Silencing branched-chain ketoacid dehydrogenase or treatment with branched-chain ketoacids ex vivo inhibits muscle insulin signaling

Abstract

AbbreviationsAcadm, Acyl-Coenzyme A dehydrogenase, medium chain; BCAAs, branched-chain amino acids; BCKAs, branched-chain keto acids; BCATm, mitochondrial branched chain aminotransferase; BCKDH, branchedchain α-keto acid dehydrogenase; BCKDK, branched-chain keto acid dehydrogenase kinase; BT2, 3,6dichlorobenzo [b]thiophene-2-carboxylic acid; Cyclo, Cyclophilin; DIO, diet induced obesity; Glut, glucose transporter; Hadh, Hydroxyacyl-Coenzyme A dehydrogenase; HFD, high fat diet; HFHS, high fat high sucrose diet; Hmgcs1, 3-hydroxy-3-methylglutaryl-Coenzyme A synthase 1; IR, insulin resistance; Ivd2, Isovaleryl-CoA dehydrogenase; KIC, α-ketoisocaproic acid; KLF15, Kruppel like factor-15; mTOR, mammalian target of rapamycin; Mut, methylmalonyl-Coenzyme A mutase; PDH, pyruvate dehydrogenase; PPM1K, mitochondrial-targeted protein phosphatase; Rer1, Retention in endoplasmic reticulum sorting receptor 1; Rpl41, Ribosomal protein L41; Rpl7l1, Ribosomal protein L7-like 1; T2D, type 2 diabetes 3 Abstract Branched-chain α-keto acids (BCKAs) are downstream catabolites of branched-chain amino acids (BCAAs). Mitochondrial oxidation of BCKAs is catalyzed by branched-chain ketoacid dehydrogenase (BCKDH), an enzyme sensitive to inhibitory phosphorylation by BCKD kinase (BCKDK). Emerging studies show that defective BCAA catabolism and elevated BCKAs levels correlate with glucose intolerance and cardiac dysfunction. However, if/how BCKDH and BCKDK exert control on the availability and flux of intramyocellular BCKAs and if BCKA reprograms nutrient metabolism by influencing insulin action remains unexplored. We observed altered BCAA catabolizing enzyme expression in the murine heart and skeletal muscle during physiological fasting and diet-induced obesity and after ex vivo exposure of C2C12 cells to increasing concentration of saturated fatty acid, palmitate. BCKAs per se impaired insulin-induced AKT phosphorylation and AKT activity in skeletal myotubes and cardiomyocytes. In skeletal muscle cells, mTORC1 and protein translation signaling was enhanced by BCKA with concomitant suppression of mitochondrial respiration. Lowering intracellular BCKA levels by genetic and pharmacological activation of BCKDHA enhanced insulin signaling and activated pyruvate dehydrogenase, an effector of glucose oxidation and substrate metabolism. Our findings suggest that BCKAs profoundly influence muscle insulin function, providing new insight into the molecular nexus of BCAA metabolism and signaling with cellular insulin action and respiration.