Maintaining HNF6 expression prevents AdHNF3β-mediated decrease in hepatic levels of Glut-2 and glycogen

Abstract

The hepatocyte nuclear factor 3 (HNF-3) proteins are members of the Forkhead Box (Fox) family of transcription factors that play important roles in regulating expression of genes involved in cellular proliferation, differentiation, and metabolic homeostasis. In previous studies we increased liver expression of HNF-3β by using either transgenic mice (transthyretin HNF-3β) or recombinant adenovirus infection (AdHNF3β), and observed diminished hepatic levels of glycogen, and glucose transporter 2 (Glut-2), as well as the HNF-6, HNF-3, HNF-1α, HNF-4α, and C/EBPα transcription factors. We conducted the present study to determine whether maintaining HNF-6 protein expression during AdHNF3β infection prevents reduction of hepatic levels of glycogen and the earlier-mentioned genes. Here, we show that AdHNF3β- and AdHNF6-infected mouse liver displayed increased hepatic levels of glycogen, Glut-2, HNF-3γ, HNF-1α, and HNF-4α at 2 and 3 days postinfection (PI). Furthermore, restoration of hepatic glycogen levels after AdHNF3β and AdHNF6 coinfection was associated with increased Glut-2 expression. AdHNF6 infection alone caused a 2-fold increase in hepatic Glut-2 levels, suggesting that HNF 6 stimulates in vivo transcription of the Glut-2 gene. DNA binding assays showed that only recombinant HNF-6 protein, but not the HNF-3 proteins, binds to the mouse - 185 to - 144 bp Glut-2 promoter sequences. Cotransfection assays in human hepatoma (HepG2) cells with either HNF-3 or HNF-6 expression vectors show that only HNF-6 provided significant transcriptional activation of the Glut-2 promoter. In conclusion, these studies show that the hepatic Glut-2 promoter is a direct target for HNF-6 transcriptional activation.