The hotdog-fold acyl-CoA thioesterases (ACOTs) are enzymes typically related to lipid metabolism and are widely spread across the tree of life, but many members of this group are poorly characterized. In this work, through in silico analyzes we identified the MICOS-associated mitochondrial proteins of unknown function Mrx3 and Fmp10 as two putative ACOTs in Saccharomyces cerevisiae with structural homology to the mammalian thioesterases Them4 and Them5, which possess a conserved motif that is catalytic toward thioester hydrolysis. Lipidomics analyses revealed that the mrx3Δ and fmp10Δ mutants have lower free fatty acid (FFA) levels with concomitant accumulation of storage lipids, besides a decreased pool of cardiolipin species. During our functional investigation, we observed that the absence of either protein partially restores respiratory capacity in the leu5Δ strain, whose mutation has been previously linked to very slow respiratory growth due to severe Coenzyme A (CoA) depletion in the mitochondrial matrix. We verified that the respiratory defect of the leu5Δ mutant is also related to a defect in the respiratory chain given the low NADH-dependent oxygen consumption rate and is specifically linked to defective Complex IV activity. The strain's growth defect was exacerbated by deletion of the glycine transporter HEM25 and was ameliorated by either δ-aminolevulinic acid (ALA, a heme precursor) or iron supplementation. These results may indicate that the leu5Δ strain has a subtle heme deficiency. We propose a model in which Mrx3 and Fmp10 control the mitochondrial acyl-CoA/CoA ratio through a thioesterase activity, which finely modulates the mitochondrial membrane lipidome, with a concomitant effect in heme biosynthesis. In the leu5Δ mutant, there is a disturbed metabolic homeostasis. Deleting these hotdog-fold proteins cause increased acyl-CoA levels and partially restores metabolic balance, potentially by repartitioning carbon flux through the tricarboxylic acid (TCA) cycle to replenish succinyl-CoA using the limited available CoA pool, which may be sufficient to restore heme biosynthesis. This mechanism highlights the critical role of CoA compartmentalization and the underlying connection of distinct metabolic pathways.
Costa-Lima et al. (Wed,) studied this question.