Fig. 1: ChREBPβ is the main nuclear isoform after prolonged exposure to high concentrations of glucose.

a, b Ins-1 cells were cultured in low (2 mM) or high (20 mM) glucose for 24 h. Cells were fixed and immunostained with the C-terminus antibody for ChREBP, which recognizes both ChREBPα and ChREBPβ (a), or the N-terminus antibody for ChREBP, specific for ChREBPα (b). The results are representative of three independent experiments. c, d Islets from floxed ChREBPβ mice were isolated and transduced with adenoviruses expressing LacZ as a control, or Cre recombinase to excise ChREBPβ and cultured in 6 mM or 20 mM glucose. After 48 h, cells were fixed and immunostained with the N-terminal (c) or C-terminal (d) antibody; nuclei were stained with DAPI. Shown is a representative result of 3 independent experiments. e, f Ins-1 cells were transduced with an adenovirus expressing flag-tagged ChREBPα and treated with 2 mM (e) or 20 mM glucose (f) for the indicated times. Cells were fixed and immunostained with an anti-Flag antibody and nuclei were stained with DAPI. The results from (f) were quantified in (g). The results are representative of three independent experiments, *, mean, P < 0.05. h, i Ins-1 cells were cultured overnight in 2 mM glucose and then glucose was added to a total of 20 mM for the indicated times; chromatin was isolated and processed for chromatin immunoprecipitation using antibodies recognizing the C-terminus, the N-terminus, or IgG. DNA was amplified by qPCR using primers specific for regions near the ChoREs on the Pklr (h), or Mlxipl (ChREBPβ) (i) gene promoters. The data are the means ± SEM of the percent input after subtraction of the IgG control. n = 4; *p < 0.05; ***P < 0.0005; ****P < 0.0001 using two-way ANOVA.