Nutrient-dependent regulation of β-cell KATP channels is controlled by the isoforms of pyruvate kinase and the source of phosphoenolpyruvate
Pyruvate kinase (PK) and the phosphoenolpyruvate (PEP) cycle play key roles in nutrient-stimulated KATP channel closure and insulin secretion. To identify the PK isoforms involved, we generated mice lacking {beta}-cell PKm1, PKm2, and mitochondrial PEP carboxykinase (PCK2) that generates mitochondrial PEP. Glucose metabolism generates both glycolytic and mitochondrially-derived PEP, which triggers KATP closure through local PKm1 and PKm2 signaling at the plasma membrane. Amino acids, which generate mitochondrial PEP without producing glycolytic fructose 1,6-bisphosphate to allosterically activate PKm2, signal through PKm1 to raise ATP/ADP, close KATP channels, and stimulate insulin secretion. Raising cytosolic ATP/ADP with amino acids is insufficient to close KATP channels in the absence of PK activity or PCK2, indicating that KATP channels are regulated by mitochondrially-derived PEP that provides ATP via plasma membrane-associated PK, but not via mitochondrially-derived ATP. Following membrane depolarization, the PEP cycle is also involved in an "off-switch" that facilitates KATP channel reopening and Ca2+ extrusion, as shown by PK activation experiments and {beta}-cell PCK2 deletion that prolonged Ca2+ oscillations and increased insulin secretion. In conclusion, the differential response of PKm1 and PKm2 to the glycolytic and mitochondrial sources of PEP influences the {beta}-cell nutrient response, and controls the oscillatory cycle regulating insulin secretion.