FOXO1 represses peroxisome proliferator-activated receptor-γ1 and -γ2 gene promoters in primary adipocytes -: A novel paradigm to increase insulin sensitivity

FOXO1 and peroxisome proliferator-activated receptor-gamma (PPAR gamma) are crucial transcription factors that regulate glucose metabolism and insulin responsiveness in insulin target tissues. We have shown that, in primary rat adipocytes, both factors regulate transcription of the insulin-responsive GLUT4 gene and that PPAR gamma 2 detachment from the GLUT4 promoter upon thiazolidinedione binding up-regulates GLUT4 gene expression, thus increasing insulin sensitivity (Armoni, M., Kritz, N., Harel, C., Bar-Yoseph, F., Chen, H., Quon, M. J., and Karnieli, E. (2003) J. Biol. Chem. 278, 30614-30623). However, the mechanisms regulating PPAR gamma gene transcription are largely unknown. We studied the effects of FOXO1 on human PPAR gamma gene expression in primary rat adipocytes and found that both genes are endogenously expressed. FOXO1 coexpression dose-dependently repressed transcription from either the PPAR gamma 1 or PPAR gamma 2 promoter reporter by 65%, whereas insulin (100 nM, 20-24 h) either partially or completely reversed this effect. Phosphorylation-defective FOXO1 mutants T24A, S256A, S319A, and T24A/S256A/S319A still repressed the PPAR gamma 1 promoter and partially lost their effects on the PPAR gamma 2 promoter in either basal or insulin-stimulated cells. Use of DNA binding-defective FOXO1(H215R) indicated that this domain is crucial for FOXO1 repression of the PPAR gamma 2 (but not PPAR gamma 1) promoter. Progressive 5'-deletion and gel retardation analyses revealed that this repression involves direct and specific binding of FOXO1 to the PPAR gamma 2 promoter; chromatin immunoprecipitation analysis confirmed that this binding occurs in cellulo. We suggest a novel paradigm to increase insulin sensitivity in adipocytes in which FOXO1 repression of PPAR gamma, the latter being a repressor of the GLUT4 promoter, consequently leads to GLUT4 derepression/up-regulation, thus enhancing cellular insulin sensitivity. The newly identified FOXO1-binding site on the PPAR gamma 2 promoter may serve as a therapeutic target for type 2 diabetes.