Generation of human ovarian cancer OVCA429 cells with Dox-inducible expression of the constitutively-active mutant ALK3 receptor (ALK3QD). (A) Phosphorylation of Smad1 (P-Smad1) and expression of ID1 mRNA are induced in serum-starved OVCA429 cells treated with 10 ng/mL BMP4 for 30 min and 90 min, respectively. Total Smad1 and actin were used as protein loading controls, and GAPDH for RNA loading control. (B) Expression of HA-tagged constitutively-active ALK3 receptor (ALK3QD) was observed by Western analysis in two independent 429T-ALK3QD stable cell clones (A44 and A54) after 24 h treatment with Dox, as compared with control cells (T7Hyg4). Actin was used as a control for protein loading. (C) Activated BMP signalling was confirmed by Northern analysis of ID1 and ID3 mRNA expression from 429T-A44 and 429T-A54 cells, and 429T control cells, treated with Dox for 24 h, or left untreated. GAPDH served as a control for RNA loading.