Fig. 5. The depletion of SUPT5H gene expression induces cell cycle arrest in MDA-MB-231 cells. (A) Cells transfected with siRNA-SUPT5H only (knockdown), siRNA-SUPT5H plus the siRNA resistant SUPT5H ORF (Δ2), non-silencing siRNA-scr or siRNA-scr plus empty vector (EV) were stained with propidium iodide after 72 hours of transfection and subjected to FACS analysis to determine cell-cycle distribution. Depletion of SUPT5H by siRNA-SUPT5H induced the S-phase cell cycle arrest, and the effect of siRNA-SUPT5H was rescued by engineered SUPT5H gene construct. (B) Quantification of p21 gene expression in SUPT5H silenced MDA-MB-231 cells using real-time qPCR. (A, B) data represented as means ± SD (n=3). *p<0.05, **p<0.01, not significant, ns (C) Western blot analysis of cell-cycle regulatory proteins in MDA-MB-231 cells transfected with siRNA-SUPT5H or siRNA-scr (representative images are shown from two independent experiments).