SLC25A32 (also known as MFT/MFTC, the mitochondrial folate transporter) is a member of the SLC25 mitochondrial carrier family: a ~35 kDa multi-pass protein of the mitochondrial inner membrane with three Solcar repeats and six transmembrane helices. It was originally cloned as the carrier that imports folates (tetrahydrofolate) into the mitochondrial matrix, where folate cofactors support mitochondrial one-carbon metabolism. Subsequent biochemical and mouse-genetic work established that its primary transported substrate is flavin adenine dinucleotide (FAD): it translocates FAD across the inner membrane into the matrix, supplying the redox cofactor to mitochondrial flavoenzymes involved in fatty-acid beta-oxidation, amino-acid and choline metabolism, and the electron transport chain. In particular it provides FAD to dihydrolipoamide dehydrogenase (DLD) of the glycine cleavage system, linking it to glycine/formate handling and, indirectly, to folate-mediated one-carbon metabolism. Its effect on mitochondrial folate levels is now understood to be largely an indirect consequence of this FAD-dependent metabolic role. Loss of function causes autosomal-recessive riboflavin-responsive exercise intolerance with combined respiratory-chain/flavoenzyme deficiency, and is associated with folate-resistant, formate-preventable neural tube defects.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0055085 transmembrane transport | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: Correct but very general phylogenetic (IBA) annotation. SLC25A32 is a mitochondrial carrier that mediates transmembrane transport, so this is not wrong, but it is far less informative than the specific FAD transport process. Keep as a true but non-core parent term. |
| GO:0005739 mitochondrion | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: Correct localization at the organelle level, supported by phylogeny and by experimental IDA/HTP annotations. Redundant with the more specific mitochondrial inner membrane term, so keep as non-core. |
| GO:0015230 FAD transmembrane transporter activity | IBA GO_REF:0000033 | ACCEPT | Summary: Core molecular function, here inferred phylogenetically from the FLX1/SLC25A32 FAD-carrier clade. Consistent with the experimental IMP annotations and with UniProt. Accept. |
| GO:0005743 mitochondrial inner membrane | IEA GO_REF:0000044 | ACCEPT | Summary: Correct core localization, mapped electronically from the UniProt subcellular-location vocabulary. Strongly supported by experimental (IGI/IDA/ISS) and TAS annotations. Accept. |
| GO:0006862 nucleotide transport | IEA GO_REF:0000002 | MARK AS OVER ANNOTATED | Summary: Over-generalized InterPro-derived annotation. FAD is a flavin dinucleotide, so a generic "nucleotide transport" term is a distant, uninformative ancestor of the true FAD transport function and does not reflect the substrate specificity of SLC25A32. |
| GO:0015215 nucleotide transmembrane transporter activity | IEA GO_REF:0000117 | MARK AS OVER ANNOTATED | Summary: Over-generalized ARBA-derived molecular-function annotation. The specific and well-supported activity is FAD transmembrane transporter activity (GO:0015230); this generic nucleotide-carrier term is an uninformative parent. If retained it should be replaced by the specific FAD term. Proposed replacements: FAD transmembrane transporter activity |
| GO:0015884 folic acid transport | IEA GO_REF:0000108 | KEEP AS NON CORE | Summary: Inter-ontology inference from the folic acid transporter activity annotation. SLC25A32 was originally defined as the mitochondrial folate transporter, and it does influence mitochondrial folate levels, but mouse genetics show its disease mutations block FAD (not folate) uptake, and UniProt now regards folate handling as an indirect effect. Keep as a real but non-core process. |
| GO:0055085 transmembrane transport | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: Correct but very general InterPro-derived transport annotation, duplicating the IBA call. True but non-core relative to the specific FAD transport process. |
| GO:0005739 mitochondrion | IEA GO_REF:0000120 | KEEP AS NON CORE | Summary: Correct organelle-level localization from combined automated methods; redundant with the more specific inner-membrane term and with experimental IDA annotations. Keep as non-core. |
| GO:0015230 FAD transmembrane transporter activity | IEA GO_REF:0000107 | ACCEPT | Summary: Core molecular function, transferred electronically from the experimentally characterized mouse ortholog (Q8BMG8). Consistent with human IMP evidence and UniProt. Accept. |
| GO:1990548 mitochondrial FAD transmembrane transport | IEA GO_REF:0000107 | ACCEPT | Summary: Core biological-process counterpart of the FAD transporter activity, transferred from the mouse ortholog and matching the human IMP evidence (PMID:35727412, PMID:16165386). Accept. |
| GO:0046655 folic acid metabolic process | TAS Reactome:R-HSA-196757 | KEEP AS NON CORE | Summary: Reactome curator-asserted process reflecting SLC25A32's role in folate/pterine metabolism. SLC25A32 contributes to mitochondrial folate homeostasis and folate-mediated one-carbon metabolism, but does so largely indirectly via FAD-dependent flavoenzymes. Real association; keep as non-core. |
| GO:0005743 mitochondrial inner membrane | IGI PMID:15140890 A mutation inactivating the mitochondrial inner membrane fol... | ACCEPT | Summary: Experimental (genetic-interaction) support for inner-membrane localization: the glyB CHO mutation is a point mutation in the inner mitochondrial membrane folate carrier that abolishes mitochondrial folate transport. Core localization. Accept. |
