Phosphoenolpyruvate cycling via mitochondrial phosphoenolpyruvate carboxykinase links anaplerosis and mitochondrial GTP with insulin secretion

J Biol Chem. 2009 Sep 25;284(39):26578-90. doi: 10.1074/jbc.M109.011775. Epub 2009 Jul 27.

Abstract

Pancreatic beta-cells couple the oxidation of glucose to the secretion of insulin. Apart from the canonical K(ATP)-dependent glucose-stimulated insulin secretion (GSIS), there are important K(ATP)-independent mechanisms involving both anaplerosis and mitochondrial GTP (mtGTP). How mtGTP that is trapped within the mitochondrial matrix regulates the cytosolic calcium increases that drive GSIS remains a mystery. Here we have investigated whether the mitochondrial isoform of phosphoenolpyruvate carboxykinase (PEPCK-M) is the GTPase linking hydrolysis of mtGTP made by succinyl-CoA synthetase (SCS-GTP) to an anaplerotic pathway producing phosphoenolpyruvate (PEP). Although cytosolic PEPCK (PEPCK-C) is absent, PEPCK-M message and protein were detected in INS-1 832/13 cells, rat islets, and mouse islets. PEPCK enzymatic activity is half that of primary hepatocytes and is localized exclusively to the mitochondria. Novel (13)C-labeling strategies in INS-1 832/13 cells and islets measured substantial contribution of PEPCK-M to the synthesis of PEP. As high as 30% of PEP in INS-1 832/13 cells and 41% of PEP in rat islets came from PEPCK-M. The contribution of PEPCK-M to overall PEP synthesis more than tripled with glucose stimulation. Silencing the PEPCK-M gene completely inhibited GSIS underscoring its central role in mitochondrial metabolism-mediated insulin secretion. Given that mtGTP synthesized by SCS-GTP is an indicator of TCA flux that is crucial for GSIS, PEPCK-M is a strong candidate to link mtGTP synthesis with insulin release through anaplerotic PEP cycling.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Blotting, Western
  • Cell Line, Tumor
  • Cells, Cultured
  • Citric Acid Cycle
  • Guanosine Triphosphate / metabolism*
  • Insulin / metabolism*
  • Insulin Secretion
  • Insulinoma / enzymology
  • Insulinoma / metabolism
  • Insulinoma / pathology
  • Islets of Langerhans / cytology
  • Islets of Langerhans / enzymology
  • Islets of Langerhans / metabolism
  • Mice
  • Mitochondria / enzymology*
  • Mitochondria / metabolism
  • Models, Biological
  • Phosphoenolpyruvate / metabolism*
  • Phosphoenolpyruvate Carboxylase / genetics
  • Phosphoenolpyruvate Carboxylase / metabolism*
  • RNA Interference
  • Rats
  • Rats, Sprague-Dawley
  • Reverse Transcriptase Polymerase Chain Reaction
  • Signal Transduction
  • Succinate-CoA Ligases / metabolism

Substances

  • Insulin
  • Phosphoenolpyruvate
  • Guanosine Triphosphate
  • Phosphoenolpyruvate Carboxylase
  • Succinate-CoA Ligases