gsr-1 encodes the only glutathione reductase of Caenorhabditis elegans, an FAD-containing homodimeric flavoenzyme of the class-I pyridine nucleotide-disulfide oxidoreductase family that uses NADPH to reduce glutathione disulfide (GSSG) to reduced glutathione (GSH). Alternative first exons/start codons produce a mitochondrial isoform (GSR-1a) with a 14-residue N-terminal targeting sequence and a cytosolic isoform (GSR-1b). By keeping the glutathione pool reduced, GSR-1 underpins tolerance of oxidative stress (juglone, paraquat), lifespan, reduction of cuticle disulfides during molting together with thioredoxin reductase TRXR-1, mitochondrial integrity and proteostasis. The cytosolic isoform is essential: loss-of-function mutants are maternal-effect embryonic lethal and are rescued by cytosolic but not mitochondrial GSR-1. gsr-1 is expressed widely (pharynx, intestine, hypodermis, muscle) and is induced by oxidants and starvation through SKN-1.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0000303 response to superoxide | IMP PMID:23593298 The glutathione reductase GSR-1 determines stress tolerance ... | MODIFY | Summary: gsr-1 RNAi animals die on the redox cycler juglone and are also sensitive to paraquat and, less strongly, to cumene hydroperoxide and tert-butyl hydroperoxide. Reason: The phenotype shows GSR-1 is needed to tolerate oxidative stress, but it is not specific to superoxide (organic hydroperoxides also affect the knockdown) and glutathione reductase does not act on superoxide itself; it regenerates GSH consumed while countering the oxidants. The better-supported claim is the broader cellular response to oxidative stress, which the gene already carries by IBA and which overexpression (increased juglone resistance) also supports. Proposed replacements: cellular response to oxidative stress Supporting Evidence: PMID:23593298 Among the 67 GSH-metabolism genes tested only C46F11.2 was found to be absolutely essential for juglone tolerance PMID:23593298 GSR-1 was also found to be crucial for the tolerance against other pro-oxidants including paraquat and to a lesser extent cumene hydroperoxide and tert-butylhydroperoxide PMID:23593298 Transgenic Pgsr-1::gsr-1::GFP worms exhibited a significantly higher resistance towards juglone than the control strain |
| GO:0004362 glutathione-disulfide reductase (NADPH) activity | IBA GO_REF:0000033 | ACCEPT | Summary: GSR-1 is the sole glutathione reductase of C. elegans and sits inside the PANTHER PTHR42737 glutathione reductase clade; the IBA node is seeded by experimentally characterized GRs from yeast to human and by gsr-1 itself. Reason: The IBA is consistent with direct biochemical evidence for this gene (recombinant GSR-1 reduces GSSG with NADPH). The target appearing among the WITH/FROM sources reflects its own experimental grounding. Core molecular function. Supporting Evidence: PMID:23593298 The protein catalysed the reduction of GSSG by using NADPH as an electron donor. file:worm/gsr-1/gsr-1-uniprot.txt Belongs to the class-I pyridine nucleotide-disulfide oxidoreductase family. |
| GO:0004362 glutathione-disulfide reductase (NADPH) activity | IDA PMID:23593298 The glutathione reductase GSR-1 determines stress tolerance ... | ACCEPT | Summary: Recombinant His-tagged GSR-1 (the 459-aa GSR-1b form) is an active homodimer that reduces GSSG using NADPH, with Km 34.1 uM for GSSG and 12.9 uM for NADPH. Reason: Direct enzyme assay with kinetic parameters; core function. Supporting Evidence: PMID:23593298 The protein catalysed the reduction of GSSG by using NADPH as an electron donor. PMID:23593298 The Km values for GSSG and NADPH were found to be 34.1 |
| GO:0004362 glutathione-disulfide reductase (NADPH) activity | IDA PMID:27117030 Glutathione reductase gsr-1 is an essential gene required fo... | ACCEPT | Summary: Recombinant GSR-1b reduces GSSG dose-dependently, whereas the tm3574 deletion protein lacking most of the NADPH domain is inactive; GSR-1b has no thioredoxin reductase activity. Reason: Independent direct assay confirming the NADPH-dependent GSSG reductase activity and its specificity versus thioredoxin; core function. Supporting Evidence: PMID:27117030 we show that wild type GSR-1b is able to reduce GSSG in a dose-dependent manner PMID:27117030 GSR-1b did not function as thioredoxin reductase when using either yeast TRX3 or worm TRX-2 as substrates |
