Glucose-6-phosphatase catalytic subunit 2 negatively regulates glucose oxidation and insulin secretion in pancreatic β-cells

Elevated fasting blood glucose (FBG) is associated with increased risks of developing type 2 diabetes (T2D) and cardiovascular-associated mortality. G6PC2 is predominantly expressed in islets, encodes a glucose-6-phosphatase catalytic subunit that converts glucose-6-phosphate (G6P) to glucose, and has been linked with variations in FBG in genome-wide association studies. Deletion of G6pc2 in mice has been shown to lower FBG without affecting fasting plasma insulin levels in vivo. At 5 mM glucose, pancreatic islets from G6pc2 knockout (KO) mice exhibit no glucose cycling, increased glycolytic flux, and enhanced glucose-stimulated insulin secretion (GSIS). However, the broader effects of G6pc2 KO on beta-cell metabolism and redox regulation are unknown. Here we used CRISPR/Cas9 gene editing and metabolic flux analysis in beta TC3 cells, a murine pancreatic beta-cell line, to examine the role of G6pc2 in regulating glycolytic and mitochondrial fluxes. We found that deletion of G6pc2 led to X60% increases in glycolytic and citric acid cycle (CAC) fluxes at both 5 and 11 mM glucose concentrations. Furthermore, intracellular insulin content and GSIS were enhanced by approximately two-fold, along with increased cytosolic redox potential and reductive carboxylation flux. Normalization of fluxes relative to net glucose uptake revealed upregulation in two NADPHproducing pathways in the CAC. These results demonstrate but also, independently, citric acid cycle activity in beta-cells. Overall, our findings implicate G6PC2 as a potential therawhich could benefit individuals with prediabetes, T2D, and obesity.