Suppression of Glutamine:Fructose-6-phosphate amidotransferase-1 inhibits adipogenesis in 3T3-L1 adipocytes

O‐linked N‐acetylglucosamine (O‐GlcNAc) protein modification has been implicated in the regulation of signaling pathways, cell function, and gene expression. Glutamine:fructose‐6‐phosphate amidotransferase‐1 (GFAT‐1) is the rate‐limiting enzyme in the hexosamine biosynthetic pathway (HBP), which generates the sugar nucleotide UDP‐GlcNAc, where this nucleotide acts as the donor for O‐GlcNAc modification. In this study, we determined whether GFAT‐1 regulates adipogenesis in adipocytes. 3T3‐L1 preadipocytes were differentiated using medium containing high glucose, insulin, dexamethasone, and isobutylmethylxanthine. Cells were harvested 4, 8, and 12 h and 1, 2, 3, 4, 6, and 8 days after the initiation of differentiation. Global level of O‐GlcNAc modification increased 4 h after induction and persisted for 8 days of observation. GFAT‐1 mRNA and protein expression was also upregulated beginning 4 h after induction. Pharmacological inhibition of GFAT‐1 or GFAT‐1 siRNA treatment blocked the increase in O‐GlcNAcylation and the formation of lipid droplets in adipocytes. GFAT‐1 may regulate the expression of C/EBPβ, PPARγ, SREBP‐1, fatty acid synthase, S3‐12, perilipin, or adipophilin during adipogenesis. Our results suggest that GFAT‐1 plays a critical role in modulating adipogenesis via the regulation of protein O‐GlcNAcylation in adipocytes. J. Cell. Physiol. 227: 108–115, 2012. © 2011 Wiley Periodicals, Inc.