| GO:0004362 glutathione-disulfide reductase (NADPH) activity | IEA GO_REF:0000120 | ACCEPT | Summary: Electronic mapping (InterPro IPR006322, EC 1.8.1.7, RHEA:11740) to glutathione-disulfide reductase activity. Reason: The IPR006322 family signature is specific for glutathione reductases and the activity is directly demonstrated for this protein; the electronic annotation is correct. Supporting Evidence: file:worm/gsr-1/gsr-1-uniprot.txt IPR006322; Glutathione_Rdtase_euk/bac. PMID:23593298 The protein catalysed the reduction of GSSG by using NADPH as an electron donor. |
| GO:0005737 cytoplasm | IEA GO_REF:0000044 | MODIFY | Summary: UniProt subcellular-location mapping to cytoplasm. Reason: Correct but less specific than the evidence: GSR-1b is a soluble cytosolic protein (diffuse GFP signal), and the gene already carries cytosol (IDA, IBA). Refine to cytosol. Proposed replacements: cytosol Supporting Evidence: PMID:27117030 a diffuse GFP labeling indicating a cytoplasmic localization was obtained PMID:23593298 only strong cytosolic GFP expression was observed, possibly obscuring lesser granular mitochondrial GFP signals |
| GO:0005739 mitochondrion | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic inference of mitochondrial location from the GR clade (yeast Glr1, Aspergillus, and gsr-1 itself). Reason: The organelle claim holds: the 14-residue N-terminal extension of GSR-1a is a functional mitochondrial targeting sequence that co-localizes GFP with TOMM-20::mRFP, and GSR-1a::GFP gives a punctate mitochondrial pattern. The IBA node is consistent with the experimental data on this gene. Mitochondrial matrix is added separately as a NEW sub-compartment row. Supporting Evidence: PMID:27117030 Together, we conclude that the C. elegans gsr-1 gene encodes both mitochondrial (GSR-1a) and cytoplasmic (GSR-1b) isoforms. PMID:27117030 the additional 14 amino acids of GSR-1a constitute a functional MTS on its own PMID:27117030 colocalizing with the TOMM-20::mRFP mitochondrial marker |
| GO:0005739 mitochondrion | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt subcellular-location keyword mapping (SL-0173) to mitochondrion. Reason: The organelle claim holds: the 14-residue N-terminal extension of GSR-1a is a functional mitochondrial targeting sequence that co-localizes GFP with TOMM-20::mRFP, and GSR-1a::GFP gives a punctate mitochondrial pattern. The UniProt mitochondrial location rests on similarity to human GSR and the transit peptide; the worm GFP data confirm it. Supporting Evidence: PMID:27117030 Together, we conclude that the C. elegans gsr-1 gene encodes both mitochondrial (GSR-1a) and cytoplasmic (GSR-1b) isoforms. PMID:27117030 colocalizing with the TOMM-20::mRFP mitochondrial marker |
| GO:0005739 mitochondrion | ISS GO_REF:0000024 | ACCEPT | Summary: ISS transfer of mitochondrial location from human GSR (P00390). Reason: The organelle claim holds: the 14-residue N-terminal extension of GSR-1a is a functional mitochondrial targeting sequence that co-localizes GFP with TOMM-20::mRFP, and GSR-1a::GFP gives a punctate mitochondrial pattern. Similarity transfer is confirmed by direct worm data. Supporting Evidence: PMID:27117030 Together, we conclude that the C. elegans gsr-1 gene encodes both mitochondrial (GSR-1a) and cytoplasmic (GSR-1b) isoforms. PMID:27117030 the additional 14 amino acids of GSR-1a constitute a functional MTS on its own |
| GO:0005829 cytosol | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic inference of cytosolic activity for the GR clade. Reason: Consistent with worm data: the GSR-1b isoform lacks the MTS and is cytosolic, and restoring GSR-1 only in the cytoplasm rescues embryonic lethality, showing that the activity that matters for viability is cytosolic. Supporting Evidence: PMID:27117030 a diffuse GFP labeling indicating a cytoplasmic localization was obtained PMID:27117030 cytoplasmic GSR-1b is essential for embryonic development |
