DFR1 encodes dihydrofolate reductase (DHFR, EC 1.5.1.3), a 211-residue NADPH-dependent enzyme that reduces 7,8-dihydrofolate to 5,6,7,8-tetrahydrofolate. This is the final step of de novo tetrahydrofolate biosynthesis, and the same reaction recycles the dihydrofolate released by thymidylate synthase (Cdc21p) back into the one-carbon pool. Dfr1p is the methotrexate-sensitive DHFR of yeast, and multicopy DFR1 confers strong methotrexate resistance. dfr1 mutants require dTMP, adenine, histidine and methionine, the end products of folate-dependent one-carbon metabolism. They also fail to use folinic acid in rich medium and cannot grow on glycerol, consistent with a mitochondrial role. DHFR activity and GFP-tagged Dfr1p are found in both the cytoplasm and mitochondria. Dfr1p was also recovered in a proteome-wide screen for RNA-binding proteins.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0003729 mRNA binding | IDA PMID:21124907 A screen for RNA-binding proteins in yeast indicates dual fu... | KEEP AS NON CORE | Summary: Tsvetanova et al. protein-microarray screen for RNA-binding proteins found many metabolic enzymes binding RNA, and Dfr1p was among the enzymes selected for follow-up. Vertebrate DHFR binds its own mRNA, but no regulatory function of RNA binding is known for yeast Dfr1p, so this is a possible moonlighting activity rather than core function. Supporting Evidence: PMID:21124907 we selected eleven proteins with catalytic activities (Dfr1p, Gre3p, Map1p, Mdh1p, Mdh3p, Meu1p, Pfk2p, Phr1p, Pot1p, Pre10p, Ymr1p) |
| GO:0004146 dihydrofolate reductase activity | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from DHFR node PTN000167322 (DFR1/SGD:S000005762 is an experimental source). Core activity. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt Reaction=(6S)-5,6,7,8-tetrahydrofolate + NADP(+) = 7,8-dihydrofolate + |
| GO:0004146 dihydrofolate reductase activity | IDA PMID:2838385 Nucleotide sequence of the dihydrofolate reductase gene of S... | ACCEPT | Summary: Fling et al. sequenced DFR and found DHFR activity overproduced ~40-fold from multicopy plasmid, establishing that the gene encodes DHFR; sequence related to chicken and E. coli DHFR. Core activity. Supporting Evidence: PMID:2838385 DHFR was overproduced about 40-fold in S. cerevisiae when the cloned gene was present in the vector YEp24. PMID:2838385 The polypeptide was significantly related to the DHFRs of chicken liver and Escherichia coli. |
| GO:0004146 dihydrofolate reductase activity | IEA GO_REF:0000120 | ACCEPT | Summary: InterPro/ARBA/RHEA:15009/EC 1.5.1.3 mapping; correct. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt Reaction=(6S)-5,6,7,8-tetrahydrofolate + NADP(+) = 7,8-dihydrofolate + |
| GO:0004146 dihydrofolate reductase activity | IMP PMID:1427091 The phenotype of a dihydrofolate reductase mutant of Sacchar... | ACCEPT | Summary: Huang et al. constructed a dfr1 mutant that phenocopies methotrexate treatment (requirements for dTMP, adenine, histidine, methionine), consistent with loss of DHFR activity. Core activity. Supporting Evidence: PMID:1427091 The mutant has auxotrophic growth requirements for the C1 metabolites dTMP, adenine, histidine and methionine, similar to those of wild-type (wt) strains grown in the presence of methotrexate (MTX). |
| GO:0004146 dihydrofolate reductase activity | IMP PMID:2827121 Molecular characterization of the Saccharomyces cerevisiae d... | ACCEPT | Summary: Lagosky et al. showed multicopy DFR1 confers dramatically enhanced resistance to the DHFR inhibitor methotrexate, consistent with Dfr1p being the methotrexate target DHFR. Core activity. Supporting Evidence: PMID:2827121 Yeast strains containing the cloned DFR1 gene on multicopy number shuttle vector plasmids show dramatically enhanced methotrexate resistance. |
| GO:0004146 dihydrofolate reductase activity | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA for EC 1.5.1.3 in ALL-CHORISMATE-PWY-1 (superpathway of chorismate metabolism); correct activity for Dfr1p. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt Reaction=(6S)-5,6,7,8-tetrahydrofolate + NADP(+) = 7,8-dihydrofolate + |
