Depleting glioblastoma cells of very long-chain acyl-CoA synthetase 3 (ACSVL3) produces metabolic alterations in non-lipid pathways

Abstract

ABSTRACTKnockout (KO) of the fatty acid-activation enzyme very long-chain acyl-CoA synthetase 3 (ACSVL3; SLC27A3) in U87MG glioblastoma cells reduced their malignant growth properties both in vitro and in xenografts. These U87-KO glioma cells grew at a slower rate, became adherence-dependent, and were less invasive than parental U87 cells. U87-KO cells produced fewer, slower-growing subcutaneous and intracranial tumors when implanted in NOD-SCID mice. Thus, depleting U87MG cells of ACSVL3 restored these cells to a phenotype more like that of normal astrocytes. To understand the mechanisms underlying these beneficial changes, we investigated several possibilities, including the effects of ACSVL3 depletion on carbohydrate metabolism. Proteomic and metabolomic profiling indicated that ACSVL3 KO produced changes in glucose and energy metabolism. Even though protein levels of glucose transporters GLUT1 and GLUT3 were reduced by KO, cellular uptake of labeled 2-deoxyglucose was unaffected. Glucose oxidation to CO2was reduced nearly 7-fold by ACSVL3 depletion, and the cellular glucose level was 25% higher in KO cells. Glycolytic enzymes were upregulated by KO, but metabolic intermediates were essentially unchanged. Surprisingly, lactate production and the levels of lactate dehydrogenase isozymes LDHA and LDHB were elevated by ACSVL3 KO. The activity of the pentose phosphate pathway was found to be lower in KO cells. Citric acid cycle enzymes, electron transport chain complexes, and ATP synthase protein levels were all reduced by ACSVL3 depletion. Mitochondria were elongated in KO cells, but had a more punctate morphology in U87 cells. The mitochondrial potential was unaffected by lack of ACSVL3. We conclude that the beneficial effects of ACSVL3 depletion in human glioblastoma cells may result in part from alterations in diverse metabolic processes that are not directly related to role(s) of this enzyme in fatty acid and/or lipid metabolism. (Supported by NIH 5R01NS062043 and KKI institutional funds.)