Abstract
Pancreatic β-cell dysfunction contributes to onset and progression of type 2 diabetes. In this state β-cells become metabolically inflexible, losing the ability to select between carbohydrates and lipids as substrates for mitochondrial oxidation. These changes lead to β-cell dedifferentiation. We have proposed that FoxO proteins are activated through deacetylation-dependent nuclear translocation to forestall the progression of these abnormalities. However, how deacetylated FoxO exert their actions remains unclear. To address this question, we analyzed islet function in mice homozygous for knock-in alleles encoding deacetylated FoxO1 (6KR). Islets expressing 6KR mutant FoxO1 have enhanced insulin secretion in vivo and ex vivo and decreased fatty acid oxidation ex vivo Remarkably, the gene expression signature associated with FoxO1 deacetylation differs from wild type by only ∼2% of the >4000 genes regulated in response to re-feeding. But this narrow swath includes key genes required for β-cell identity, lipid metabolism, and mitochondrial fatty acid and solute transport. The data support the notion that deacetylated FoxO1 protects β-cell function by limiting mitochondrial lipid utilization and raise the possibility that inhibition of fatty acid oxidation in β-cells is beneficial to diabetes treatment.
Highlights
Three gene families were conspicuous by their extensive changes: 9 of the 22 members of the Rho guanine nucleotide exchange factors (Arhgefs) decreased, whereas 29 of the 41 subunits of the mitochondrial NADH dehydrogenase and 20 of the 53 members of the ribosomal protein L family (Rpl) increased
We were gratified to see that the top regulator identified in re-fed 6KR islets was the MODY gene Hnf1␣, which we have previously shown to be among key FoxO targets required for metabolic flexibility [10]
We found no difference in islets incubated in 2.8 mM glucose but a nearly 50% decrease in 6KR islets incubated in 16.8 mM glucose compared with WT (Fig. 2D)
Summary
Animals—We have described generation, characterization, and genotyping protocols for this mouse strain in a previous publication [12]. Each array was performed with pooled islets from three mice per genotype. For -cell morphometry, 4 pancreatic sections from 4 mice of each genotype were sampled 150 m apart and used for a two-tailed paired Student’s t test analysis. Glucose and Lipid Metabolism—After overnight recovery and 2-h starvation, 20 islets were preincubated in KRBH for 45 min at 2.8 mM glucose followed by incubation in KRBH containing 0.5 Ci of D-[5-3H]glucose (16 Ci/mmol) and 1Ci/ml D-[U-14C]glucose (250 mCi/mmol) at 2.8, 8.3, and 16.7 mM glucose. In experiments with purified islets, samples were incubated in either 5 mM or 20 mM glucose for 24 h before mRNA measurements. In some instances islets were transduced with adenovirus expressing GFP, FoxO1ADA, or FoxO16KR for 24 h [7] and incubated in 11 mM glucose before isolating mRNA. We used two-tailed Student’s t test, one-way ANOVA or two-way ANOVA for data analysis and the customary threshold of p Ͻ 0.05 to declare statistically significant differences
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