The impact of WDR77 on the AR cistrome in prostate cancer cells

Androgen receptor (AR) drives prostate cancer (CaP) even after androgen deprivation therapy (ADT) failed. Inhibiting AR’s transcription factor function to overcome acquired resistance to ADT is an attractive therapeutic avenue. However, which AR-coregulator interaction should be targeted is not clear, AR-coregulator complex higher order structures and stoichiometry are unknown, and how manipulating AR-coregulator interactions impacts the AR cistrome is not understood. Here, we examine interactions between AR and WD repeat 77 (WDR77, non-catalytic component of the methylosome complex), which are enriched in ADT-resistant CaP and mediate CaP cell survival. We performed CUT&RUN assays defined AR and WDR77 cistromes, the WDR77-dependence of the AR cistrome and the overlap between the AR and WDR77 cistromes in CaP cells. In a first experiment, LNCaP cells were seeded in RPMI1640 medium supplemented with 9% FBS without added antibiotics. The next day, cells were transfected with siRNAs targeting WDR77 or non-targeting control siRNA. The next day, medium was changed to medium supplemented with 9% charcoal-stripped FBS. After 80hours, medium was refreshed and cells were treated with 5nM R1881 or vehicle (ethanol) for another 16h. CUT&RUN assays were performed using an antibody targeting AR. In a second experiment, LNCaP cells were seeded in RPMI1640 medium supplemented with 9% charcoal-stripped FBS. Two days later, medium was refreshed and cells were treated with 5nM R1881 or vehicle (ethanol) for another 16h. CUT&RUN assays were performed using an antibody targeting WDR77.