| GO:0004146 dihydrofolate reductase activity | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA for EC 1.5.1.3 in FOLSYN-PWY-1 (superpathway of tetrahydrofolate biosynthesis and salvage); correct activity for Dfr1p. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt Reaction=(6S)-5,6,7,8-tetrahydrofolate + NADP(+) = 7,8-dihydrofolate + |
| GO:0004146 dihydrofolate reductase activity | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA for EC 1.5.1.3 in PWY-6614 (tetrahydrofolate biosynthesis); correct activity for Dfr1p. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt Reaction=(6S)-5,6,7,8-tetrahydrofolate + NADP(+) = 7,8-dihydrofolate + |
| GO:0004146 dihydrofolate reductase activity | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA for EC 1.5.1.3 in PWY3O-20 (folate polyglutamylation); correct activity for Dfr1p. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt Reaction=(6S)-5,6,7,8-tetrahydrofolate + NADP(+) = 7,8-dihydrofolate + |
| GO:0004146 dihydrofolate reductase activity | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA for EC 1.5.1.3 in PWY3O-45 (folate biosynthesis II); correct activity for Dfr1p. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt Reaction=(6S)-5,6,7,8-tetrahydrofolate + NADP(+) = 7,8-dihydrofolate + |
| GO:0004146 dihydrofolate reductase activity | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA for EC 1.5.1.3 in PWY3O-697 (folate interconversions); correct activity for Dfr1p. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt Reaction=(6S)-5,6,7,8-tetrahydrofolate + NADP(+) = 7,8-dihydrofolate + |
| GO:0005737 cytoplasm | HDA PMID:14562095 Global analysis of protein localization in budding yeast. | ACCEPT | Summary: Huh et al. GFP survey: Dfr1p in cytoplasm (with a mitochondrial pool). Consistent with older biochemical data showing DHFR activity in both cytoplasm and mitochondria. Supporting Evidence: PMID:15169867 In yeast the biochemical analysis had only revealed that dihydrofolate reductase (DHFR), which catalyzes the last step in THF synthesis is present in both the cytoplasm and mitochondria |
| GO:0005739 mitochondrion | HDA PMID:14562095 Global analysis of protein localization in budding yeast. | ACCEPT | Summary: Huh et al. GFP survey scored a mitochondrial pool of Dfr1p; consistent with biochemical detection of DHFR in both cytoplasm and mitochondria and with the respiratory defect of dfr1 mutants. Supporting Evidence: PMID:15169867 In yeast the biochemical analysis had only revealed that dihydrofolate reductase (DHFR), which catalyzes the last step in THF synthesis is present in both the cytoplasm and mitochondria PMID:1427091 suggesting that the DFR1 gene product is also required for mitochondrial function |
| GO:0005739 mitochondrion | IBA GO_REF:0000033 | ACCEPT | Summary: IBA for mitochondrial activity from PTN000556119 (human DHFR/DHFR2 mitochondrial pools). Yeast evidence independently supports a mitochondrial DHFR pool from a single DFR1 gene, so the inherited location is consistent. Supporting Evidence: PMID:15169867 In yeast the biochemical analysis had only revealed that dihydrofolate reductase (DHFR), which catalyzes the last step in THF synthesis is present in both the cytoplasm and mitochondria |
| GO:0005829 cytosol | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA location from ALL-CHORISMATE-PWY-1 (superpathway of chorismate metabolism). The major Dfr1p pool is cytoplasmic/cytosolic, where DHF is reduced for cytosolic one-carbon metabolism. Supporting Evidence: PMID:15169867 In yeast the biochemical analysis had only revealed that dihydrofolate reductase (DHFR), which catalyzes the last step in THF synthesis is present in both the cytoplasm and mitochondria |
| GO:0005829 cytosol | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA location from FOLSYN-PWY-1 (superpathway of tetrahydrofolate biosynthesis and salvage). The major Dfr1p pool is cytoplasmic/cytosolic, where DHF is reduced for cytosolic one-carbon metabolism. Supporting Evidence: PMID:15169867 In yeast the biochemical analysis had only revealed that dihydrofolate reductase (DHFR), which catalyzes the last step in THF synthesis is present in both the cytoplasm and mitochondria |
