KLK3-KLK2 locus showing the occupancy of p-S5-RNA PolII (Pol II), AR and TOP1 (all tested with and without DHT treatment). (C) GRO-seq analysis of the effect of TOP1 knockdown on nascent RNA levels shown as a heatmap for 579 (out of 644, which were upregulated by DHT treatment) with the most affected AR-enhancers at the top. (D) Heatmap showing DHT-induced TOP1 sequencing tags density increase around 644 AR-enhancer binding sites (centered on AR). (E) Boxplot: siTOP1 reduced transcription at ~ 80% of DHT-up-regulated AR-enhancers. *P 2.2e-16 (Wilcoxon test). (F) Boxplot: The response to DHT of 368 DHT-up-regulated genes was reduced after TOP1 knockdown by siRNA. (G) Knockdown of TOP1 affects the induction of both eRNA and mRNA. LNCaP cells, hormone-starved for 1 day and transfected with the indicated siRNA, were stimulated with 100 nM DHT for 1h (eRNA) or 5 h (mRNA) 48 h post transfection. Quantitative RT-PCR was performed with SYBR Green using reverse-transcribed RNA. Data represent mean ± s.d.; (n=3), **P<0.01. (H) Recruitment of ATR to the KLK3 and KLK2 enhancers following DHT stimulation of starved cells. Data represent mean ± s.d.; (n=3). **P<0.01. See also Figure S1 and Table S1.