Transforming growth factor-β inhibits adipocyte differentiation by Smad3 interacting with CCAAT/enhancer-binding protein (C/EBP) and repressing C/EBP transactivation function

Abstract

Transforming growth factor (TGF)-β is a potent inhibitor of adipocyte differentiation. To identify which adipocyte transcription factors might be targeted by TGF-β, we overexpressed key adipogenic transcription factors, C/EBPβ, C/EBPδ, or peroxisome proliferator-activated receptor (PPAR) γ in NIH3T3 cells and tested the ability of TGF-β to block adipogenesis. We show that TGF-β inhibits adipocyte differentiation driven by either C/EBPβ or C/EBPδ without affecting C/EBP protein expression levels, suggesting that these C/EBPs are a direct target of TGF-β action. Because TGF-β inhibits adipogenesis by signaling through Smad3, we examined physical and functional interactions of Smad3 and Smad4 with C/EBPβ, C/EBPδ, and PPARγ2. C/EBPβ and C/EBPδ were found to physically interact with Smad3 and Smad4, and Smad3 cooperated with Smad4 and TGF-β signaling to repress the transcriptional activity of C/EBPs. Thus, repression of the activity of C/EBPs by Smad3/4 at C/EBP binding sites inhibited transcription from the PPARγ2 and leptin promoters. In contrast, PPARγ interacted only very weakly with Smad3 and its transcriptional activity was not repressed by Smad3/4 or in response to TGF-β. Smad3/4 did not reduce the ability of C/EBP to bind to its cognate DNA sequence, but repressed transcription by inhibiting the transactivation function of C/EBP.