Department of Biochemistry and Cancer Biology, University of Toledo College of Medicine, Toledo, Ohio, USA.
PLoS One. 2013 Jun 17;8(6):e65531. doi: 10.1371/journal.pone.0065531. Print 2013.
Differentiation of Embryonic Stem Cells 1 (Dies1) was recently identified as a novel type I immunoglobulin (IgG) domain-containing plasma membrane protein important for effective differentiation of a murine pluripotent embryonic stem cell line. In this setting, Dies1 enhances bone morphogenetic protein 4 (BMP4) signaling. Here we show Dies1 transcript expression is induced ∼225-fold during in vitro adipogenesis of 3T3-L1 murine preadipocytes. Immunocytochemical imaging using ectopic expression of Flag-tagged Dies1 in 3T3-L1 adipocytes revealed localization to the adipocyte plasma membrane. Modulation of adipocyte phenotype with with tumor necrosis factor-α (TNFα) treatment or by siRNA knockdown of the master pro-adipogenic transcription factor peroxisome proliferator activated receptor gamma (PPARγ) resulted in a 90% and 60% reduction of Dies1 transcript levels, respectively. Moreover, siRNA-mediated Dies1 knockdown in 3T3-L1 preadipocytes inhibited adipogenic conversion. Such cultures had a 35% decrease in lipid content and a 45%-65% reduction in expression of key adipocyte transcripts, including that for PPARγ. The standard protocol for full in vitro adipogenic conversion of committed preadipocytes, such as 3T3-L1, does not include BMP4 treatment. Thus we posit the positive role of Dies1 in adipogenesis, unlike that for Dies1 in differentiation of embryonic stem cells, does not include its pro-BMP4 effects. In support of this idea, 3T3-L1 adipocytes knocked down for Dies1 did not evidence decreased phospho-Smad1 levels upon BMP4 exposure. qPCR analysis of Dies1 transcript in multiple murine and human tissues reveals high enrichment in white adipose tissue (WAT). Interestingly, we observed a 10-fold induction of Dies1 transcript in WAT of fasted vs. fed mice, suggesting a role for Dies1 in nutritional response of mature fat cells in vivo. Together our data identify Dies1 as a new differentiation-dependent adipocyte plasma membrane protein whose expression is required for effective adipogenesis and that may also play a role in regard to nutritional status in WAT.
胚胎干细胞 1（Dies1）的分化最近被确定为一种新型的 I 型免疫球蛋白（IgG）结构域包含的质膜蛋白，对于有效分化小鼠多能胚胎干细胞系非常重要。在这种情况下，Dies1 增强了骨形态发生蛋白 4（BMP4）信号。在这里，我们显示 Dies1 转录物的表达在体外 3T3-L1 鼠前脂肪细胞的脂肪生成过程中被诱导约 225 倍。在 3T3-L1 脂肪细胞中异位表达 Flag 标记的 Dies1 的免疫细胞化学成像显示其定位于脂肪细胞膜。用肿瘤坏死因子-α（TNFα）处理或通过 siRNA 敲低主前脂肪生成转录因子过氧化物酶体增殖物激活受体γ（PPARγ）来调节脂肪细胞表型，导致 Dies1 转录物水平分别降低 90%和 60%。此外，3T3-L1 前脂肪细胞中的 siRNA 介导的 Dies1 敲低抑制了脂肪生成转化。这种培养物的脂质含量减少了 35%，关键脂肪细胞转录物的表达减少了 45%-65%，包括 PPARγ。用于完全体外诱导分化的标准方案承诺前脂肪细胞，如 3T3-L1，不包括 BMP4 处理。因此，我们假设 Dies1 在脂肪生成中的积极作用，与它在胚胎干细胞分化中的作用不同，不包括其促 BMP4 作用。支持这一观点，敲低 Dies1 的 3T3-L1 脂肪细胞在暴露于 BMP4 时并未显示磷酸化 Smad1 水平降低。对多个小鼠和人类组织中的 Dies1 转录物进行 qPCR 分析显示，白色脂肪组织（WAT）高度富集。有趣的是，我们观察到禁食与进食小鼠的 WAT 中 Dies1 转录物的诱导增加了 10 倍，这表明 Dies1 在体内成熟脂肪细胞对营养的反应中起作用。总之，我们的数据将 Dies1 确定为一种新的分化依赖性脂肪细胞质膜蛋白，其表达是有效脂肪生成所必需的，并且可能在 WAT 的营养状态方面也发挥作用。
