Summary Brown fat can increase energy expenditure and protect against obesity through a specialized program of uncoupled respiration. We show here by in vivo fate mapping that brown but not white fat cells arise from precursors that express myf5, a gene previously thought to be expressed only in the myogenic lineage. Notably, the transcriptional regulator, PRDM16 controls a bidirectional cell fate switch between skeletal myoblasts and brown fat cells. Loss of PRDM16 from brown fat precursors causes a loss of brown fat characteristics and promotes muscle differentiation. Conversely, ectopic expression of PRDM16 in myoblasts induces their differentiation into brown fat cells. PRDM16 stimulates brown adipogenesis by binding to PPARγ and activating its transcriptional function. Finally, PRDM16-deficient brown fat displays an abnormal morphology, reduced thermogenic gene expression and elevated expression of muscle-specific genes. Taken together, these data indicate that PRDM16 specifies the brown fat lineage from a progenitor that expresses myoblast markers and is not involved in white adipogenesis.

The white adipose organ is composed of both subcutaneous and several intra-abdominal depots. Excess abdominal adiposity is a major risk factor for metabolic disease in rodents and humans, while expansion of subcutaneous fat does not carry the same risks. Brown adipose produces heat as a defense against hypothermia and obesity, and the appearance of brown-like adipocytes within white adipose tissue depots is associated with improved metabolic phenotypes. Thus, understanding the differences in cell biology and function of these different adipose cell types and depots may be critical to the development of new therapies for metabolic disease. Here, we found that Prdm16, a brown adipose determination factor, is selectively expressed in subcutaneous white adipocytes relative to other white fat depots in mice. Transgenic expression of Prdm16 in fat tissue robustly induced the development of brown-like adipocytes in subcutaneous, but not epididymal, adipose depots. Prdm16 transgenic mice displayed increased energy expenditure, limited weight gain, and improved glucose tolerance in response to a high-fat diet. shRNA-mediated depletion of Prdm16 in isolated subcutaneous adipocytes caused a sharp decrease in the expression of thermogenic genes and a reduction in uncoupled cellular respiration. Finally, Prdm16 haploinsufficiency reduced the brown fat phenotype in white adipose tissue stimulated by β-adrenergic agonists. These results demonstrate that Prdm16 is a cell-autonomous determinant of a brown fat-like gene program and thermogenesis in subcutaneous adipose tissues. IntroductionThe rise in the incidence of obesity has driven a public health crisis because excess adiposity predisposes to cardiovascular disease, type 2 diabetes, hypertension, stroke, and many cancers (1, 2). Since weight gain is almost always caused by chronic energy imbalance, nonsurgical therapy for obesity must reduce energy intake and/or increase energy expenditure.There are 2 major types of adipose tissues in mammals, white and brown. White adipose tissue (WAT) is highly adapted to store excess energy in the form of triglycerides. Conversely, brown adipose tissue (BAT) oxidizes chemical energy to produce heat as a defense against hypothermia and obesity. WAT develops in distinct intra-abdominal depots and in the subcutaneous layer (between the fascia and muscle). The accumulation of intra-abdominal, visceral WAT, rather than total adiposity, is most strongly correlated with elevated risk for metabolic dysfunction and cardiovascular disease (3-8). By contrast, expansion of subcutaneous WAT, even in the setting of obesity, has been suggested to promote insulin sensitivity in rodents and humans (9)(10)(11)(12)(13)(14). Implantation of subcutaneous WAT, but not visceral WAT, into the abdominal cavity of mice improves whole-body metabolism (15, 16). Moreover, subcutaneous and visceral WAT express unique gene signatures (17). These data suggest that some of the distinct metabolic effects of subcutaneous and visceral WAT are cell autonomous. However, the ...

The worldwide epidemic of obesity has increased the urgency of developing a deeper understanding of physiological systems related to energy balance and energy storage, including the mechanisms controlling the development of fat cells (adipocytes). The differentiation of committed preadipocytes to adipocytes is controlled by PPARγ and several other transcription factors 1, but the molecular basis for preadipocyte determination is not understood. Using a novel method for the quantitative analysis of transcriptional components, we identified the zinc-finger protein Zfp423 as a factor enriched in preadipose versus non-preadipose fibroblasts. Ectopic expression of Zfp423 in non-adipogenic NIH 3T3 fibroblasts robustly activates expression of PPARγ in undifferentiated cells and permits cells to undergo adipocyte differentiation under permissive conditions. ShRNA-mediated reduction of Zfp423 expression in 3T3-L1 cells blunts preadipocyte PPARγ expression and diminishes the ability of these cells to differentiate. Furthermore, both brown and white adipocyte differentiation is strikingly impaired in Zfp423-deficient mouse embryos. Zfp423 regulates PPARγ expression, in part, through amplification of the BMP signaling pathway, an effect dependent on the SMAD binding capacity of Zfp423. This study identifies Zfp423 as a transcriptional regulator of preadipocyte determination.

Fibroblastic preadipocyte cells are recruited to differentiate into new adipocytes during the formation and hyperplastic growth of white adipose tissue. Peroxisome proliferator-activated receptor ␥ (PPAR␥), the master regulator of adipogenesis, is expressed at low levels in preadipocytes, and its levels increase dramatically and rapidly during the differentiation process. However, the mechanisms controlling the dynamic and selective expression of PPAR␥ in the adipocyte lineage remain largely unknown. We show here that the zinc finger protein Evi1 increases in preadipocytes at the onset of differentiation prior to increases in PPAR␥ levels. Evi1 expression converts nonadipogenic cells into adipocytes via an increase in the predifferentiation levels of PPAR␥2, the adipose-selective isoform of PPAR␥. Conversely, loss of Evi1 in preadipocytes blocks the induction of PPAR␥2 and suppresses adipocyte differentiation. Evi1 binds with C/EBP␤ to regulatory sites in the Ppar␥ locus at early stages of adipocyte differentiation, coincident with the induction of Ppar␥2 expression. These results indicate that Evi1 is a key regulator of adipogenic competency. O besity is a major risk factor for many diseases, including type 2 diabetes, cardiovascular disease, stroke, and many cancers (10, 15). Weight gain occurs when energy intake from food chronically exceeds energy expenditure through physical activity and metabolism. Excess energy is stored as triglycerides in adipose tissue, which expands through increases in the size (hypertrophy) and/or number (hyperplasia) of adipocytes. The development and maintenance of an appropriate mass of adipose tissue are crucial for systemic metabolic health because either insufficient or excess tissue leads to insulin resistance and metabolic disease.New adipocytes are thought to arise from committed populations of fibroblastic cells resident within adipose tissues, so-called preadipocytes (reviewed in reference 6). Recent data show that these adipogenic precursors are intimately associated with the vasculature and express particular cell surface markers (16,30,41). Preadipocytes purified from adipose tissue can undergo adipogenic differentiation in culture, but there is substantial cellular heterogeneity within these isolates. Immortal preadipocyte cell lines (e.g., 3T3-L1 and 3T3-F442A cells) derived from mouse embryo fibroblasts undergo a highly conserved and efficient program of adipogenesis in culture and upon transplantation in vivo. These cell lines have provided a powerful and tractable model system to elucidate the transcriptional networks of adipogenesis.The nuclear hormone receptor peroxisome proliferator-activated receptor ␥ (PPAR␥) and members of the C/EBP (CCAATenhancer-binding protein) family of transcription factors orchestrate the adipogenic differentiation process. PPAR␥ is considered to be the "master regulator" of adipose cell differentiation since it is both necessary and sufficient for adipocyte development (1,31,42). Genome-wide analyses show that PPAR␥ binds and regulate...