Peroxisome Proliferator-activated Receptor γ Activation Modulates Cyclin D1 Transcription via β-Catenin-independent and cAMP-response Element-binding Protein-dependent Pathways in Mouse Hepatocytes

Abstract

Activation of peroxisome proliferator-activated receptor γ (PPARγ) following exposure to PPARγ-specific ligands resulted in growth inhibition in various carcinoma cell lines. Our aim was to elucidate the pathway of PPARγ2 activation-mediated modulation of cyclin D1 transcription in mouse hepatocytes. To address this we utilized stable control and PPARγ hepatocyte cell lines created via retroviral overexpression utilizing AML-12 hepatocytes. Addition of PPARγ ligand troglitazone (TZD) activated PPARγ2 in proliferating hepatocytes and resulted in growth arrest accompanied by a down-regulation of proliferating cell nuclear antigen, cyclin D1, and β-catenin expression. Furthermore activation of PPARγ2 attenuated cyclin D1 promoter activity indicating a transcriptional regulation of cyclin D1. Since β-catenin plays a pivotal role in regulating cyclin D1 transcription, we studied whether PPARγ2-mediated inhibition of cyclin D1 transcription involved β-catenin. Interestingly overexpression of either wild-type or S37A mutant β-catenin was unable to rescue PPARγ2-mediated suppression of cyclin D1 transcription, whereas overexpression of cAMP-response element-binding protein (CREB) was capable of antagonizing this inhibitory effect of PPARγ2. Additionally pretreatment with okadaic acid antagonized PPARγ2-mediated inhibition of cyclin D1 transcription without any effect on β-catenin expression. These studies also showed a TZD-mediated inhibition of total and phospho-CREBSer133 levels, CREB promoter activity, and cAMP-response element-mediated transcription in PPARγ hepatocytes. Pretreatment of PPARγ hepatocytes with okadaic acid, however, maintained higher total and phospho-CREBSer133 levels in the presence of TZD. These results indicated that PPARγ2 activation inhibited cyclin D1 transcription in hepatocytes via CREB-dependent and β-catenin-independent pathways.