Figure 5

HMGA2 acts as a master switch of EMT. (a) MCF-7 cells were treated with 10 ng/ml of OSM and the expression levels of c-Myc, Ras and HMGA2 were determined by western blot at the indicated time points. (b) MDA-231 cells were transfected with let-7b/d/g and the expression levels of c-Myc, Ras and HMGA2 were analyzed by western blot. (c) MCF-7 cells were transfected with let-7b/d/g. After transfection for 24 h, the cells were exposed to 10 ng/ml of OSM. The expression levels of c-Myc, Ras and HMGA2 were analyzed by western blot at the indicated time points. (d) MDA-231 cells were transfected with 50 or 100 nM HMGA2-specific siRNA. After transfection for 48 h, the expression levels of E-cadherin, fibronectin and HMGA2 were analyzed by western blot. (e) MCF-7 cells were transfected with HMGA2 siRNA. After transfection for 48 h, the cells were treated with 10 ng/ml of OSM. The expression of HMGA2 was analyzed by western blot at the indicated time points. (f, g) MCF-7 cells were transfected with HMGA2 siRNA. After transfection for 24 h, the cells were treated with 10 ng/ml of OSM for 4 days. The expression levels of fibronectin, E-cadherin and ZEB1 were determined by western blot (f) and miR-200c by real-time RT–PCR (g). (h, i) MCF-7 and MDA-231 cells were transfected with HMGA2 siRNA. The transfected MCF-7 cells were treated with 10 ng/ml of OSM for 4 days. The invasive activities were determined by Matrigel invasion assays after 10 h (MDA-231 cells) or 48 h (MCF-7 cells) of culture. The invading cells were stained (h) and quantified (i). (j) MCF-7/Stat3 sh cells were exposed to 10 ng/ml of OSM. The expression of HMGA2 and phosphorylation of Stat3 were analyzed by western blot at the indicated time points. Error bars indicate s.e. (N⩾3). GAPDH, glyceraldehyde 3-phosphate dehydrogenase.