The role of PPAR-γ on the adipocyte differentiation and the improvement of insulin resistance

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Peroxisome proliferator-activated receptor (PPAR) γ is a member of the nuclear receptor superfamily. It has been known to play a pivotal role in the adipocyte differentiation and insulin sensitivity, although its function has not been fully understood. The discovery of PPARγ mutants in patients with severe insulin resistance and hypertension indicates that the dominant negative effect of these mutants have a critical role in the clinical phenotypes. To elucidate the mechanism of the dominant negative effect by naturally occurring mutant PPARγ, we constructed a series of the chimeric PPARγs in which the ligand binding domain was substituted with the T3 binding domain of TRβ. These chimeras were transfected into CV-1 cells or 3T3-L1 preadipocytes together with the luciferase reporter plasmid containing PPAR responsive element (PPRE) fused to a thymidine kinase promoter. One of the chimeric receptors, PPP showed T3 dependent transcriptional activity, comparable to that of wild type PPARγ when stimulated by troglitazone. In the absence of T3, PPP inhibited the transcriptional activity of wild type PPARγ in a dominant negative fashion. Introducing a mutation into the CoR box, which is required for corepressor binding destroyed the dominant negative effect of PPP. Furthermore, the deletion of the A/B domain in PPP and mutant PPARγ, P467L, strongly enhanced the dominant negative effect. The mammalian two hybrid assay and glutathione S-transferase (GST) pull down assay revealed that deletion of the A/B domain increased the affinity of the receptor to the corepressors. The present study suggests that the A/B domain inhibits the dominant negative effect of mutant PPARγ, probably due to the inter-domain communication between the N-terminal A/B domain and the C-terminal ligand binding domain.