| GO:0005829 cytosol | IDA PMID:23593298 The glutathione reductase GSR-1 determines stress tolerance ... | ACCEPT | Summary: GSR-1::GFP expressed from its own promoter gives strong cytosolic fluorescence. Reason: Direct localization; the 2016 isoform-specific fusions confirm that GSR-1b is cytoplasmic, and the cytosolic pool is the essential one. Supporting Evidence: PMID:23593298 only strong cytosolic GFP expression was observed, possibly obscuring lesser granular mitochondrial GFP signals PMID:27117030 a diffuse GFP labeling indicating a cytoplasmic localization was obtained PMID:27117030 cytoplasmic GSR-1b is essential for embryonic development |
| GO:0006749 glutathione metabolic process | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic inference that GRs participate in glutathione metabolism. Reason: GSR-1 catalyses the NADPH-dependent reduction of GSSG to GSH, the recycling step of glutathione metabolism; it therefore performs a step of this process directly. Supporting Evidence: PMID:23593298 The protein catalysed the reduction of GSSG by using NADPH as an electron donor. file:worm/gsr-1/gsr-1-uniprot.txt Catalyzes the reduction of glutathione disulfide (GSSG) to reduced glutathione (GSH). |
| GO:0006749 glutathione metabolic process | IDA PMID:27117030 Glutathione reductase gsr-1 is an essential gene required fo... | ACCEPT | Summary: Recombinant GSR-1b reduces GSSG to GSH in vitro. Reason: GSR-1 catalyses the NADPH-dependent reduction of GSSG to GSH, the recycling step of glutathione metabolism; it therefore performs a step of this process directly. Supporting Evidence: PMID:27117030 we show that wild type GSR-1b is able to reduce GSSG in a dose-dependent manner |
| GO:0006749 glutathione metabolic process | IEA GO_REF:0000120 | ACCEPT | Summary: Electronic annotation from the IPR006322 glutathione reductase family. Reason: GSR-1 catalyses the NADPH-dependent reduction of GSSG to GSH, the recycling step of glutathione metabolism; it therefore performs a step of this process directly. Supporting Evidence: file:worm/gsr-1/gsr-1-uniprot.txt IPR006322; Glutathione_Rdtase_euk/bac. PMID:23593298 The protein catalysed the reduction of GSSG by using NADPH as an electron donor. |
| GO:0016491 oxidoreductase activity | IEA GO_REF:0000002 | MODIFY | Summary: InterPro mapping to the root oxidoreductase activity term. Reason: Correct but uninformatively general; the specific activity, glutathione-disulfide reductase (NADPH) activity, is directly demonstrated and already annotated. Proposed replacements: glutathione-disulfide reductase (NADPH) activity Supporting Evidence: PMID:23593298 The protein catalysed the reduction of GSSG by using NADPH as an electron donor. file:worm/gsr-1/gsr-1-uniprot.txt IPR006322; Glutathione_Rdtase_euk/bac. |
| GO:0016668 oxidoreductase activity, acting on a sulfur group of donors, NAD(P) as acceptor | IEA GO_REF:0000002 | MODIFY | Summary: InterPro (IPR012999 class-I active site) mapping to oxidoreductase acting on a sulfur group of donors, NAD(P) as acceptor. Reason: Correct parent of glutathione-disulfide reductase (NADPH) activity, which is directly demonstrated for this protein; refine to the specific term. Proposed replacements: glutathione-disulfide reductase (NADPH) activity Supporting Evidence: PMID:23593298 The protein catalysed the reduction of GSSG by using NADPH as an electron donor. file:worm/gsr-1/gsr-1-uniprot.txt Belongs to the class-I pyridine nucleotide-disulfide oxidoreductase family. |
| GO:0034599 cellular response to oxidative stress | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic inference of a role in the cellular response to oxidative stress. Reason: Strongly supported in worm: gsr-1 knockdown abolishes juglone tolerance and sensitizes to paraquat and hydroperoxides, overexpression increases juglone resistance, GSSG accumulates under juglone stress after knockdown, and gsr-1 expression is induced by juglone through SKN-1. GSR-1 does part of the work, by regenerating the GSH consumed by antioxidant reactions. Supporting Evidence: PMID:23593298 Among the 67 GSH-metabolism genes tested only C46F11.2 was found to be absolutely essential for juglone tolerance PMID:23593298 GSR-1 was also found to be crucial for the tolerance against other pro-oxidants including paraquat and to a lesser extent cumene hydroperoxide and tert-butylhydroperoxide PMID:23593298 Transgenic Pgsr-1::gsr-1::GFP worms exhibited a significantly higher resistance towards juglone than the control strain PMID:23593298 Simultaneously, the GSSG-concentration altered from 2.0 to about 6.0 nmol/mg protein in control worms and gsr-1(RNAi) worms exposed to juglone stress |
