A. TALEN engineering strategy. tAR-2 TALENs were designed to cut AR (red arrows)
and delete the 22Rv1 duplication. B. An mCherry/split GFP reporter harboring a
tAR-2 cut site (CS) was used to identify, sort, and plate transfected cells with
tAR-2 activity C. Cells were cultured in steroid-deplete medium and treated with
1 nM dihydrotestosterone (DHT), 10 μM enzalutamide (Enza), or vehicle
control as indicated. Blots were probed with antibodies specific for AR (N20),
AR-V7, or ERK2 (loading control). D. Growth of cells transfected with indicated
siRNAs was assessed by crystal violet staining. Bars represent mean fold growth
on Day 6 relative to Day 0 (n = 12, error bars = 95%
confidence intervals). **** p≤0.0001,
** p ≤ 0.01, ns p>0.05, Mann Whitney U tests. Bottom
panels illustrate western blots of lysates from transfected cells, probed with
antibodies as in C.