Fig. 4

Short-term HDAC1 knockdown downregulates HDAC7. a Real-time quantitative RT-PCR analysis confirmed a significant siRNA-mediated knockdown of HDAC7 (4.7- and 8.0-fold) and HDAC1 (37.5- and 29.8-fold) mRNAs respectively in nsTC-like HMLER2 (gray bars) and CSC-like BPLER2 (black bars) cells at 72 h post transfection. HDAC1, MYC, and VDR mRNA levels decreased 1.7- to 2.3-fold with HDAC7 specific siRNAs (siH7) in BPLER2 but not HMLER2 cells. As for VDR and c-MYC, HDAC7, mRNA levels did not change significantly with HDAC1-specific siRNAs (siH1) transient transfection in both cell populations. Transcript levels were normalized to ACTB and B2M and expressed relative to values in nontargeting (NT) siRNAs control. Results of real-time quantitative RT-PCR from three independent repeats were plotted in a bar chart using logarithmic scale. Error bars represent standard deviations. b HDAC7 protein expression was reduced with transient transfection of siH1 for 72 h in HMLER2 and BPLER2 cells, but no change was observed in HDAC1 protein expression with transient transfection of siH7. Nontargeting siRNAs (NT) were used as control. No change in c-MYC and VDR expression or total H3K27ac and H3K9/14ac (H3ac) levels was found at 72 h after transient transfection with either siH1 or siH7. Immunoblot of whole cell lysates were probed with the indicated antibodies