A glucocorticoid- and diet-responsive pathway toggles adipocyte precursor cell activity in vivo.

Obesity is driven by excess caloric intake, which leads to the expansion of adipose tissue by hypertrophy and hyperplasia. Adipose tissue hyperplasia results from the differentiation of adipocyte precursor cells (APCs) that reside in adipose depots. Investigation into this process has elucidated a network of mostly transcription factors that drive APCs through the differentiation process. Using in vitro and in vivo approaches, our study revealed a signaling pathway that inhibited the initiation of the adipocyte differentiation program. Mouse adipocytes secreted the extracellular protease ADAMTS1, which triggered the production of the cytokine pleiotrophin (PTN) through the Wnt/ß-catenin pathway, and promoted proliferation rather than differentiation of APCs. Glucocorticoid exposure in vitro or in vivo reduced ADAMTS1 abundance in adipocytes. In addition, mice fed a high-fat diet showed decreased Adamts1 expression in the visceral perigonadal adipose depot, which expanded by adipogenesis in response to the diet, and increased Adamts1 expression in the subcutaneous inguinal adipose depot, which did not induce adipogenesis. Similar to what occurred in mouse subcutaneous adipose tissue, diet-induced weight gain increased the expression of ADAMTS1, PTN, and certain Wnt target genes in the subcutaneous adipose depot of human volunteers, suggesting the relevance of this pathway to physiological adipose tissue homeostasis and the pathogenesis of obesity. Thus, this pathway functions as a toggle on APCs, regulating a decision between differentiation and proliferation and coordinating the response of adipose tissue to systemic cues.

Characterization of Cre recombinase activity for in vivo targeting of adipocyte precursor cells.

The increased incidence of obesity and metabolic disease underscores the importance of elucidating the biology of adipose tissue development. The recent discovery of cell surface markers for prospective identification of adipose precursor cells (APCs) in vivo will greatly facilitate these studies, yet tools for specifically targeting these cells in vivo have not been identified. Here, we survey three transgenic mouse lines, Fabp4-Cre, PdgfRα-Cre, and Prx1-Cre, precisely assessing Cre-mediated recombination in adipose stromal populations and mature tissues. Our data provide key insights into the utility of these tools to modulate gene expression in adipose tissues. In particular, Fabp4-Cre is not effective to target APCs, nor is its activity restricted to these cells. PdgfRα-Cre directs recombination in the vast majority of APCs, but also targets other populations. In contrast, adipose expression of Prx1-Cre is chiefly limited to subcutaneous inguinal APCs, which will be valuable for dissection of APC functions among adipose depots.

Adipose circadian clocks: coordination of metabolic rhythms by clock genes, steroid hormones, and PPARs.

A central clock consisting of interconnected positive and negative feedback gene loops operates in the brain, tying rhythmic activity to the 24-h day. The central clock entrains similar feedback loops present in most peripheral tissues to coordinate metabolic gene expression among organs and with feeding activity for more efficient utilization of resources. Recent studies are beginning to elucidate the intricate feedback mechanisms among central and peripheral clocks and their roles in activity and metabolic homeostasis. Adipose tissue serves as a major energy storage organ and releases paracrine and endocrine hormones to signal energy status to other organs. Within the adipose tissue, the transcriptional feedback regulation between clock genes and nuclear hormone receptors, together with direct protein associations among these molecules, ensures the expression of metabolic genes at the appropriate time. This review will summarize the important components and mechanisms of adipose clock entrainment, particularly highlighting instructive studies carried out in mice. This research not only illustrates the intricate connections between clocks and metabolism but also provides potential mechanisms to correct abnormalities induced by disrupted sleep or poor diet.

Circadian rhythm gene period 3 is an inhibitor of the adipocyte cell fate.

