The isoforms of pyruvate kinase act as nutrient sensors for the β-cell K_ATP channel

Abstract

As nutrient sensors for the organism, pancreatic β-cells use metabolism as a signaling pathway to elicit insulin secretion. Phosphoenolpyruvate (PEP), the substrate for pyruvate kinase (PK), may be a critical signaling intermediate based on its ability to locally control ATP-sensitive K+ (KATP) channels on the plasma membrane. Using isoform-specific deletion, we show that constitutively-active PKm1 is sufficient for KATP closure. Yet, it is the minor but allosterically-tunable PKm2 isoform that enables glucose-dependent regulation of the KATP channel microcompartment. In contrast to glucose, PKm1 and PKm2 have non-overlapping responses to amino acids, which generate PEP via the mitochondrial PCK2 enzyme. β-cell deletion of PCK2 blocked amino acid regulation of KATP and impacted Ca2+ influx (via PKm1) and extrusion (via PKm2). Shifting the β-cell from PKm2 to PKm1 correspondingly increased secretory output. Together, these three knockout models indicate that the source of PEP and the isoforms of PK are key determinants of the β-cell nutrient response.

β Cell–specific deletion of Zfp148 improves nutrient-stimulated β cell Ca2+ responses

“…RNA-Seq of islets from β-Zfp148 KO and control mice identified numerous genes that control nutrient responses. We demonstrated that 1 or more of these pathways converge to enhance PEP cycling and, consequently, pyruvate kinase-derived ATP production (Figures 2 and 5, and Supplemental Figures 2 and 3), the direct activator of K ATP channel closure and a known regulator of insulin secretion (22,37).…”

“…Glutamate dehydrogenase (GDH) plays a key role in amino acid-induced stimulation of insulin secretion (36). β Cells express high levels of mitochondrial phosphoenolpyruvate carboxykinase (Pck2), and deletion of this gene dramatically reduces insulin secretion and blocks amino acid-stimulated Ca 2+ influx at low glucose levels (23,37). Activators of pyruvate kinases stimulate insulin secretion (22).…”

“…22). Thus, with a restricted glycolytic PEP supply, the PEP substrate used by pyruvate kinase to generate the ATP that closes the ATP-sensitive K + (K ATP ) channel derives predominantly, if not exclusively, from mitochondrial PEP production via PCK2 (23,37).…”

“…The role of GDH in this pathway was revealed through the discovery that people with mutations resulting in constitutively active GDH have hyperinsulinemia and hyperammonemia (36,59). Together with subsequent work demonstrating a critical role for mitochondrial PEPCK (encoded by Pck2) in amino acid-stimulated insulin secretion, the formation of PEP and its cycling via PK compose the pathway mediating amino acid-stimulated insulin secretion (22,23,37). The enhanced sensitivity to amino acid-induced Ca 2+ elevation we observed in β-Zfp148 KO islets is consistent with elevated PEP cycling.…”

“…Enzymes involved in metabolism of serine and folate (SHMT2, ALDH1L2, MTHFD2, and FOLR1; Supplemental Figure 7) may alter TCA cycling through production of redox intermediates (72,73). Finally, elevated mitochondrial PEPCK (PCK2) increases conversion of OAA to PEP (22,23,37,74). Collectively, these enzyme changes support a model whereby the loss of Zfp148 results in increased mitochondrial PEP production and flux through pyruvate kinase, with increased ATP, resulting in more rapid closure of K ATP channels and enhanced insulin secretion.…”