Abstract
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.
Highlights
White adipose tissue (WAT) is the major organ of energy storage in vertebrates, where excess energy is present as triglyceride within adipocyte lipid droplets [1,2]
With the exception of WT-brown adipose tissue (BAT) described below, were purchased from The American Type Culture Collection (ATCC, Manassas VA). 293T cells were cultured in Dulbecco’s Modified Eagle Medium (DMEM) with 10% fetal calf serum (FCS). 3T3-L1 preadipocytes were grown in DMEM supplemented with 10% calf serum
Since our studies describe the role of this gene in differentiation, we will refer to it as Differentiation of Embryonic Stem Cells 1 (Dies1)
Summary
White adipose tissue (WAT) is the major organ of energy storage in vertebrates, where excess energy is present as triglyceride within adipocyte lipid droplets [1,2]. With insufficient ability to store triglyceride in WAT, either due to exceeding storage capacity as in obesity, or limited WAT storage capacity as occurs in some lipodystrophies, a lipotoxic and proinflammatory state is established [5]. Under these circumstances, free fatty acids can no longer be safely sequestered as triglyceride in the lipid droplet of white adipocytes [6]. Non-adipocytes are ill-equipped to handle excess lipid and lipoapoptosis and other detrimental responses ensue [5]
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