Figure 6.

The combined effect of androgen receptor (AR) and CCAAT enhancer binding protein-α (C/EBPα) on global gene regulation. (a, c) LNCaP cells were transfected by nucleofection with either C/EBPα or vector. After 48 h, the cells were treated with either R1881(1 nM) or vehicle for 6 h. The cells were then harvested either for western blot analysis using antibody to AR or C/EBPα (a) or to obtain total RNA (c). In panel c, the mRNA profile was determined using replicate samples by Affymetrix microarray analysis and genes that were induced ≥twofold by R1881 in 6 h were selected; the data are plotted to show the effect of ectopic C/EBPα on the fold activation by R1881 of each gene; a value of 1 on the y axis indicates that R1881 stimulation was unaffected by C/EBPα. (b, d) LNCaP cells were transfected by nucleofection with C/EBPα or vector. After 24h, the cells were infected with AR short hairpin RNA (shRNA) lentivirus or non-target control lentivirus; the cells were then grown in fresh hormone-free media. The cells were then treated for 48h with either vehicle or R1881 (1 nM) as shown in (b). The cells were harvested at 72 h after nucleofection for either western blot analysis (b) or to obtain total RNA (d). In panel d, the mRNA profile was determined using replicate samples by Affymetrix microarray analysis; genes whose expression was unaffected by R1881/AR were selected; the data are plotted to show the increase in expression in the presence of R1881+AR+C/EBPα compared with C/EBPα alone.