ml347 alk1 2 inhibitor tocris  (Tocris)

Journal: Neoplasia (New York, N.Y.)

Article Title: Epigenetic Regulation of GDF2 Suppresses Anoikis in Ovarian and Breast Epithelia 1

doi: 10.1016/j.neo.2015.11.003

Figure Lengend Snippet: ALK3 and ALK6 are required for GDF2-induced SMAD1/5 phosphorylation. (A-B) Western blotting for pSMAD1/5 activation in PA1 and MCF10A cells in the presence and absence of dorsomorphin 1 μM (+) or 3 μM (++), SB431542 3 μM (+) or 5 μM (++), or ML347 500 nM (+) or 1 mM (++) as indicated with and without GDF2 (10 ng/ml) as indicated (quantification of pSMAD1/5 levels presented in Supplementary Figure S2 C ). (C) Immunoblotting of pSMAD1/5 in PA1 cells in the presence of shRNAs to ALK2, ALK3, ALK6, and BMPRII without and with GDF2 treatment (10 ng/ml) for 30 minutes. (D) QRT-PCR analyses of (C) to confirm reduced expression of ALK2, ALK3, ALK6, and BMPRII expression as indicated. (E) Kinase inactive ALK3 and ALK6 inhibit SMAD1/5 phosphorylation. Western blotting as indicated in MCF10A and PA1 cells in the presence of either mock transfected or HA-tagged kinase inactive ALK3 (ALK3 K-R) or ALK6 (ALK6 K-R) and treated with GDF2 for the time points indicated. Actin was the loading control. (F) Dorsomorphin inhibits SMAD1/5 transcriptional activation. BRE-luciferase reporter activity in indicated cells in the absence (GDF2 alone) or presence of 1 μM dorsomorphin (GDF2+DM). Fold induction of luciferase activity compared with DMSO-treated control cells is presented.

Article Snippet: ALK1/2 inhibitor ML347 (#4945) was from Tocris Bioscience.

Techniques: Phospho-proteomics, Western Blot, Activation Assay, Quantitative RT-PCR, Expressing, Transfection, Control, Luciferase, Activity Assay