Received for publication December 23, 2005.
Revised February 15, 2006.
Accepted for publication March 13, 2006.

A novel vitamin D derivative activates bone morphogenetic protein (BMP) signaling in MCF10 breast epithelial cells

Abstract

We investigated the action of 1,25-dihydroxyvitamin D₃ (1,25(OH)₂D₃) as well as a novel Gemini vitamin D₃ analog Ro-438-3582 [1,25-dihydroxy-20S-21(3-hydroxy-3-methyl-butyl)-23-yne-26,27-hexafluorocholecalciferol; Ro3582] and a classical vitamin D₃ analog Ro-26-2198 (1,25-dihydroxy-16,23Z-diene-26,27-hexafluoro-19-nor-cholecalciferol; Ro2198) in modulating the transforming growth factor- (TGF-)/bone morphogenetic protein (BMP) system in MCF10 immortalized breast epithelial cells. We found that 1,25(OH)₂D₃, Ro3582 and Ro2198 all enhanced BMP/Smad signaling by increasing the phosphorylation of receptor-regulated Smads. Ro3582 was more active than Ro2198 but both were considerably more active than 1,25(OH)2D3. Ro3582 enhanced BMP/Smad signaling by (a) inducing the phosphorylation of receptor-regulated Smads (Smad1/5), (b) increasing the accumulation of phosphorylated Smad1/5 in the nucleus, and (c) activating BMP-mediated transcription in MCF10 breast epithelial cells. Furthermore, Ro3582 induced the synthesis of BMP-2 and BMP-6 mRNA and protein, and the expression of Smad6 mRNA in MCF10 breast epithelial cells was inhibited. The induction of phospho-Smad1/5 by Ro3582 was inhibited by treatment with the BMP antagonist, Noggin, while neutralizing antibody to TGF- did not block the induction of phospho-Smad1/5 by Ro3582. Treatment with Noggin also blocked the effect of Ro3582 on nuclear accumulation of phospho-Smad1/5 and the induction of BMP-2 and BMP-6 mRNA synthesis. These results indicate that the activation of BMP/Smad signaling by the Gemini vitamin D3 analog Ro3582 may be through the production of BMP ligands including BMP-2 and BMP-6 and/or down-regulation of the inhibitory Smad6. This is the first report to show that 1,25(OH)₂D₃ and its derivatives activate BMP/Smad specific signaling in human breast epithelial cells.

Key words: Growth hormone, Vitamin D, Signaling network analyses, Fluorescence techniques, Regulation - transcriptional

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