Acat1 (CG10932) is the single Drosophila melanogaster ortholog of human mitochondrial acetoacetyl-CoA thiolase (ACAT1 / "T2"; EC 2.3.1.9), a member of the thiolase family with the canonical Cys-His-Cys catalytic triad. It is a mitochondrial, CoA-dependent thiolase that catalyzes the reversible Claisen condensation/thiolysis at the acetoacetyl-CoA node (2 acetyl-CoA <-> acetoacetyl-CoA + CoA), placing it centrally in ketone body metabolism (both ketolysis and ketogenesis). By analogy to human T2 it also cleaves the 2-methyl-branched intermediate 2-methylacetoacetyl-CoA, the final thiolytic step of L-isoleucine catabolism. In vivo, Acat1 is required in adult mushroom body neurons for the mitochondrial oxidation of imported ketone bodies to acetyl-CoA, supplying energy that sustains associative memory during starvation. Despite the shared historical nickname "ACAT1", this mitochondrial thiolase is distinct from the endoplasmic-reticulum cholesterol-esterifying sterol O-acyltransferase (SOAT1/SOAT2, EC 2.3.1.26, also called "ACAT"); Acat1 has no cholesterol O-acyltransferase activity.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0003985 acetyl-CoA C-acetyltransferase activity | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic (IBA) assignment of the acetyl-CoA C-acetyltransferase (acetoacetyl-CoA thiolase, EC 2.3.1.9) activity across the ACAT1/T2 orthology group. This is the core catalytic function of Acat1, concordant with the thiolase-family domain architecture and the DIOPT 15/15 orthology to human ACAT1. Reason: Core molecular function, supported by IBA, EC-based IEA, ISS from human T2, and the canonical thiolase catalytic triad (Cys109/His368/Cys396) in the UniProt record. Supporting Evidence: PMID:35177854 the orthologue of the human mitochondrial acetoacetyl-CoA thiolase ACAT1, CG10932 |
| GO:0003985 acetyl-CoA C-acetyltransferase activity | IEA GO_REF:0000003 | ACCEPT | Summary: Electronic assignment of the core acetyl-CoA C-acetyltransferase activity via EC 2.3.1.9 mapping. Redundant with the IBA and ISS annotations but correct. Reason: Correctly captures the core catalytic function; EC 2.3.1.9 is asserted in the UniProt record. |
| GO:0003985 acetyl-CoA C-acetyltransferase activity | ISS GO_REF:0000024 | ACCEPT | Summary: Curator ISS transfer of the acetyl-CoA C-acetyltransferase activity from the experimentally characterized human ortholog ACAT1 (UniProtKB:P24752). Concordant with the IBA/IEA annotations and the shared thiolase catalytic machinery. Reason: Core molecular function; ortholog transfer from human T2 is appropriate given the 15/15 DIOPT orthology and conserved catalytic residues. Supporting Evidence: PMID:35177854 the orthologue of the human mitochondrial acetoacetyl-CoA thiolase ACAT1, CG10932 |
| GO:0016453 C-acetyltransferase activity | ISS GO_REF:0000024 | KEEP AS NON CORE | Summary: ISS annotation to the more general parent term C-acetyltransferase activity. Correct but less informative than the specific child GO:0003985. Reason: Correct parent term; the specific acetyl-CoA C-acetyltransferase activity (GO:0003985) is the preferred core molecular-function annotation. |
| GO:0016746 acyltransferase activity | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: InterPro-based assignment of the broad acyltransferase-activity parent term from the thiolase domain signatures. Reason: Technically correct but uninformative; superseded by the specific thiolase term GO:0003985. |
| GO:0016747 acyltransferase activity, transferring groups other than amino-acyl groups | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: InterPro-based assignment of an intermediate acyltransferase parent term from the thiolase InterPro signatures. Reason: Correct parent term; the specific acetyl-CoA C-acetyltransferase activity better captures the molecular function. |
| GO:0005739 mitochondrion | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic localization of Acat1 to the mitochondrion across the ACAT1/T2 orthology group. Concordant with the direct organelle-proteomics evidence and with the mitochondrial-matrix localization of human T2. Reason: Core cellular localization; the mitochondrion is the compartment where this thiolase acts, and the fly experimental study describes Acat1 generating acetyl-CoA "in the mitochondria". Supporting Evidence: PMID:35177854 KBs are used by ACAT1 to generate acetyl-CoA in the mitochondria for energy |
