Mitochondrial GTP Links Nutrient Sensing to β Cell Health, Mitochondrial Morphology, and Insulin Secretion Independent of OxPhos

Abstract

SUMMARY Mechanisms coordinating pancreatic β cell metabolism with insulin secretion are essential for glucose homeostasis. One key mechanism of β cell nutrient sensing uses the mitochondrial GTP (mtGTP) cycle. In this cycle, mtGTP synthesized by succinyl-CoA synthetase (SCS) is hydrolyzed via mitochondrial PEPCK (PEPCK-M) to make phosphoenolpyruvate, a high-energy metabolite that integrates TCA cycling and anaplerosis with glucose-stimulated insulin secretion (GSIS). Several strategies, including xenotopic overexpression of yeast mitochondrial GTP/GDP exchanger (GGC1) and human ATP and GTP-specific SCS isoforms, demonstrated the importance of the mtGTP cycle. These studies confirmed that mtGTP triggers and amplifies normal GSIS and rescues defects in GSIS both in vitro and in vivo. Increased mtGTP synthesis enhanced calcium oscillations during GSIS. mtGTP also augmented mitochondrial mass, increased insulin granule number, and membrane proximity without triggering de-differentiation or metabolic fragility. These data highlight the importance of the mtGTP signal in nutrient sensing, insulin secretion, mitochondrial maintenance, and β cell health.

Pyruvate kinase controls signal strength in the insulin secretory pathway

“…Paradoxically, PK recruitment occurs in parallel with the slowing of glycolytic flux. However, this recruitment occurs in the setting of continuous GK flux, when FBP levels rise and flux through PC increases, rerouting pyruvate flux through PEPCK-M (Jesinkey et al, 2019;Kibbey et al, 2007;Stark et al, 2009). Prior estimations of PEP cycling showed rates could reach ~40% of the total PK flux in islets (Stark et al, 2009).…”

“…Even as PK is sufficient to close KATP channels (Figure 1), the insulin secretion and calcium imaging data confirm that when glucose is the substrate then PK is dependent on glycolytic PEP that is under the metabolic control of GK (Figure 3). But importantly, PEP can arise from both the glycolytic enolase reaction as well as from anaplerosis contributing to PEPCK-M mediated cataplerosis known as the PEP cycle ( Figure 4A) (Jesinkey et al, 2019;Stark et al, 2009). To understand the mechanism by which PEP cycling interacts with PK, we utilized substrates that supply PK with PEP independently of glycolysis.…”

“…PEPCK-M is dependent on mitochondrial GTP (mtGTP) produced from the GTP-specific isoform of the TCA cycle enzyme succinyl CoA synthetase (SCS). Both the GTP isoform (SCS-GTP) and the ATP isoform (SCS-ATP) are catalytically dependent on a shared alpha subunit, so silencing or overexpressing one or the other isoform can modulate mtGTP synthesis and therefore PEPCK-M activity (Jesinkey et al, 2019;Kibbey et al, 2007). To determine whether PKa requires the PEP cycle to stimulate insulin secretion, we took advantage of INS1 832/13 cells stably overexpressing SCS-GTP or SCS-ATP (Jesinkey et al, 2019).…”

“…Glucose carbons that transit through PC generate 40% of cytosolic PEP through the cataplerotic mitochondrial PEP carboxykinase (PEPCK-M) reaction (Stark et al, 2009). This 'PEP cycle' has been linked to insulin secretion (Jesinkey et al, 2019;Stark et al, 2009) and provides a mechanism distinct from OxPhos for cytosolic ATP/ADP generation via pyruvate kinase (PK), which is allosterically activated by fructose 1,6-bisphosphate (FBP) prior to membrane depolarization (Merrins et al, 2013(Merrins et al, , 2016.…”

Endocrine-exocrine signaling drives obesity-associated pancreatic ductal adenocarcinoma

“…Islets from C57/B6 wild-type and ob/ob mice (n=4 mice per group) were isolated and perfused as previously described with a few minor changes, which are noted (Jesinkey et al, 2019). Isolated islets were recovered overnight and were perfused the following day in DMEM (D5030, Sigma) supplemented with 24mM NaHCO3, 10mM HEPES, 2.5mM glucose, 2mM glutamine and 0.2% BSA.…”