O-GlcNAcylation Increases ChREBP Protein Content and Transcriptional Activity in the Liver

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FIG. 1.
FIG. 1.

ChREBP interacts with OGT and is O-GlcNAcylated in a dynamic manner in HEK293T cells and in mouse hepatocytes. A: Schematic diagram of the UDP-GlcNAc pathway. O-GlcNAcylation is an important posttranslational modification that has been proposed to be a nutrient sensor because the donor sugar, UDP-GlcNAc, receives input from multiple metabolic pathways. Activity of OGT, the enzyme that transfers the monosaccharide to serine/threonine residues, depends upon UDP-GlcNAc concentrations. Of glucose that enters the cells, 2–5% is used for production of the donor sugar nucleotide. The addition of GlcNH2 directly enters the hexosamine biosynthetic pathway. PUGNAc is an inhibitor of the OGA that hydrolyses the sugar. B: HEK239T cells were cotransfected with OGT (1 μg) and ChREBPwt plasmids (1 μg) and incubated for 24 h under high glucose conditions (G25). Immunoprecipitation (IP) of ChREBP was analyzed by immunoblotting with an OGT antibody. β-Actin was used as loading control. Representative Western blots (Wb) are shown. n = 3 independent experiments. C: ChREBP-overexpressing HEK293T cells were incubated under low glucose medium plus 5 mM GlcNH2 or transfected with an OGT-expressing vector or both. Representative Western blots are shown. n = 3 independent experiments. D: Overexpression of OGA (1 μg) in HEK293T cells cultured in 25 mM glucose leads to a decrease in global and transfected ChREBPOG. Representative Western blots are shown. n = 3 independent experiments. E: Immunoprecipitated ChREBP from primary mouse hepatocytes incubated for 24 h in either low glucose (G5) or high glucose plus insulin (G25i) was immunoblotted with an OGT antibody in order to evaluate the interaction between the two endogenous proteins. β-Actin was used as a loading control. Representative Western blots are shown. n = 3 independent experiments. (A high-quality color representation of this figure is available in the online issue.)

This Article

  1. Diabetes vol. 60 no. 5 1399-1413