BETA2 activates transcription from the upstream glucokinase gene promoter in islet β-cells and gut endocrine cells

Glucokinase (GK) gene transcription initiates in the islet (beta-cell), gut, and brain from promoter sequences residing similar to35 kbp upstream from those used in liver. Expression of betaGK is controlled in beta-cells by cell-enriched (i.e. pancreatic duodenal homeobox 1 [PDX-1]) and ubiquitously (i.e., Pal) distributed factors that bind to and activate from conserved sequence motifs within the upstream promoter region (termed PGK). Here, we show that a conserved E-box element also contributes to control in the islet and gut. PGK promoter-driven reporter gene activity was diminished by mutating the specific sequences involved in E-box-mediated basic helix-loop-helix factor activator binding in islet beta-cells and enteroendocrine cells. Gel shift assays demonstrated that the PGK and insulin gene E-box elements formed the same cell-enriched (BETA2: E47) and generally distributed (upstream stimulatory factor [USF]) protein-DNA complexes. PGK E-box-driven activity was stimulated in cotransfection assays performed in baby hamster kidney (BHK) cells with BETA2 and E47, but not USF. Chromatin immunoprecipitation assays performed with BETA2 antisera showed that BETA2 occupies the upstream promoter region of the endogenous PGK gene in beta-cells. We propose that BETA2 (also termed NeuroD1) regulates PGK promoter activity.