Construction of HOXB13-manipulated LNCaP prostate cancer cells: A, HOXB13 was induced by doxycycline in S2 and S4 cells compared to no expression in the Tet-on control cells. B, Recombinant HOXB13 in S4 the LNCaP cells was quantitated by densitometry. The endogenous HOXB13 level was arbitrarily set as one fold. C, Western blot analysis showed clones of HOXB13-suppressed LNCaP cells in addition to scrambled DNA-transfected LNCaP. D-E, By the reporter transcription assay, the effect on androgen-stimulated AR activity was tested in the Tet-on and S4 cells and HOXB13-suppressed clones. Cells were transiently transfected with 100 ng of pGL-ARE4-Luc and 2 ng of ranilla with or without 10 nM R1881. When needed, doxycycline was added with R1881 to induce HOXB13. Luciferase assays were performed 48 h post-transfection. Values indicate fold induction (RLU with androgen/RLU without androgen). Each bar represents the mean ± S.D.