Peroxisome proliferator-activated receptor-γ represses GLUT4 promoter activity in primary adipocytes, and rosiglitazone alleviates this effect

Abstract

The synthetic thiazolidinedione ligands of peroxisome proliferator-activated receptor-γ (PPARγ) improve insulin sensitivity in type II diabetes and induce GLUT4 mRNA expression in fat and muscle. However, the molecular mechanisms involved are still unclear. We studied the regulatory effects of PPARγ and its ligands on GLUT4 gene expression in primary rat adipocytes and CHO-K1 cells cotransfected with PPARγ and the GLUT4 promoter reporter. PPARy1 and PPARy2 repressed the activity of the GLUT4 promoter in a dose-dependent manner. Whereas this repression was augmented by the natural ligand 15Δ-prostaglandin J 2, it was completely alleviated by rosiglitazone (Rg). Ligand binding-defective mutants PPARγ1-L468A/E471A and PPARγ2-L496A/E499A retained the repression effect, which was unaffected by Rg, whereas the PPARγ2-S112A mutant exhibited a 50% reduced capacity to repress GLUT4 promoter activity. The -66/+163 bp GLUT4 promoter region was sufficient to mediate PPARγ inhibitory effects. The PPARγ/retinoid X receptor-α heterodimer directly bound to this region, whereas binding was abolished in the presence of Rg. Thus, we show that PPARγ represses transcriptional activity of the GLUT4 promoter via direct and specific binding of PPARγ/retinoid X receptor-α to the GLUT4 promoter. This effect requires an intact Ser112 phosphorylation site on PPARγ and is completely alleviated by Rg, acting via its ligand-binding domain. These data suggest a novel mechanism by which Rg exerts its antidiabetic effects via detaching PPARγ from the GLUT4 gene promoter, thus leading to increased GLUT4 expression and enhanced insulin sensitivity.