| GO:0005739 mitochondrion | HTP PMID:34800366 Quantitative high-confidence human mitochondrial proteome an... | KEEP AS NON CORE | Summary: High-throughput mitochondrial-proteome localization to the mitochondrion. Correct at the organelle level and consistent with other evidence; less specific than the inner-membrane annotation, so keep as non-core. |
| GO:0015230 FAD transmembrane transporter activity | IMP PMID:35727412 Mitochondrial FAD shortage in SLC25A32 deficiency affects fo... | ACCEPT | Summary: Strongest experimental support for the core molecular function. Patient-derived knock-in and knockout mice showed that SLC25A32 mutations specifically block mitochondrial FAD uptake while leaving folate uptake intact, directly demonstrating FAD transporter activity. Accept as core. |
| GO:1990548 mitochondrial FAD transmembrane transport | IMP PMID:35727412 Mitochondrial FAD shortage in SLC25A32 deficiency affects fo... | ACCEPT | Summary: Core biological process, experimentally supported by the same mouse models: loss of SLC25A32 function blocks mitochondrial FAD uptake, impairing downstream flavoenzymes. Accept as core. |
| GO:0008517 folic acid transmembrane transporter activity | TAS Reactome:R-HSA-200680 | KEEP AS NON CORE | Summary: Reactome curator-asserted folate (THF) transporter activity, reflecting the original identification of SLC25A32 as the mitochondrial folate transporter. UniProt now notes this was superseded as the primary function by FAD transport, but folate transporter activity remains a documented, curator-supported activity of the protein. Keep as non-core rather than removing an experimental/curator TAS call; the specific FAD activity is the core function. |
| GO:0008517 folic acid transmembrane transporter activity | TAS Reactome:R-HSA-200720 | KEEP AS NON CORE | Summary: Duplicate Reactome TAS folate transporter activity annotation (matrix-to-cytosol direction of the same reversible THF transport reaction). Same assessment as R-HSA-200680: real curator assertion, but non-core relative to FAD transport. |
| GO:0005739 mitochondrion | IDA PMID:29666258 Formate rescues neural tube defects caused by mutations in S... | KEEP AS NON CORE | Summary: Direct experimental localization to the mitochondrion; the truncation variant retained normal mitochondrial localization in this study. Correct organelle-level call, redundant with the inner-membrane term. Keep as non-core. |
| GO:0005743 mitochondrial inner membrane | ISS GO_REF:0000024 | ACCEPT | Summary: Sequence-similarity transfer of the inner-membrane localization from an ortholog. Consistent with experimental IGI/IDA evidence and UniProt. Correct core localization. Accept. |
| GO:1990548 mitochondrial FAD transmembrane transport | IMP PMID:16165386 Identification of the human mitochondrial FAD transporter an... | ACCEPT | Summary: Experimental support for mitochondrial FAD transport: SLC25A32 (MFT) functionally complemented the yeast FLX1 FAD-carrier mutant, identifying it as the human mitochondrial FAD transporter. Core biological process. Accept. |
| GO:0005739 mitochondrion | IDA PMID:21956163 Effect of chronic alcohol exposure on folate uptake by liver... | KEEP AS NON CORE | Summary: Direct-assay localization to mitochondria from a study of carrier-mediated mitochondrial folate uptake (rat liver / HepG2). Correct organelle-level call; redundant with inner-membrane term. Keep as non-core. |
| GO:0005743 mitochondrial inner membrane | TAS Reactome:R-HSA-200680 | ACCEPT | Summary: Reactome curator-asserted inner-membrane localization, agreeing with experimental and UniProt evidence. Core localization. Accept. |
| GO:0005743 mitochondrial inner membrane | TAS Reactome:R-HSA-200720 | ACCEPT | Summary: Duplicate Reactome TAS inner-membrane localization (matrix-to-cytosol reaction). Same core, correct assessment as R-HSA-200680. Accept. |
| GO:0005743 mitochondrial inner membrane | NAS PMID:10978331 Retrovirally mediated complementation of the glyB phenotype.... | ACCEPT | Summary: Author-asserted (NAS) inner-membrane localization from the original cloning paper, which described the protein as an inner mitochondrial wall transporter and restored mitochondrial folate accumulation. Correct core localization, corroborated by later experimental evidence. Accept. |
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Download this section (compressed HTML)Q: Does SLC25A32 transport mitochondrial folate directly at all, or is its influence on mitochondrial folate pools entirely secondary to FAD-dependent one-carbon flux (e.g. via the glycine cleavage system and formate)?
Q: What accounts for the phenotypic and biochemical variability across SLC25A32 patients and mouse models (riboflavin-responsive exercise intolerance versus lethal neural tube defects), and how do specific residues (e.g. K235, Y174) map onto FAD versus residual folate transport?
Experiment: Reconstitute purified human SLC25A32 into proteoliposomes and directly measure transport kinetics for FAD versus tetrahydrofolate/5-methyl-THF (and FMN/riboflavin), to quantify the relative substrate specificity that distinguishes the FAD-carrier from folate-carrier models.
Experiment: Perform substrate-specific flux tracing (labeled FAD precursors and labeled folate/one-carbon units) in SLC25A32-null versus rescued human cells to dissect which mitochondrial folate changes are direct transport effects versus downstream metabolic consequences.
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