Activation of CCAAT/enhancer-binding protein (C/EBP) α expression by C/EBPβ during adipogenesis requires a peroxisome proliferator-activated receptor-γ-associated repression of HDAC1 at the C/ebpα gene promoter

Abstract

Studies have shown that CCAAT/enhancer-binding protein β (C/EBPβ) can stimulate adipogenesis in noncommitted fibroblasts by activating expression of peroxisome proliferator-activated receptor-γ (PPARγ). Other investigations have established a role for C/EBPα as well as PPARγ in orchestrating the complex program of adipogenic gene expression during terminal preadipocyte differentiation. Consequently, it is important to identify factors regulating transcription of the C/ebpα gene. In this study, we demonstrated that inhibition of PPARγ activity by exposure of 3T3-L1 preadipocytes to a potent and selective PPARγ antagonist inhibits adipogenesis but also blocks the activation of C/EBPα expression at the onset of differentiation. Ectopic expression of C/EBPβ in Swiss 3T3 mouse fibroblasts (Swiss-LAP cells) induces PPARγ expression without any significant enhancement of C/EBPα expression. Treatment of Swiss-LAP cells with a PPARγ agonist induces adipogenesis, which includes activation of C/EBPα expression. To further establish a role for PPARγ in regulating C/EBPα expression, we expressed C/EBPβ in PPARγ-deficient mouse embryo fibroblasts (MEFs). The data show that C/EBPβ is capable of inducing PPARγ in Pparγ+/-MEFs, which leads to activation of adipogenesis, including C/EBPα expression following exposure to a PPARγ ligand. In contrast, C/EBPβ is not able to induce C/EBPα expression or adipogenesis in Pparγ-/- MEFs. Chromatin immunoprecipitation analysis reveals that C/EBPβ is bound to the minimal promoter of the C/ebpα gene in association with HDAC1 in unstimulated Swiss-LAP cells. Exposure of the cells to a PPARγ ligand dislodges HDAC1 from the proximal promoter of the C/ebpα gene, which involves degradation of HDAC1 in the 26 S proteasome. These data suggest that C/EBPβ activates a single unified pathway of adipogenesis involving its stimulation of PPARγ expression, which then activates C/EBPα expression by dislodging HDAC1 from the promoter for degradation in the proteasome. © 2006 by The American Society for Biochemistry and Molecular Biology, Inc.