Glucocorticoids rapidly and robustly induce cell fate decisions in various multipotent cells, although the precise mechanisms of these important cellular events are not understood. Here we showed that glucocorticoids repressed Per3 expression and that this repression was critical for advancing mesenchymal stem cells to the adipocyte fate. Exogenous expression of Per3 inhibited adipogenesis, whereas knocking out Per3 enhanced that fate. Moreover, we found that PER3 formed a complex with PPARγ and inhibited PPARγ-mediated transcriptional activation via Pparγ response elements. Consistent with these findings, Per3 knock-out mice displayed alterations in body composition, with both increased adipose and decreased muscle tissue compared with wild-type mice. Our findings identify Per3 as potent mediator of cell fate that functions by altering the transcriptional activity of PPARγ.

Serotonergic transcription of human FEV reveals direct GATA factor interactions and fate of Pet-1-deficient serotonin neuron precursors.

Altered expression of the human FEV (fifth Ewing variant) ETS transcription factor gene impacts the level of CNS serotonin (5-HT) neuron gene expression and maternal nurturing. However, the regulatory mechanisms that determine FEV expression are poorly understood. Here, we investigated the cis-regulatory control of FEV to begin to identify the upstream transcription factors that restrict FEV expression to 5-HT neurons. We find that sequences extending only 275 bp upstream of the FEV 5' untranslated region are sufficient to direct FEV transgene expression to embryonic 5-HT neurons, although sequences farther upstream are required for maintenance in adult 5-HT neurons. Two highly conserved consensus GATA factor binding sites within the 275 bp region interact with GATA factors in vitro. Chromatin immunoprecipitations with embryonic hindbrain demonstrated Gata-2 interactions with the orthologous mouse Pet-1 ETS cis-regulatory region. Mutagenesis of GATA sites revealed that one or the other site is required for serotonergic FEV transgene expression. Unexpectedly, FEV-LacZ transgenes enabled determination of 5-HT neuron precursor fate in the adult Pet-1(-/-) dorsal and median raphe nuclei and thus provided additional insight into FEV/Pet-1 function. Comparable numbers of FEV-LacZ-positive cells were detected in Pet-1(+/-) and Pet-1(-/-) adult dorsal raphe nuclei, indicating that the majority of mutant serotonergic precursors are not fated to apoptosis. However, B7 dorsal raphe cells were aberrantly distributed, suggesting a role for FEV/Pet-1 in their midline organization. Our findings identify a direct transcriptional interaction between Gata-2 and FEV and a unique marker for new insight into FEV/Pet-1 function in 5-HT neuron development.

A differentially autoregulated Pet-1 enhancer region is a critical target of the transcriptional cascade that governs serotonin neuron development.

The Pet-1 [pheochromocytoma 12 ETS (E26 transformation-specific)] gene plays a critical role in the development of serotonin (5-HT)-modulated behaviors via its control of embryonic 5-HT neuron differentiation. Pet-1 transcription is induced exclusively in 5-HT neuron postmitotic precursors before the appearance of transmitter, and its restricted expression is maintained in the adult. However, the mechanisms that direct Pet-1 expression to this single CNS neuronal cell type are unknown. Here, we show, using transgenic methods, that genomic sequences upstream, but not downstream or within the Pet-1-coding region, are sufficient for 5-HT neuron-specific transgene expression. Enhancer sequences within a 40 kb upstream fragment directed position-independent lacZ (beta-D-galactosidase) transgene expression to the developing hindbrain before the appearance of 5-HT. Moreover, virtually all of the 5-HT neurons in the adult were lacZ positive in all of the lines examined. Transgene expression in 5-HT neurons was maintained when the 40 kb fragment was truncated on its 5' end to either 12 or 1.8 kb, although position independence was then lost. Analysis of transgene expression in Pet-1 null mice indicated that Pet-1 was required to maintain the activity of the Pet-1 enhancer region in a subset of 5-HT neurons. These findings suggest that a conserved 1.8 kb region immediately flanking the Pet-1-coding region is a critical genomic target of the transcriptional cascade that governs 5-HT neuron development and provide additional evidence for 5-HT neuron heterogeneity at the genetic level. We discuss the potential application of the Pet-1 transgenes reported here to the selective genetic manipulation of 5-HT neurons.