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VARS2 Depletion Leads to Activation of the Integrated Stress Response and Disruptions in Mitochondrial Fatty Acid Oxidation.

ABSTRACT: Mutations in mitochondrial aminoacyl-tRNA synthetases (mtARSs) have been reported in patients with mitochondriopathies: most commonly encephalopathy, but also cardiomyopathy. Through a GWAS, we showed possible associations between mitochondrial valyl-tRNA synthetase (VARS2) dysregulations and non-ischemic cardiomyopathy. We aimed to investigate the possible consequences of VARS2 depletion in zebrafish and cultured HEK293A cells. Transient VARS2 loss-of-function was induced in zebrafish embryos using Morpholinos. The enzymatic activity of VARS2 was measured in VARS2-depleted cells via northern blot. Heterozygous VARS2 knockout was established in HEK293A cells using CRISPR/Cas9 technology. BN-PAGE and SDS-PAGE were used to investigate electron transport chain (ETC) complexes, and the oxygen consumption rate and extracellular acidification rate were measured using a Seahorse XFe96 Analyzer. The activation of the integrated stress response (ISR) and possible disruptions in mitochondrial fatty acid oxidation (FAO) were explored using RT-qPCR and western blot. Zebrafish embryos with transient VARS2 loss-of-function showed features of heart failure as well as indications of CNS and skeletal muscle involvements. The enzymatic activity of VARS2 was significantly reduced in VARS2-depleted cells. Heterozygous VARS2-knockout cells showed a rearrangement of ETC complexes in favor of complexes III₂, III₂ + IV, and supercomplexes without significant respiratory chain deficiencies. These cells also showed the enhanced activation of the ISR, as indicated by increased eIF-2α phosphorylation and a significant increase in the transcript levels of ATF4, ATF5, and DDIT3 (CHOP), as well as disruptions in FAO. The activation of the ISR and disruptions in mitochondrial FAO may underlie the adaptive changes in VARS2-depleted cells.

SUBMITTER: Kayvanpour E

PROVIDER: S-EPMC9267100 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:AimsCardiomyopathy and arrhythmias can be severe presentations in patients with inherited defects of mitochondrial long-chain fatty acid β-oxidation (FAO). The pathophysiological mechanisms that underlie these cardiac abnormalities remain largely unknown. We investigated the molecular adaptations to a FAO deficiency in the heart using the long-chain acyl-CoA dehydrogenase (LCAD) knockout (KO) mouse model.Methods and resultsWe observed enrichment of amino acid metabolic pathways and of ATF4 target genes among the upregulated genes in the LCAD KO heart transcriptome. We also found a prominent activation of the eIF2α/ATF4 axis at the protein level that was independent of the feeding status, in addition to a reduction of cardiac protein synthesis during a short period of food withdrawal. These findings are consistent with an activation of the integrated stress response (ISR) in the LCAD KO mouse heart. Notably, charging of several transfer RNAs (tRNAs), such as tRNAGln was decreased in LCAD KO hearts, reflecting a reduced availability of cardiac amino acids, in particular, glutamine. We replicated the activation of the ISR in the hearts of mice with muscle-specific deletion of carnitine palmitoyltransferase 2.ConclusionsOur results show that perturbations in amino acid metabolism caused by long-chain FAO deficiency impact cardiac metabolic signalling, in particular the ISR. These results may serve as a foundation for investigating the role of the ISR in the cardiac pathology associated with long-chain FAO defects.Translational Perspective: The heart relies mainly on mitochondrial fatty acid β-oxidation (FAO) for its high energy requirements. The heart disease observed in patients with a genetic defect in this pathway highlights the importance of FAO for cardiac health. We show that the consequences of a FAO defect extend beyond cardiac energy homeostasis and include amino acid metabolism and associated signalling pathways such as the integrated stress response.

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VARS2 Depletion Leads to Activation of the Integrated Stress Response and Disruptions in Mitochondrial Fatty Acid Oxidation.

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