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. 2014 Jul 26;16(6):489–500. doi: 10.1016/j.neo.2014.05.008

Figure 3.

Figure 3

sTβRIII inhibits BMP signaling in mammary epithelial cells. (A and B) Western blot analysis of pSmad1/5/8 signaling in HMEC and MCF10A cells serum starved overnight and pretreated with sTβRIII (60 ng/ml) or conditioned (CM) from MDA-MB-231- TβRIII cells prior to BMP2 (B2) and 4 (B4) treatment. (C) Western blot analysis of pSmad1/5/8 signaling in MDA-MB-231-Neo and TβRIII stable cells serum starved and pre-treated with recombinant sTβRIII (60 ng/ml) overnight and subsequently treated with BMP4 at the indicated dosages. (D) Western blot analysis of pSmad1/5/8 signaling in MDA-MB-231-Neo and TβRIII stable cells serum starved and treated with CM from MDA-MB-231-TβRIII cells and subsequently treated with BMP2 and 4 at the indicated dosages. (E) Western blot analysis of pSmad1/5/8 signaling in HMECs serum starved overnight and treated with pre-treated with recombinant sTβRIII at the indicated dosages. Cells were treated with 10nM BMP2 for 10 minutes. (F) Western blot analysis of pSmad1/5/8 signaling in MDA-MB-231 cells serum starved overnight and treated with pre-treated with recombinant sTβRIII at the indicated dosages. Cells were treated with 10nM BMP2 for 10 minutes. (G) Western blot analysis of pSmad1/5/8 in HMECs adenovirally infected with GFP or TβRIII-GFP serum starved overnight and pretreated with 25 μM TAPI-2 prior to treatment with 10 nM BMP2 for 10 minutes. (H) Western blot analysis of pSmad1/5/8 signaling in MDA-MB-231-Neo and TβRIII cells serum starved and pretreated overnight with 25 μM TAPI-2 prior to treatment with 10nM BMP2 for 10 minutes. Total Smad1 and β-actin levels are shown as loading controls for westerns. All experiments were independently performed at least 3 times and representative data are shown.