ASNA-1 (ZK637.5) is the C. elegans ortholog of yeast Get3 and human ASNA1/TRC40/GET3, a cytosolic P-loop ATPase of the ArsA/GET3 family. As a zinc-bridged homodimer it binds the hydrophobic C-terminal transmembrane domain of newly made tail-anchored (TA) proteins and, in an ATP-driven cycle, delivers them to the ER membrane receptor WRB-1 for insertion; in worms it is required for ER targeting of the TA protein SEC-61beta. ASNA-1 is broadly expressed (neurons, intestine, germline and other tissues) and is found in the cytoplasm, at the ER and in Golgi puncta of neurons. It exists in reduced and oxidized (chaperone-like) redox states; the reduced form is needed for TA targeting and cisplatin resistance. Null mutants arrest at the first larval stage, secrete less DAF-28/insulin, and are hypersensitive to cisplatin, arsenite and antimonite. The insulin-secretion role is genetically separable from TA targeting and involves an interaction with the ER chaperone ENPL-1/GRP94, but its mechanism is unknown. Purified ASNA-1 has ATPase activity that is stimulated by arsenite and antimonite, a property retained from the bacterial ArsA ancestor.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005515 protein binding | IPI PMID:14704431 A map of the interactome network of the metazoan C. elegans. | REMOVE | Summary: Large-scale yeast two-hybrid interaction between ASNA-1 and CEE-1 (Q19824) from the first C. elegans interactome map. Reason: Bare protein binding is uninformative and is removed under the protein-binding policy. Removal does not mean the reported interaction is false. The CEE-1 interaction has no functional follow-up and does not define an informative molecular function. |
| GO:0005515 protein binding | IPI PMID:19123269 Empirically controlled mapping of the Caenorhabditis elegans... | REMOVE | Summary: Repeat yeast two-hybrid detection of the ASNA-1/CEE-1 interaction in the empirically controlled C. elegans interactome map. Reason: Bare protein binding is uninformative and is removed under the protein-binding policy. Removal does not mean the interaction is false. The functionally relevant partners of ASNA-1 (TA substrates such as SEC-61beta and the receptor WRB-1) are captured by the protein carrier activity and GET complex terms. |
| GO:0005524 ATP binding | IEA GO_REF:0000120 | ACCEPT | Summary: ASNA-1 has a P-loop (Walker A) ATP-binding motif and the conserved ArsA/GET3 ATPase fold; ATP binding drives dimer closure and TA substrate capture. Reason: ATP binding is a required step of the ArsA/GET3 catalytic cycle and is supported by family membership and the conserved P-loop. The IDA-supported ATP hydrolysis activity implies ATP binding. Supporting Evidence: file:worm/asna-1/asna-1-uniprot.txt Belongs to the arsA ATPase family. file:worm/asna-1/asna-1-uniprot.txt ATP binding drives the homodimer towards the closed dimer state |
| GO:0005737 cytoplasm | IEA GO_REF:0000044 | ACCEPT | Summary: ArsA/GET3 ATPases are soluble cytosolic targeting factors. In worms ASNA-1 is seen in the cytoplasm, at the ER and in Golgi puncta. Reason: Correct location. Cytoplasm is kept, not narrowed to cytosol, because worm ASNA-1 is seen at several cytoplasmic sites (cytosol, ER surface, neuronal Golgi), all within cytoplasm. Supporting Evidence: file:worm/asna-1/asna-1-uniprot.txt Recognizes and selectively binds the transmembrane domain of TA proteins in the cytosol. PMID:36480541 cerevisiae GET3 /mammalian ASNA1 are essential cytosolic ATPases |
| GO:0005783 endoplasmic reticulum | IEA GO_REF:0000120 | ACCEPT | Summary: The UniRule/SubCell mapping places ASNA-1 at the ER, where it docks on its membrane receptor to release TA substrates. In worms ASNA-1::GFP was imaged with the ER marker mCherry::SP12 and binds the ER receptor WRB-1. Reason: Checked the basis of this synthesis row. The ER claim holds. The HAMAP rule reflects the conserved docking of GET3 on the ER receptor, and worm-specific data agree (ASNA-1::GFP imaged with an ER marker; co-IP with ER-localized WRB-1). The association is peripheral and transient (cytosolic face of the ER membrane), so no sub-compartment row is added. The evidence for a stable ER-membrane location is a single line (receptor docking). Supporting Evidence: PMID:33883621 Confocal imaging merge of animals co-expressing mCherry::SP12 with either WRB-1::GFP or ASNA-1::GFP. PMID:33883621 By means of Co-IP/MS/MS analysis, we detected WRB-1 as an interaction partner of ASNA-1::GFP |
| GO:0016887 ATP hydrolysis activity | IBA GO_REF:0000033 | ACCEPT | Summary: The PAINT node for the ArsA/GET3 ATPase family carries ATP hydrolysis activity, grounded on yeast Get3, human ASNA1 and worm ASNA-1 itself. Reason: Core catalytic activity, conserved across the family. That worm ASNA-1 is among the donors reflects its own experimental (IDA) evidence, not circularity. Supporting Evidence: PMID:17419726 The ATPase activity of the ASNA-1 protein was dependent on the presence of As(III) or Sb(III). |
| GO:0016887 ATP hydrolysis activity | IDA PMID:17289575 ASNA-1 positively regulates insulin secretion in C. elegans ... | ACCEPT | Summary: Kao et al. 2007 characterize asna-1 as encoding an ATPase. Only the abstract is cached, but the curator read the full text and the activity agrees with independent biochemistry. Reason: Experimental annotation consistent with the purified-protein ATPase assays of PMID:17419726 and with the conserved GET3 ATPase cycle. Accepted on the curator's reading of the full text. Supporting Evidence: PMID:17289575 asna-1 encodes an ATPase that functions nonautonomously to regulate growth. |
