Phosphorylation of CCAAT/enhancer-binding protein α regulates GLUT4 expression and glucose transport in adipocytes

The transcription factor CCAAT/enhancer-binding protein alpha(C/EBP alpha) is required during adipogenesis for development of insulin-stimulated glucose uptake. Modes for regulating this function of C/EBP alpha have yet to be determined. Phosphorylation of C/EBP alpha on Ser-21 has been implicated in the regulation of granulopoiesis and hepatic gene expression. To explore the role of Ser-21 phosphorylation on C/EBP alpha function during adipogenesis, we developed constructs in which Ser-21 was mutated to alanine (S21A) to model dephosphorylation. In two cell culture models deficient in endogenous C/EBP alpha, enforced expression of S21A-C/EBP alpha resulted in normal lipid accumulation and expression of many adipogenic markers. However, S21A-C/EBP alpha had impaired ability to activate the Glut4 promoter specifically, and S21A-C/EBP alpha expression resulted in diminished GLUT4 and adiponectin expression, as well as reduced insulin-stimulated glucose uptake. No defects in insulin signaling or GLUT4 vesicle trafficking were identified with S21A-C/EBP alpha expression, and when exogenous GLUT4 expression was enforced to normalize expression in S21A-C/EBP alpha cells, insulin-responsive glucose transport was reconstituted, suggesting that the primary defect was a deficit in GLUT4 levels. Mice in which endogenous C/EBP alpha was replaced with S21A-C/EBP alpha displayed reduced GLUT4 and adiponectin protein expression in epididymal adipose tissue and increased blood glucose compared with wild-type littermates. These results suggest that phosphorylation of C/EBP alpha on Ser-21 may regulate adipocyte gene expression and whole body glucose homeostasis.