A novel ChREBP isoform in adipose tissue regulates systemic glucose metabolism

Abstract

The prevalence of obesity and type 2 diabetes is increasing worldwide and threatens to shorten lifespan. Impaired insulin action in peripheral tissues is a major pathogenic factor. Insulin stimulates glucose uptake in adipose tissue through the GLUT4 (also known as SLC2A4) glucose transporter, and alterations in adipose tissue GLUT4 expression or function regulate systemic insulin sensitivity. Downregulation of human and mouse adipose tissue GLUT4 occurs early in diabetes development. Here we report that adipose tissue GLUT4 regulates the expression of carbohydrate-responsive-element-binding protein (ChREBP; also known as MLXIPL), a transcriptional regulator of lipogenic and glycolytic genes. Furthermore, adipose ChREBP is a major determinant of adipose tissue fatty acid synthesis and systemic insulin sensitivity. We find a new mechanism for glucose regulation of ChREBP: glucose-mediated activation of the canonical ChREBP isoform (ChREBP-α) induces expression of a novel, potent isoform (ChREBP-β) that is transcribed from an alternative promoter. ChREBP-β expression in human adipose tissue predicts insulin sensitivity, indicating that it may be an effective target for treating diabetes.

Figure 1. Genetically altering adipose tissue glucose flux regulates the expression of ChREBP and its lipogenic targets

a, mRNA expression of fatty acid synthetic enzymes, and b, lipogenic transcription factors in perigonadal fat from 6-week-old female mice (n=10–14 per group). *P<0.05 compared to respective controls. c, ChREBP and Glut4 mRNA correlate highly in PG WAT from control and AG4KO mice (n=27). Values are means ± S.E.

Figure 2. ChREBP is essential for the effects of adipose tissue Glut4 on adiposity, DNL, and glucose homeostasis

a, DNL measured in vivo in fed, 4-month-old male mice PG=perigonadal, SC=subcutaneous, (n=5–6 per group). b, Quantification of western blots of FAS and ACC in SC fat from fed, 6-month-old females (n=9–10 per group). For a and b, *P<0.05 versus same ChREBP genotype, different AG4OX genotype. # p<0.05 versus same AG4OX genotype, different ChREBP genotype. c, Body weights in male mice on chow (n=5–7 per group). *P<0.05 versus all other groups at the indicated time. d, Body composition in 8-week-old, female mice (for d–g; n=10–12 per group). *P<0.01 versus all others. e, Glucose tolerance test and f, insulin tolerance test. *P<0.05 versus all others; #P<0.05 versus WT and ChREBP KO. g, Glycemia following food removal * P<0.05 versus all others; # P<0.05 versus ChREBP KO and AG4OX-ChREBP KO; †P<0.05 versus AG4OX; Values are means ± S.E.

Figure 3. ChREBP is regulated in mouse and human adipose tissue in pathologic conditions

a, Body weight in chow- versus HFD-WT and AG4OX mice (9-week-old, n=8 per group), *P<0.05 WT-Chow compared to all others at the same time point. b, Glucose tolerance test in 7-week-old males (n=8 per group). *P<0.001 versus all others; c, Fatty acid synthesis measured in vivo in 5–6 month-old males (n=7 per group). Last panel, plasma insulin: HFD increases insulin (†P=0.01 by 2-way ANOVA). mRNA expression of d, ChREBP and SREBP-1c and e, ACC and FAS in SC fat from 4-month-old male mice (n=8–14 per group). For panels c–e, *P<0.05 compared to same diet, different genotype. #P<0.05 compared to same genotype, different diet. f, SC fat ChREBP mRNA correlates with insulin sensitivity in non-diabetic, normal glucose-tolerant humans (n=123). g, ChREBP and Glut4 mRNA expression in SC fat correlate in this group. h, SC fat ChREBP mRNA correlates highly with insulin sensitivity in obese, non-diabetic, BMI-matched humans (n=38). i, ChREBP and Glut4 mRNA expression do not correlate in this group. Values are means ± S.E.

Figure 4. Expression of the novel ChREBPβ isoform is regulated in a glucose- and ChREBP-dependent manner

a, Model of ChREBPα and ChREBPβ gene structure with indication of splice sites and translational start sites (ATG). b, Regulation of ChREBPα and β mRNA expression in perigonadal fat and liver of 10-week-old, female mice with fasting and refeeding (n=6/group). *P<0.05 compared to fed group. c, ChREBPα and β mRNA expression in perigonadal fat from 6-week-old female AG4OX and AG4KO compared to littermate controls (n=10–14 per group). *P<0.05 versus respective control. d, Glucose regulation of exon 1b promoter-luciferase reporter and indicated ChREBPβ mutants, co-transfected with ChREBPα and Mlx (n=3/group). *P<0.05 compared to non-mutated exon 1b-luciferase construct in the same glucose. #P<0.05 compared to the same construct in low glucose. e, ChREBPα and β induce an ACC ChoRE-luciferase reporter compared to pGL3_basic control vector in both low and high glucose. *P<0.05 compared to ChREBPα in the same glucose, #P<0.05 compared to ChREBPα, low glucose. Values are means ± S.E.

Figure 5. Adipose tissue ChREBPβ expression predicts insulin sensitivity

a, ChREBPα and β mRNA expression in SC fat of 4-month-old male mice on chow or HFD (n=10–14 per group). *P<0.05 compared to Chow-fed. b, mRNA expression of ChREBPβ in SC fat correlates more highly with insulin sensitivity (% increase in insulin-stimulated glucose uptake over basal measured during a euglycemic-hyperinsulinemic clamp procedure) than ChREBPα in obese, non-diabetic, BMI-matched humans (n=38). Values are means ± S.E.

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