Phosphoenolpyruvate Cycling via Mitochondrial Phosphoenolpyruvate Carboxykinase Links Anaplerosis and Mitochondrial GTP with Insulin Secretion

• Published in: The Journal of biological chemistry Vol. 284; no. 39; pp. 26578 - 26590
• Main Authors: Stark, Romana, Pasquel, Francisco, Turcu, Adina, Pongratz, Rebecca L., Roden, Michael, Cline, Gary W., Shulman, Gerald I., Kibbey, Richard G.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 25.09.2009; American Society for Biochemistry and Molecular Biology
• Subjects: 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; Metabolism and Bioenergetics; Mice; Mitochondria - enzymology; Mitochondria - metabolism; Models, Biological; Phosphoenolpyruvate - metabolism; Phosphoenolpyruvate Carboxylase - genetics; Phosphoenolpyruvate Carboxylase - metabolism; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; RNA Interference; Signal Transduction; Succinate-CoA Ligases - metabolism
• ISSN: 0021-9258; 1083-351X; 1083-351X
• DOI: 10.1074/jbc.M109.011775

Pancreatic β-cells couple the oxidation of glucose to the secretion of insulin. Apart from the canonical KATP-dependent glucose-stimulated insulin secretion (GSIS), there are important KATP-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 13C-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.