Figure 1. Overexpression of EZH2 enhances invasion.

(a) Ectopic expression of EZH2 induces invasion of primary prostate epithelial cells and benign immortalized breast cell lines. A reconstituted basement membrane invasion chamber assay (Boyden chamber assay) was used to assess the invasive potential of primary prostate and benign breast epithelial cell lines infected with EZH2, EZH2ΔSET or control adenovirus. EZH2 infected cells were also treated with the histone deacetylase inhibitor SAHA (500nM). Representative fields of invaded and stained cells are shown (left). Invasion was quantitated using colorimetry (absorbance at 560 nm, right). All p values were calculated between EZH2 and vector treated samples. (b) The SET domain mutant of EZH2 inhibits cancer cell invasion. DU145 cells, which express high levels of endogenous EZH2, were infected with EZH2, EZH2ΔSET, and control adenoviruses. Invasion was quantitated using colorimetry. The p value was calculated between EZH2ΔSET and vectors. (c) Cell invasion is attenuated by EZH2 knockdown. Boyden chamber invasion assay using DU145 cells treated with siRNA duplexes targeting EZH2. Inset demonstrates knockdown of EZH2 protein by RNA interference. All p values were calculated between control and EZH2 knockdown clones. (d) Stable knockdown of EZH2 decreases invasiveness of DU145 cells. DU145 cells were stably transfected with EZH2 shRNA and assessed by invasion assay. Three stable clones exhibiting knockdown of EZH2 are shown. Inset demonstrates knockdown of EZH2 protein by RNA interference.