| GO:0005829 cytosol | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA location from PWY-6614 (tetrahydrofolate biosynthesis). The major Dfr1p pool is cytoplasmic/cytosolic, where DHF is reduced for cytosolic one-carbon metabolism. Supporting Evidence: PMID:15169867 In yeast the biochemical analysis had only revealed that dihydrofolate reductase (DHFR), which catalyzes the last step in THF synthesis is present in both the cytoplasm and mitochondria |
| GO:0005829 cytosol | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA location from PWY3O-20 (folate polyglutamylation). The major Dfr1p pool is cytoplasmic/cytosolic, where DHF is reduced for cytosolic one-carbon metabolism. Supporting Evidence: PMID:15169867 In yeast the biochemical analysis had only revealed that dihydrofolate reductase (DHFR), which catalyzes the last step in THF synthesis is present in both the cytoplasm and mitochondria |
| GO:0005829 cytosol | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA location from PWY3O-45 (folate biosynthesis II). The major Dfr1p pool is cytoplasmic/cytosolic, where DHF is reduced for cytosolic one-carbon metabolism. Supporting Evidence: PMID:15169867 In yeast the biochemical analysis had only revealed that dihydrofolate reductase (DHFR), which catalyzes the last step in THF synthesis is present in both the cytoplasm and mitochondria |
| GO:0005829 cytosol | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA location from PWY3O-697 (folate interconversions). The major Dfr1p pool is cytoplasmic/cytosolic, where DHF is reduced for cytosolic one-carbon metabolism. Supporting Evidence: PMID:15169867 In yeast the biochemical analysis had only revealed that dihydrofolate reductase (DHFR), which catalyzes the last step in THF synthesis is present in both the cytoplasm and mitochondria |
| GO:0035999 folate cycle | RCA GO_REF:0000123 | MARK AS OVER ANNOTATED | Summary: RCA from the THF superpathway. DHFR regenerates THF from DHF (from thymidylate synthesis or de novo synthesis), but it does not transfer C1 units between THF molecules as tetrahydrofolate interconversion is defined. Its process is tetrahydrofolate biosynthesis / dihydrofolate metabolism. Reason: Better captured by GO:0046654, which is a parent of this term and already annotated. |
| GO:0046417 chorismate metabolic process | RCA GO_REF:0000123 | REMOVE | Summary: RCA from the superpathway of chorismate metabolism; DHFR acts on dihydrofolate, nowhere near chorismate. Superpathway inflation. Reason: No chorismate chemistry. |
| GO:0046452 dihydrofolate metabolic process | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from DHFR node; DHFR consumes dihydrofolate. Correct. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt Reaction=(6S)-5,6,7,8-tetrahydrofolate + NADP(+) = 7,8-dihydrofolate + |
| GO:0046452 dihydrofolate metabolic process | IMP PMID:1427091 The phenotype of a dihydrofolate reductase mutant of Sacchar... | ACCEPT | Summary: dfr1 mutants are auxotrophic for C1 end products, consistent with failure to reduce dihydrofolate. Correct. Supporting Evidence: PMID:1427091 The mutant has auxotrophic growth requirements for the C1 metabolites dTMP, adenine, histidine and methionine, similar to those of wild-type (wt) strains grown in the presence of methotrexate (MTX). |
| GO:0046654 tetrahydrofolate biosynthetic process | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from DHFR node; DHFR catalyses the final step of THF biosynthesis and THF regeneration. Core process. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt PATHWAY: Cofactor biosynthesis; tetrahydrofolate biosynthesis; 5,6,7,8- |
| GO:0046654 tetrahydrofolate biosynthetic process | IEA GO_REF:0000120 | ACCEPT | Summary: InterPro/UniPathway UPA00077 mapping (THF from DHF, step 1/1). Core process. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt PATHWAY: Cofactor biosynthesis; tetrahydrofolate biosynthesis; 5,6,7,8- |
