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. Author manuscript; available in PMC: 2021 Feb 28.
Published in final edited form as: Nat Genet. 2020 Aug 31;52(10):1011–1017. doi: 10.1038/s41588-020-0681-7

Figure 2. LSD1 inhibition broadly impairs AR recruitment and suppresses AR transcriptional activity.

Figure 2.

a, b, c, AR ChIP-seq analyses were performed in LNCaP cells treated with vehicle, DHT (10 nM for 4 h), or DHT (4 h) with pretreated GSK2879552 (50 μM, 0.5 or 48 h). (a) overlap of AR peaks in treated cells, (b) heatmap view of AR and FOXA1 binding intensity at AR binding sites, and (c) the mean of AR binding intensity were shown (Veh vs. GSK-4h: P = 8.8 × 10−33; Veh vs. GSK-48h: P = 9.9 × 10−267). d, e, ChIP-qPCR for (d) AR or (e) p300 at AR-mediated enhancers in LNCaP cells treated with/out DHT (10 nM, 4 h) and pretreated with GSK2879552 (48 h). f, RT-PCR for KLK3 in LNCaP cells treated with 1–10 nM DHT (24 h) and pretreated with GSK2879552 (0–50 μM, 24 h). g, h, i, j, LNCaP cells were maintained in the medium containing vehicle or 1 μM GSK2879552 for ~2 weeks. The following experiments were performed: (g) qRT-PCR for KLK3; (h) immunoblotting for KLK3; (i) ChIP-qPCR for FOXA1; (j) ChIP-qPCR for AR (with 10 nM DHT). k, RNA-seq analyses were done in these long-term treated cells (10 nM DHT, 24 h) in comparison with parental LNCaP cells treated with/out DHT (24 h) and pretreated with 50 μM GSK2879552 (24 h). Androgen-upregulated genes were identified from parental LNCaP cells using 2-fold cut-off (DHT/Vehicle). The heatmap for the change in expression in response to GSK2879552 treatment was shown. l, Cell density was examined under the indicated conditions (mean±SD). m, LNCaP cells stably expressing doxycycline-inducible AR-V7 (V5 tagged) (LNCaP-tetARV7) were subjected to immunoblotting. n, ChIP-qPCR for V5 binding in LNCaP-tetARV7 cells treated with doxycycline versus doxycycline plus GSK2879552 (10 μM, 24 h). o, qRT-PCR for AR-V7-regulated genes in these cells. p, ChIP-qPCR for AR-V7 binding in CWR22-RV1 cells treated with GSK2879552 (2.5 μM, 24 h). q, ChIP-qPCR for AR-V7 binding in the LSD1-KO line versus the control line.