| GO:0042395 ecdysis, collagen and cuticulin-based cuticle | IGI PMID:21199936 Selenoprotein TRXR-1 and GSR-1 are essential for removal of ... | KEEP AS NON CORE | Summary: gsr-1 RNAi in a trxr-1 mutant background arrests molting with partial detachment of old cuticle; cuticle disulfides remain oxidized and exogenous GSH reduces them and induces apolysis. Reason: The cached paper is abstract-only, but the abstract supports a genetic interaction (IGI with trxr-1) and a mechanism in which GSR-1-generated GSH reduces cuticle disulfides, so GSR-1 contributes a step (reducing equivalents) to old-cuticle removal. This is a tissue-specific use of the core GSSG reductase activity rather than a core function; deferring to the curator who read the full text. Supporting Evidence: PMID:21199936 molting involves the regulated reduction of cuticle components driven by TRXR-1 and GSR-1 PMID:21199936 when trxr-1 and gsr-1 function is reduced, disulfide groups in the cuticle remain oxidized file:worm/gsr-1/gsr-1-deep-research-manual.md these are downstream consequences rather than separate molecular activities. |
| GO:0045454 cell redox homeostasis | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic inference that GRs act in cell redox homeostasis. Reason: Maintaining a reduced glutathione pool is the core physiological role of GR; consistent with worm genetics. Supporting Evidence: PMID:27117030 our data support a prominent role of the glutathione reductase/glutathione pathway in maintaining redox homeostasis in the nematode file:worm/gsr-1/gsr-1-uniprot.txt Catalyzes the reduction of glutathione disulfide (GSSG) to reduced glutathione (GSH). |
| GO:0045454 cell redox homeostasis | IEA GO_REF:0000120 | ACCEPT | Summary: Electronic annotation from the GSHR/TRXR-like family (IPR046952). Reason: Correct for this protein; consistent with experimental data. Supporting Evidence: PMID:27117030 our data support a prominent role of the glutathione reductase/glutathione pathway in maintaining redox homeostasis in the nematode PMID:23593298 The protein catalysed the reduction of GSSG by using NADPH as an electron donor. |
| GO:0045454 cell redox homeostasis | IMP PMID:27117030 Glutathione reductase gsr-1 is an essential gene required fo... | ACCEPT | Summary: gsr-1 loss-of-function mutants are maternal-effect embryonic lethal, short-lived and stress sensitive with mitochondrial dysfunction; cytoplasmic GSR-1b rescues. Reason: The authors interpret the phenotypes as loss of glutathione redox homeostasis, and the enzyme directly performs GSSG reduction, so it participates in (not merely is required for) cell redox homeostasis. Core process. Supporting Evidence: PMID:27117030 our data support a prominent role of the glutathione reductase/glutathione pathway in maintaining redox homeostasis in the nematode PMID:27117030 cytoplasmic GSR-1b is essential for embryonic development PMID:27117030 decreased gsr-1 levels cause mitochondrial stress and dysfunction |
| GO:0050660 flavin adenine dinucleotide binding | IBA GO_REF:0000033 | ACCEPT | Summary: FAD binding inferred from family (IBA) and InterPro domains (IEA). Reason: GR is a flavoenzyme; UniProt annotates one FAD per subunit and conserved FAD-binding residues, and the tm3574 analysis shows the FAD/NADPH domains are required for activity. FAD binding is intrinsic to the catalytic mechanism. Supporting Evidence: file:worm/gsr-1/gsr-1-uniprot.txt Note=Binds 1 FAD per subunit. file:worm/gsr-1/gsr-1-uniprot.txt Belongs to the class-I pyridine nucleotide-disulfide oxidoreductase family. |