| GO:0046654 tetrahydrofolate biosynthetic process | IMP PMID:1427091 The phenotype of a dihydrofolate reductase mutant of Sacchar... | ACCEPT | Summary: dfr1 mutant shows the C1 auxotrophies expected from loss of THF production. Core process. Supporting Evidence: PMID:1427091 The mutant has auxotrophic growth requirements for the C1 metabolites dTMP, adenine, histidine and methionine, similar to those of wild-type (wt) strains grown in the presence of methotrexate (MTX). |
| GO:0046654 tetrahydrofolate biosynthetic process | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA from PWY-6614 (tetrahydrofolate biosynthesis); DHFR catalyses its last step. Core process. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt PATHWAY: Cofactor biosynthesis; tetrahydrofolate biosynthesis; 5,6,7,8- |
| GO:0046655 folic acid metabolic process | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: IBA from the DHFR node (seeded by rat, E. coli, zebrafish DHFR). DHFRs can reduce folic acid to DHF, the route by which exogenous folic acid enters metabolism; yeast dfr1 mutants also have unexplained folinic acid utilisation defects. Plausible but not the core role. Supporting Evidence: PMID:1427091 The inability of the mutants to metabolize FA suggests that the DFR1 gene product may have a role in folate metabolism in addition to its well-characterized function in the reduction of dihydrofolate. |
| GO:0046655 folic acid metabolic process | RCA GO_REF:0000123 | KEEP AS NON CORE | Summary: RCA from PWY3O-20 (folate polyglutamylation). Acceptable only in the sense that DHFR can reduce folic acid; its principal substrate is dihydrofolate. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt Reaction=(6S)-5,6,7,8-tetrahydrofolate + NADP(+) = 7,8-dihydrofolate + |
| GO:0046655 folic acid metabolic process | RCA GO_REF:0000123 | KEEP AS NON CORE | Summary: RCA from PWY3O-697 (folate interconversions). Acceptable only in the sense that DHFR can reduce folic acid; its principal substrate is dihydrofolate. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt Reaction=(6S)-5,6,7,8-tetrahydrofolate + NADP(+) = 7,8-dihydrofolate + |
| GO:0046656 folic acid biosynthetic process | RCA GO_REF:0000123 | MODIFY | Summary: RCA from PWY3O-45 (folate biosynthesis II). The pathway product in yeast is tetrahydrofolate, of which DHFR catalyses the last step; folic acid itself is not synthesised. Reason: Use tetrahydrofolate biosynthetic process. Proposed replacements: tetrahydrofolate biosynthetic process |
| GO:0050661 NADP binding | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: IBA from DHFR node; NADPH is the hydride donor of DHFR. Mechanistically correct, subsumed by the DHFR activity. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt Reaction=(6S)-5,6,7,8-tetrahydrofolate + NADP(+) = 7,8-dihydrofolate + |
| GO:0050661 NADP binding | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: InterPro IPR012259 (DHFR) mapping to NADP binding; correct cofactor binding, part of the DHFR activity. Supporting Evidence: file:yeast/DFR1/DFR1-uniprot.txt Reaction=(6S)-5,6,7,8-tetrahydrofolate + NADP(+) = 7,8-dihydrofolate + |
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Download this section (compressed HTML)Q: How is a single DFR1 gene product distributed between cytosol and mitochondria (alternative translation start, dual targeting, or import of a fraction)?
Q: Why do dfr1 null mutants fail to utilise folinic acid, and does Dfr1p have a role beyond DHF reduction?
Experiment: Protease-protection assays on isolated mitochondria with tagged Dfr1p, N-terminal mutagenesis of potential alternative start codons, and testing whether a cytosol-restricted Dfr1p rescues the dfr1 glycerol-growth defect.
Hypothesis: A fraction of Dfr1p is imported into mitochondria via a weak or alternative N-terminal targeting signal
Type: mitochondrial import / complementation
Dihydrofolate reductase involved in tetrahydrofolate biosynthesis; required for respiratory metabolism; mutation is functionally complemented by human DHFR
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