| GO:0005739 mitochondrion | HDA PMID:19317464 Mapping organelle proteins and protein complexes in Drosophi... | ACCEPT | Summary: High-throughput organelle-proteomics (LOPIT) mapping of Drosophila embryo proteins assigns Acat1 to the mitochondrion. Direct mass-spectrometry-based localization consistent with the IBA annotation and the mitochondrial identity of the ACAT1/T2 subfamily. Reason: Core mitochondrial localization supported by an experimental (HDA) organelle-mapping dataset. Supporting Evidence: PMID:19317464 we apply LOPIT, a mass-spectrometry based technique that simultaneously maps proteins to specific subcellular compartments, to Drosophila embryos |
| GO:0005739 mitochondrion | ISM PMID:22758915 An inventory of peroxisomal proteins and pathways in Drosoph... | ACCEPT | Summary: Sequence-model (ISM) mitochondrial prediction. Redundant with the direct HDA and phylogenetic mitochondrial annotations and consistent with them; the mitochondrion is the core localization of this thiolase. Reason: Correct core compartment, agreeing with the HDA (LOPIT) and IBA mitochondrial annotations for the same term. |
| GO:0005777 peroxisome | ISM PMID:22758915 An inventory of peroxisomal proteins and pathways in Drosoph... | MARK AS OVER ANNOTATED | Summary: Sequence-model (ISM) peroxisomal prediction from a Drosophila peroxisomal-proteome inventory. This conflicts with the mitochondrial identity of the ACAT1/T2 subfamily (human T2 is a mitochondrial-matrix protein) and with the direct LOPIT and phylogenetic mitochondrial annotations for Acat1. The cited paper is abstract-only and states only that "five of these predicted peroxisomal proteins" had their localization confirmed, without indicating whether Acat1 was among them, so the prediction cannot be verified and is contradicted by stronger mitochondrial evidence. Reason: Computational targeting-signal prediction with no verification for Acat1 and contradicted by the mitochondrial localization of the ACAT1/T2 thiolase subfamily; peroxisomal thiolase activity in flies belongs to distinct ACAA1-type thiolases, not the ACAT1/T2 ortholog. Supporting Evidence: PMID:22758915 The subcellular localization of five of these predicted peroxisomal proteins was confirmed. |
| GO:0046952 ketone body catabolic process | ISS GO_REF:0000024 | ACCEPT | Summary: ISS transfer from human ACAT1 (P24752) placing Acat1 in ketone body catabolism (ketolysis: acetoacetyl-CoA + CoA -> 2 acetyl-CoA). Directly corroborated in Drosophila by the demonstration that Acat1 is required in mushroom body neurons for mitochondrial oxidation of imported ketone bodies to acetyl-CoA. Reason: Core biological process. The single experimental fly study identifies Acat1 as a key enzyme of ketone-body oxidation, matching this catabolic term. Supporting Evidence: PMID:35177854 KBs are used by ACAT1 to generate acetyl-CoA in the mitochondria for energy PMID:35177854 we targeted a key enzyme of KB oxidation |
| GO:0042182 ketone catabolic process | ISS GO_REF:0000024 | KEEP AS NON CORE | Summary: ISS annotation to the broader parent "ketone catabolic process". Correct in direction (ketolysis) but less specific than ketone body catabolic process (GO:0046952). Reason: Correct but superseded by the more specific ketone body catabolic process for the core role. |
| GO:0006550 L-isoleucine catabolic process | ISS GO_REF:0000024 | ACCEPT | Summary: ISS transfer from human ACAT1 (P24752). Human T2 uniquely cleaves the 2-methyl-branched intermediate 2-methylacetoacetyl-CoA, the final thiolytic step of L-isoleucine catabolism; the conserved fly ortholog is expected to perform the same step. Reactome (fly) places Acat1 in "Branched-chain amino acid catabolism" (R-DME-70895), consistent with this assignment. Reason: Core biological process inferred from the well-characterized substrate specificity of the human ortholog and supported by fly pathway curation, though no direct fly isoleucine-flux assay exists. Supporting Evidence: PMID:35177854 the orthologue of the human mitochondrial acetoacetyl-CoA thiolase ACAT1, CG10932 |
