The forkhead transcription factors, FoxC2 and FoxO1, regulate adipocyte differentiation

In the search for transcription factors that regulate the adipogenic process (formation of fat cells), we investigated two members of the forkhead box family, FoxC2 and FoxO1. Here we show expression of FoxC2 inhibits differentiation in 3T3-L1 preadipocytes. This block is characterized by an extensive decrease in protein expression associated with the function of the mature fat cell. Since the expression of these proteins lies downstream of peroxisome proliferator activated receptor-gamma (PPARgamma), we overexpressed PPARgamma in Swiss mouse fibroblasts to promote adipocyte differentiation. We show that FoxC2 blocks the ability of PPARgamma to induce adipogenic gene expression in the presence of a PPARgamma ligand. Interestingly, the expression of fatty acid binding protein (aP2) escapes this inhibitory action. Finally, we show that FoxC2 does not affect the ability of PPARgamma to bind to or transactivate from a PPARgamma response element, in vitro. However FoxC2 blocks adipogenesis by inhibiting the capacity of PPARgamma to promote the expression of a subset of adipogenic genes. Expression of another forkhead, FoxO1, increases during adipogenesis. However, during the early phase of adipogenesis, FoxO1 is excluded from the nucleus as a result of insulin-induced phosphorylation. Constitutively active FoxO1 which remains in the nucleus, inhibits the adipogenic potential of CCAAT enhancer binding protein-beta (C/EBPbeta), an early adipogenic transcription factor. This block is likely mediated through inhibition of C/EBPbeta's capacity to induce PPARgamma expression. Expression of wild type FoxO1 which is subject to phosphorylation, does not confer this inhibitory effect, suggesting that the phospho-regulation of FoxO1 is required for early adipogenesis to proceed. During terminal adipogenesis, FoxO1 nuclear expression increases, however, overexpression of constitutively active FoxO1 in a terminal adipogenic model attenuates adipogenesis downstream of PPARgamma. Interestingly, wild type FoxO1 seems to enhance adipogenesis in this PPARgamma driven system. In addition, expression of either wild type or constitutively active FoxO1 promotes differentiation in other models of terminal adipogenesis. These data suggest that the retention of FoxO1 in the cytoplasm is required for preadipocytes to enter into terminal adipogenesis. Furthermore, FoxO1 may function to promote aspects of terminal adipogenesis.