Bekeova_2023_bioRxiv__

Acyl-Coenzyme A (acyl-CoA) thioesters are compartmentalized intermediates that participate in in multiple metabolic reactions within the mitochondrial matrix. The limited availability of free CoA (CoASH) in the matrix raises the question of how the local acyl-CoA concentration is regulated to prevent trapping of CoASH from overload of any specific substrate. Acyl-CoA thioesterase-2 (ACOT2) hydrolyzes long-chain acyl-CoAs to their constituent fatty acids and CoASH, and is the only mitochondrial matrix ACOT refractory to inhibition by CoASH. Thus, we reasoned that ACOT2 may constitutively regulate matrix acyl-CoA levels. Acot2 deletion in murine skeletal muscle (SM) resulted in acyl-CoA build-up when lipid supply and energy demands were modest. When energy demand and pyruvate availability were elevated, lack of ACOT2 activity promoted glucose oxidation. This preference for glucose over fatty acid oxidation was recapitulated in C2C12 myotubes with acute depletion of Acot2 , and overt inhibition of beta-oxidation was demonstrated in isolated mitochondria from Acot2 -depleted glycolytic SM. In mice fed a high fat diet, ACOT2 enabled the accretion of acyl-CoAs and ceramide derivatives in glycolytic SM, and this was associated with worse glucose homeostasis compared to when ACOT2 was absent. These observations suggest that ACOT2 supports CoASH availability to facilitate beta-oxidation in glycolytic SM when lipid supply is modest. However, when lipid supply is high, ACOT2 enables acyl-CoA and lipid accumulation, CoASH sequestration, and poor glucose homeostasis. Thus, ACOT2 regulates matrix acyl-CoA concentration in glycolytic muscle, and its impact depends on lipid supply.