| GO:0006085 acetyl-CoA biosynthetic process | ISS GO_REF:0000024 | KEEP AS NON CORE | Summary: ISS annotation reflecting that thiolytic cleavage of acetoacetyl-CoA yields acetyl-CoA. This is a reaction-level restatement of the thiolase chemistry (and matches the fly observation that Acat1 generates acetyl-CoA in neurons) rather than a distinct physiological process. Reason: Chemically defensible product of the reaction, but the physiological process core is ketone body catabolism / isoleucine catabolism rather than generic acetyl-CoA biosynthesis. Supporting Evidence: PMID:35177854 KBs are used by ACAT1 to generate acetyl-CoA in the mitochondria for energy |
| GO:0046951 ketone body biosynthetic process | NAS PMID:35177854 Glia fuel neurons with locally synthesized ketone bodies to ... | KEEP AS NON CORE | Summary: NAS annotation to ketogenesis. The reversible thiolase can in principle contribute the acetoacetyl-CoA-forming (condensation) step of ketone-body synthesis, and Silva et al. describe ketogenesis via "the successive actions of a thiolase, the HMGS and the HMG-CoA lyase". However, the direct experimental evidence in that paper for Acat1/CG10932 is its neuronal ketone-body OXIDATION (catabolic) role; the ketogenic (biosynthetic) direction is inferred rather than demonstrated for this gene. Reason: Biologically plausible via the reversible thiolase chemistry, but not the direction directly demonstrated for Acat1 in the fly; the well-supported core is ketone body catabolism. Supporting Evidence: PMID:35177854 acetyl-CoA that will be used to generate acetoacetate by the successive actions of a thiolase, the HMGS and the HMG-CoA lyase |
| GO:0042181 ketone biosynthetic process | NAS PMID:35177854 Glia fuel neurons with locally synthesized ketone bodies to ... | KEEP AS NON CORE | Summary: NAS annotation to the broader parent "ketone biosynthetic process" (ketogenesis direction). Same considerations as GO:0046951: plausible for a reversible thiolase but not the direction directly demonstrated for Acat1 in the fly study. Reason: Broad parent of ketone body biosynthetic process; inferred ketogenic direction, non-core relative to the demonstrated catabolic role. Supporting Evidence: PMID:35177854 acetyl-CoA that will be used to generate acetoacetate by the successive actions of a thiolase, the HMGS and the HMG-CoA lyase |
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Download this section (compressed HTML)Q: In Drosophila, does Acat1 act physiologically in the ketogenic (acetoacetyl-CoA condensation) direction as well as the demonstrated ketolytic (oxidation) direction, i.e. is the thiolase step of glial ketogenesis in Silva et al. 2022 actually catalyzed by CG10932 or by a different thiolase?
Q: Is the peroxisomal localization prediction for Acat1 (ISM, PMID:22758915) ever experimentally realized, or is Acat1 exclusively mitochondrial as expected for the ACAT1/T2 subfamily?
Experiment: Assay recombinant Drosophila Acat1 for acetoacetyl-CoA thiolase (EC 2.3.1.9) activity in both directions and test cleavage of 2-methylacetoacetyl-CoA, alongside a negative control for cholesterol O-acyltransferase (EC 2.3.1.26) activity.
Hypothesis: Acat1 has bidirectional acetoacetyl-CoA thiolase activity and can cleave the 2-methyl-branched isoleucine intermediate, but has no cholesterol-esterifying (SOAT) activity.
Experiment: Determine the endogenous subcellular localization of tagged Acat1 in Drosophila tissues by fluorescence microscopy and organelle fractionation, comparing mitochondrial versus peroxisomal markers.
Hypothesis: Acat1 localizes to the mitochondrial matrix, and the peroxisomal prediction (PMID:22758915) is a targeting-signal artifact not realized in vivo.
Experiment: Metabolically profile ketone-body and isoleucine-pathway intermediates (acetoacetyl-CoA turnover, 2-methylacetoacetyl-CoA, tiglylglycine-equivalents) in Acat1-knockdown versus control flies under fed and starved conditions.
Hypothesis: Acat1 loss impairs both neuronal ketolysis and the final thiolytic step of isoleucine catabolism, producing accumulation of 2-methyl-branched intermediates analogous to human beta-ketothiolase deficiency.
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