Ebf2 is a selective marker of brown and beige adipogenic precursor cells (RNA-seq)

Brown adipocytes, muscle and dorsal dermis descend from precursor cells in the dermomyotome, but the factors that regulate commitment to the brown adipose lineage are unknown. Here, we prospectively isolated and determined the molecular profile of embryonic brown preadipose cells. Brown adipogenic precursor activity in embryos was confined to Pdgfralpha+, Myf5Cre-lineage-marked cells. RNAseq analysis identified Early B Cell Factor-2 (Ebf2) as one of the most selectively expressed genes in this cell fraction. Importantly, Ebf2-expressing cells purified from Ebf2-GFP embryos or brown fat tissue did not express myoblast or dermal cell markers and uniformly differentiated into brown adipocytes. Interestingly, Ebf2-expressing cells from white fat tissue in adult animals differentiated into brown-like (or beige) adipocytes. Loss of Ebf2 in brown preadipose cells reduced the expression levels of brown preadipose-signature genes, whereas ectopic Ebf2-expression in myoblasts activated brown preadipose-specific genes. Altogether, these results indicate that Ebf2 specifically marks and regulates the molecular profile of brown preadipose cells. Embryonic fibroblasts, isolated from dorsal body wall of E14.5 Myf5-CrE;mTmG embryos, were further fractionated based on the expression of PDGFRalpha, Itga7 and Myf5-cre (GFP). Total mRNA profiles from Myf5-cre(GFP)+PDGFRalpha+ and Myf5-cre(GFP)+PDGFRalpha-Itga7+ cells were generated by deep sequencing棕色脂肪细胞、肌肉及背部真皮起源于生皮肌节中的前体细胞，但调控棕色脂肪谱系定向分化的分子因子仍未明确。本研究前瞻性分离并解析了胚胎棕色脂肪前体细胞的分子特征。胚胎中的棕色脂肪生成前体活性局限于PDGFRα阳性（Pdgfralpha+）、经Myf5Cre谱系标记的细胞群体。通过RNA测序（RNAseq）分析，鉴定出早期B细胞因子2（Early B Cell Factor-2, Ebf2）是该细胞组分中表达特异性最高的基因之一。值得注意的是，从Ebf2启动子驱动绿色荧光蛋白（GFP）表达的转基因胚胎或棕色脂肪组织中纯化的表达Ebf2的细胞，不表达成肌细胞或真皮细胞标志物，且可均一分化为棕色脂肪细胞。有趣的是，成年动物白色脂肪组织中分离的表达Ebf2的细胞可分化为棕色样（或称米色）脂肪细胞。在棕色脂肪前体细胞中敲除Ebf2会降低棕色脂肪前体细胞特征基因的表达水平，而在成肌细胞中异位表达Ebf2则可激活棕色脂肪前体细胞特异性基因。综上，上述结果表明Ebf2可特异性标记并调控棕色脂肪前体细胞的分子特征。从胚胎第14.5天（E14.5）Myf5-Cre;mTmG胚胎的背体壁分离的胚胎成纤维细胞，进一步根据PDGFRα（Pdgfralpha）、整合素α7（Itga7）及Myf5-Cre（GFP）的表达进行分馏。对Myf5-Cre(GFP)+PDGFRα+与Myf5-Cre(GFP)+PDGFRα-Itga7+细胞的总mRNA表达谱进行了深度测序。