A detailed understanding of the processes governing adipose tissue formation will be instrumental in combating the obesity epidemic. Much progress has been made in the last two decades in defining transcriptional events controlling the differentiation of mesenchymal stem cells into adipocytes. A complex network of transcription factors and cell-cycle regulators, in concert with specific transcriptional coactivators and corepressors, respond to extracellular stimuli to activate or repress adipocyte differentiation. This review summarizes advances in this field, which constitute a framework for potential antiobesity strategies.

SUMMARY Brown adipose tissue (BAT) can disperse stored energy as heat. Promoting BAT-like features in white adipose (WAT) is an attractive, if elusive therapeutic approach to staunch the current obesity epidemic. Here we report that gain-of-function of the NAD-dependent deacetylase SirT1 or loss-of-function of its endogenous inhibitor Deleted in breast cancer-1 (Dbc1) promote “browning” of WAT by deacetylating peroxisome proliferator-activated receptor (Ppar)-γ on Lys268 and Lys293. SirT1-dependent deacetylation of Lys268 and Lys293 is required to recruit the BAT program coactivator Prdm16 to Pparγ, leading to selective induction of BAT genes and repression of visceral WAT genes associated with insulin resistance. An acetylation-defective Pparγ mutant induces a brown phenotype in white adipocytes, while an acetylated mimetic fails to induce “brown” genes, but retains the ability to activate “white” genes. We propose that SirT1-dependent Pparγ deacetylation is a form of selective Pparγ modulation of potential therapeutic import.

We demonstrate that exposure of post-confluent 3T3-L1 preadipocytes to insulin, isobutylmethylxanthine (MIX), dexamethasone (DEX), and fetal bovine serum induces a rapid but transient activation of MEK1 as indicated by extensive phosphorylation of ERK1 and ERK2 during the initial 2 h of adipogenesis. Inhibition of this activity by treating the cells with a MEK1-specific inhibitor (U0126 or PD98059) prior to the induction of differentiation significantly attenuated the expression of peroxisome proliferator-activated receptor (PPAR) ␥, CCAAT/enhancer-binding protein (C/EBP) ␣, perilipin, and adipocyte-specific fatty acid-binding protein (aP2). Treating the preadipocytes with troglitazone, a potent PPAR␥ ligand, could circumvent the inhibition of adipogenic gene expression by U0126. Fibroblast growth factor-2 (FGF-2), in the presence of dexamethasone, isobutylmethylxanthine, and insulin, induces a prolonged activation of the MEK/ERK signaling pathway, which lasts for at least 12 h post-induction, and this activity is less sensitive to the MEK inhibitors. Consequently, preadipocytes treated with U0126 in the presence of fibroblast growth factor-2 (FGF-2) express normal post-induction levels of MEK activity, and, in so doing, are capable of undergoing adipogenesis. We further show that activation of MEK1 significantly enhances the transactivation of the C/EBP␣ minimal promoter during the early phase of the differentiation process. Our results suggest that activation of the MEK/ ERK signaling pathway during the initial 12 h of adipogenesis enhances the activity of factors that regulate both C/EBP␣ and PPAR␥ expression.

The differentiation of 3T3 preadipocytes into adipocytes is accompanied by a transient induction of C/EBP␤ and C/EBP␦ expression in response to treatment of the cells with methylisobutylxanthine (MIX) and dexamethasone (DEX), respectively. In this report, we demonstrate that peroxisome proliferator-activated receptor ␥ (PPAR␥) expression in 3T3-L1 preadipocytes is induced by MIX and DEX, suggesting that C/EBP␤ and C/EBP␦ may be involved in this process. Using a tetracycline-responsive expression system, we have recently shown that the conditional ectopic expression of C/EBP␤ in NIH 3T3 fibroblasts (␤2 cells) in the presence of DEX activates the synthesis of peroxisome PPAR␥ mRNA. Subsequent exposure of these cells to PPAR activators stimulates their conversion into adipocytes; however, neither the expression of C/EBP␤ nor exposure to DEX alone is capable of inducing PPAR␥ expression in the ␤2 cell line. We find that unlike the case for 3T3 preadipocytes, C/EBP␦ is not induced by DEX in these 3T3 fibroblasts and therefore is not relaying the effect of this glucocorticoid to the PPAR␥ gene. To define the role of glucocorticoids in regulating PPAR␥ expression and the possible involvement of C/EBP␦, we have established an additional set of NIH 3T3 cell lines expressing either C/EBP␦ alone (␦23 cells) or C/EBP␦ and C/EBP␤ together (␤/␦39 cells), using the tetracyclineresponsive system. Culture of these cells in tetracycline-deficient medium containing DEX, MIX, insulin, and fetal bovine serum shows that the ␤/␦39 cells express PPAR␥ and aP2 mRNAs at levels that are almost equivalent to those observed in fully differentiated 3T3-L1 adipocytes. These levels are approximately threefold higher than their levels of expression in the ␤2 cells. Despite the fact that these ␤/␦39 cells produce abundant amounts of C/EBP␤ and C/EBP␦ (in the absence of tetracycline), they still require glucocorticoids to attain maximum expression of PPAR␥ mRNA. Furthermore, the induction of PPAR␥ mRNA by exposure of these cells to DEX occurs in the absence of ongoing protein synthesis. The ␦23 cells, on the other hand, are not capable of activating PPAR␥ gene expression when exposed to the same adipogenic inducers. Finally, attenuation of ectopic C/EBP␤ production at various stages during the differentiation process results in a concomitant inhibition of PPAR␥ and the adipogenic program. These data strongly suggest that the induction of PPAR␥ gene expression in multipotential mesenchymal stem cells (NIH 3T3 fibroblasts) is dependent on elevated levels of C/EBP␤ throughout the differentiation process, as well as an initial exposure to glucocorticoids. C/EBP␦ may function by synergizing with C/EBP␤ to enhance the level of PPAR␥ expression.

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