Insulin-producing β cells become dedifferentiated during diabetes progression. An impaired ability to select substrates for oxidative phosphorylation, or metabolic inflexibility, initiates progression from β-cell dysfunction to β-cell dedifferentiation. The identification of pathways involved in dedifferentiation may provide clues to its reversal. Here we isolate and functionally characterize failing β cells from various experimental models of diabetes and report a striking enrichment in the expression of aldehyde dehydrogenase 1 isoform A3 (ALDH+) as β cells become dedifferentiated. Flow-sorted ALDH+ islet cells demonstrate impaired glucose-induced insulin secretion, are depleted of Foxo1 and MafA, and include a Neurogenin3-positive subset. RNA sequencing analysis demonstrates that ALDH+ cells are characterized by: (i) impaired oxidative phosphorylation and mitochondrial complex I, IV and V; (ii) activated RICTOR; and (iii) progenitor cell markers. We propose that impaired mitochondrial function marks the progression from metabolic inflexibility to dedifferentiation in the natural history of β-cell failure.

Pancreatic ␤-cell dysfunction contributes to onset and progression of type 2 diabetes. In this state ␤-cells become metabolically inflexible, losing the ability to select between carbohydrates and lipids as substrates for mitochondrial oxidation. These changes lead to ␤-cell dedifferentiation. We have proposed that FoxO proteins are activated through deacetylationdependent 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 ϳ2% of the >4000 genes regulated in response to re-feeding. But this narrow swath includes key genes required for ␤-cell identity, lipid metabolism, and mitochondrial fatty acid and solute transport. The data support the notion that deacetylated FoxO1 protects ␤-cell function by limiting mitochondrial lipid utilization and raise the possibility that inhibition of fatty acid oxidation in ␤-cells is beneficial to diabetes treatment.