| GO:0050660 flavin adenine dinucleotide binding | IEA GO_REF:0000002 | ACCEPT | Summary: FAD binding inferred from family (IBA) and InterPro domains (IEA). Reason: GR is a flavoenzyme; UniProt annotates one FAD per subunit and conserved FAD-binding residues, and the tm3574 analysis shows the FAD/NADPH domains are required for activity. FAD binding is intrinsic to the catalytic mechanism. Supporting Evidence: file:worm/gsr-1/gsr-1-uniprot.txt Note=Binds 1 FAD per subunit. file:worm/gsr-1/gsr-1-uniprot.txt Belongs to the class-I pyridine nucleotide-disulfide oxidoreductase family. |
| GO:0050661 NADP binding | IEA GO_REF:0000002 | ACCEPT | Summary: NADP binding from InterPro IPR006322. Reason: NADPH is the electron donor in the demonstrated GSSG reductase reaction (Km 12.9 uM), so NADP(H) binding is directly implied by the kinetics. Supporting Evidence: PMID:23593298 The protein catalysed the reduction of GSSG by using NADPH as an electron donor. PMID:23593298 The Km values for GSSG and NADPH were found to be 34.1 |
| GO:0098869 cellular oxidant detoxification | IEA GO_REF:0000108 | KEEP AS NON CORE | Summary: Inferred by inter-ontology logic from glutathione-disulfide reductase activity. Reason: GR supports oxidant detoxification indirectly by regenerating GSH for peroxidases and other GSH-dependent enzymes; gsr-1 knockdown sensitizes worms to oxidants. Reasonable but downstream of the core activity. Supporting Evidence: PMID:23593298 Among the 67 GSH-metabolism genes tested only C46F11.2 was found to be absolutely essential for juglone tolerance PMID:23593298 GSR-1 was also found to be crucial for the tolerance against other pro-oxidants including paraquat and to a lesser extent cumene hydroperoxide and tert-butylhydroperoxide |
| GO:0005759 mitochondrial matrix | IC Q93379-1 PMID:27117030 Glutathione reductase gsr-1 is an essential gene required fo... | NEW | Summary: The mitochondrial isoform GSR-1a carries an N-terminal targeting sequence predicted to be cleaved by the two matrix proteases, placing the mature soluble enzyme in the matrix. Reason: Independent lines agree: (1) a functional N-terminal MTS predicted to be cleavable by matrix processing peptidases; (2) GSR-1 is a soluble homodimeric enzyme with no transmembrane segment; (3) the orthologous human mitochondrial GSR is in the matrix. The matrix location has not been assayed directly in worm, so this is curator inference (IC), added beside the root mitochondrion rows rather than replacing them. Supporting Evidence: PMID:27117030 This predicts that GSR-1a is located in the mitochondrial matrix while GSR-1b, lacking this putative MTS PMID:27117030 the additional 14 amino acids of GSR-1a constitute a functional MTS on its own |
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Download this section (compressed HTML)Q: What function does mitochondrial GSR-1a serve, given that it cannot rescue embryonic lethality and that TRXR-2 is not a redundant mitochondrial GSSG reductase?
Q: Is the cuticle disulfide reduction during molting catalysed by GSH generated by GSR-1 directly, or via glutaredoxins acting on cuticle proteins?
Q: Does the 14-residue presequence of GSR-1a get cleaved in vivo, placing the mature enzyme in the mitochondrial matrix?
Experiment: Measure compartment-specific glutathione redox potential (cytosolic and matrix-targeted Grx1-roGFP2) in gsr-1 mutants rescued with GSR-1b only, under normal and oxidative-stress conditions.
Hypothesis: GSR-1a is required to keep the mitochondrial glutathione pool reduced under stress, while GSR-1b controls the cytosolic pool.
Type: In vivo redox imaging
Experiment: Determine the N-terminus of mitochondrial GSR-1a by mass spectrometry and test sub-mitochondrial localization by protease protection.
Hypothesis: The GSR-1a MTS is cleaved and the mature enzyme resides in the mitochondrial matrix.
Type: Proteomics and subfractionation
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Download this section (compressed HTML)gsr-1 encodes a glutathionine reductase orthologous to human mitochondrial glutathione reductase (GSR; OMIM:138300); GSR deficiency has been associated with hemolytic anemia; loss of gsr-1 activity, together with that of trxr-1, results in molting defects characterized by a failure to remove old cuticle and reduced, disulfide groups in the cuticle that remain oxidized; gsr-1 expression is reduced in dauer larvae.
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