Critical Role for Ebf1 and Ebf2 in the Adipogenic Transcriptional Cascade

Abstract

The Ebf (O/E) family of helix-loop-helix transcription factors plays a significant role in B lymphocyte and neuronal development. The three primary members of this family, Ebf1, 2, and 3, are all expressed in adipocytes, and Ebf1 promotes adipogenesis when overexpressed in NIH 3T3 fibroblasts. Here we report that these three proteins have adipogenic potential in multiple cellular models and that peroxisome proliferatoractivated receptor ␥ (PPAR␥) is required for this effect, at least in part due to direct activation of the PPAR␥1 promoter by Ebf1. Ebf1 also directly binds to and activates the C/EBP␣ promoter, which exerts positive feedback on C/EBP␦ expression. Despite this, C/EBP␣ is dispensable for the adipogenic action of Ebf proteins. Ebf1 itself is induced by C/EBP␤ and ␦, which bind and activate its promoter. Reduction of Ebf1 and Ebf2 proteins by specific short hairpin RNA blocks differentiation of 3T3-L1 cells, suggesting a critical role for these factors and the absence of functional redundancy between members of this family. Altogether, these data place Ebf1 within the known transcriptional cascade of adipogenesis and suggest critical roles for Ebf1 and Ebf2.The last decade has seen an enormous surge of interest in the biology of adipocytes, including the developmental processes by which these cells are formed. This has been fuelled by the convergence of a worldwide epidemic of obesity and diabetes (26, 37), with the recent realization that adipose tissue is an active secretory organ regulating a wide array of physiological processes.Adipogenesis represents a complex series of transcriptional events through which multipotent mesenchymal precursor cells become committed to the adipocyte lineage and ultimately express all of the genes typical of mature fat cells (31). These transcriptional events integrate a variety of extracellular signals that direct fat cell formation in time and space. Most of our knowledge concerning the transcriptional events mediating adipogenesis has come from cultured cell lines such as 3T3-L1 and 3T3-F442A (16, 17) that can be differentiated into fat cells by empirically determined hormonal cocktails. These in vitro systems appear to recapitulate most of the developmental events that take place in vivo and offer the advantage of synchronous differentiation and ease of manipulation. Studies of these cells, as well as cultured mouse embryonic fibroblasts (MEFs) derived from mice with various targeted gene ablations (and more recently, studies from intact animals) have revealed a transcriptional cascade in which the nuclear hormone receptor peroxisome proliferator-activated receptor ␥ (PPAR␥) plays a critical role. In cultured adipocytes and fat pads in vivo, PPAR␥ is both necessary and sufficient for adipogenesis (4, 33, 41). Other transcription factors have also been shown to play an important role in adipogenesis, including the CCAAT/enhancer binding proteins C/EBP␣, C/EBP␤, and C/EBP␦ and the basic helix-loop-helix protein SREBP1c (34). In 3T3-L1 cells exposed to a proadipog...

Ebf2 is a selective marker of brown and beige adipogenic precursor cells

“…We speculate that other members of the Ebf family act to support adipogenesis in the absence of Ebf2. In line with this speculation, Ebf1 and Ebf3 can stimulate adipocyte differentiation when expressed in fibroblasts (34). It is also interesting to note that Ebf3 was identified here (Fig.…”

Gene by stress genome-wide interaction analysis and path analysis identify EBF1 as a cardiovascular and metabolic risk gene

“…It binds to and activates the C/EBPα promoter that exerts positive feedback on C/EBPα expression, and in turn induces the expression of PPARγ that joins with EBF1 and C/EBPα to induce adipocyte gene expression. 34 The mouse lipodystrophy is characterized by an increase in yellow adipose tissue in bone marrow and a marked decrease in white adipose tissue, relative to wild-type controls.…”

Adipose tissue in control of metabolism

“…Although both PRDM16 and PGC1α are the main regulators of brown and beige adipocytes, PGC1α is required for the expression of mitochondrial biogenesis and thermogenic genes but not for differentiation genes, indicating that PGC1α is essential for thermogenesis rather than adipogenesis (Uldry et al 2006). Furthermore, adipogenesis is regulated by multiple other factors including positive regulators such as BMPs and early B-cell factor (O/E1), and negative regulators such as transforming growth factor β (TGF-β) and preadipocyte factor 1 (Pref1) (Choy & Derynck 2003, Jimenez et al 2007, Margoni et al 2012). In addition, recent studies have also shown that miRNAs and long noncoding RNA (lncRNA) play important roles in regulating adipogenesis (Park et al 2015).…”