Molecular targets of a human HNF1alpha mutation responsible for pancreatic beta-cell dysfunction

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Abstract
The reverse tetracycline‐dependent transactivator system was employed in insulinoma INS‐1 cells to achieve controlled inducible expression of hepatocyte nuclear factor‐1α (HNF1α)‐P291fsinsC, the most common mutation associated with subtype 3 of maturity‐onset diabetes of the young (MODY3). Nuclear localized HNF1α‐P291fsinsC protein exerts its dominant‐negative effects by competing with endogenous HNF1α for the cognate DNA‐binding site. HNF1α controls multiple genes implicated in pancreatic β‐cell function and notably in metabolism– secretion coupling. In addition to reduced expression of the genes encoding insulin, glucose transporter‐2, l‐pyruvate kinase, aldolase B and 3‐hydroxy‐3‐methylglutaryl coenzyme A reductase, induction of HNF1α‐P291fsinsC also significantly inhibits expression of mitochondrial 2‐oxoglutarate dehydrogenase (OGDH) E1 subunit mRNA and protein. OGDH enzyme activity and [14C]pyruvate oxidation were also reduced. In contrast, the mRNA and protein levels of mitochondrial uncoupling protein‐2 were dramatically increased by HNF1α‐P291fsinsC induction. As predicted from this altered gene expression profile, HNF1α‐P291fsinsC also inhibits insulin secretory responses to glucose and leucine, correlated with impaired nutrient‐evoked mitochondrial ATP production and mitochondrial membrane hyperpolarization. These unprecedented results suggest the molecular mechanism of HNF1α‐P291fsinsC causing β‐cell dysfunction.