| GO:0016887 ATP hydrolysis activity | IDA PMID:17419726 Caenorhabditis elegans expresses a functional ArsA. | ACCEPT | Summary: Purified MBP-ASNA-1 fusion protein hydrolyses ATP. The activity is stimulated 2-fold by As(III) and 4.6-fold by Sb(III). Reason: Direct biochemical demonstration of ATPase activity with purified recombinant protein. This is the core catalytic activity of the GET3 targeting cycle. Supporting Evidence: PMID:17419726 As(III) stimulated ATPase activity by 2 +/- 0.2-fold, whereas Sb(III) stimulated it by 4.6 +/- 0.15-fold. |
| GO:0016887 ATP hydrolysis activity | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO mapping from the ArsA/GET3 ATPase signatures. Reason: Consistent with the experimental and IBA rows for the same term. Family membership is well supported. Supporting Evidence: file:worm/asna-1/asna-1-uniprot.txt Belongs to the arsA ATPase family. |
| GO:0030424 axon | IDA PMID:17289575 ASNA-1 positively regulates insulin secretion in C. elegans ... | KEEP AS NON CORE | Summary: Reporter-based localization of ASNA-1 in sensory neurons, including axons. Reason: Experimental localization from the curator's full-text reading; consistent with the later finding that ASNA-1 is broadly expressed in neurons. Axonal presence reflects a cytoplasmic protein in neurons and is not where the core TA-targeting activity is defined, so it is non-core. Supporting Evidence: PMID:17289575 asna-1 is expressed in a restricted set of sensory neurons and in insulin-producing intestinal cells. |
| GO:0032024 positive regulation of insulin secretion | IMP PMID:17289575 ASNA-1 positively regulates insulin secretion in C. elegans ... | KEEP AS NON CORE | Summary: asna-1 mutants secrete less DAF-28/insulin and overexpression mimics insulin overexpression. Later work showed ASNA-1 acts in neurons for DAF-28 secretion, interacts with ENPL-1/GRP94 and affects Golgi trafficking. Reason: The genetic requirement is well established and has been reproduced, and ASNA-1 acts cell-autonomously in DAF-28-expressing neurons. The mechanism is unknown and is genetically separable from the core TA-targeting function (wrb-1 mutants are TA-defective but secrete insulin normally). It is therefore kept as a real but non-core regulatory role. Supporting Evidence: PMID:17289575 asna-1 mutants are reduced in insulin secretion while overexpression of asna-1 mimics the effects of insulin overexpression. PMID:36939052 Consistently, ASNA-1 acted in neurons to promote DAF-28/insulin secretion. PMID:33883621 These findings strongly support the conclusion that cisplatin sensitivity and TAP targeting defects are unlinked to insulin secretion defects. |
| GO:0043025 neuronal cell body | IDA PMID:17289575 ASNA-1 positively regulates insulin secretion in C. elegans ... | KEEP AS NON CORE | Summary: Reporter-based localization of ASNA-1 in neuronal cell bodies. Reason: Experimental localization consistent with the neuronal expression and Golgi puncta reported later. It is non-core because it reflects the tissue where the cytoplasmic protein is expressed, not a functional sub-compartment. Supporting Evidence: PMID:36939052 To confirm the neuronal expression, we analyzed worms co-expressing a pan-neuronal nuclear localized tagRFP and ASNA-1::mNeonGreen::AID, and found that ASNA-1::mNeonGreen::AID was expressed in many neurons |
| GO:0043529 GET complex | IBA GO_REF:0000033 | ACCEPT | Summary: GET3 orthologs form the GET complex with the ER receptor (Get1/Get2 in yeast, WRB/CAML in mammals). Worm ASNA-1 co-immunoprecipitates WRB-1, the worm WRB homolog. Reason: The PAINT node placement is sound for a eukaryotic GET3 ortholog. Worm data (co-IP of WRB-1, which is needed for TA targeting, ER-localized and needed to keep ASNA-1 levels normal) support complex formation with the ER receptor. Supporting Evidence: PMID:33883621 Taken together, these three properties of WRB-1 satisfy the requirement to regard this protein as the ER-based ASNA-1 receptor. |
| GO:0045048 protein insertion into ER membrane | IEA GO_REF:0000104 | MODIFY | Summary: UniRule-based process term. True, but the specific branch (tail-anchored protein insertion) is known and is already annotated. Reason: The term is correct but general. ASNA-1 acts in the post-translational tail-anchored branch of ER membrane insertion, so the specific child term GO:0071816 (already carried by IBA and ISS rows) is the supported refinement. Proposed replacements: tail-anchored membrane protein insertion into ER membrane Supporting Evidence: file:worm/asna-1/asna-1-uniprot.txt ATPase required for the post-translational delivery of tail-anchored (TA) proteins to the endoplasmic reticulum. |
| GO:0071722 detoxification of arsenic-containing substance | IMP PMID:17419726 Caenorhabditis elegans expresses a functional ArsA. | KEEP AS NON CORE | Summary: asna-1 mutants are more sensitive to As(III) and Sb(III) toxicity than wild type, and the purified protein has arsenite/antimonite-stimulated ATPase activity. Reason: The mutant phenotype is experimental and the curator read the full text, so the row is kept. In bacteria ArsA powers the ArsB arsenite efflux pump. Eukaryotic GET3 orthologs have been repurposed for TA protein targeting, and no arsenite transporter partner of ASNA-1 has been identified in worms. The sensitivity may reflect a direct role or an indirect effect of losing TA-protein biogenesis or redox-sensor function. The role is peripheral to the core function. Supporting Evidence: PMID:17419726 These results provide evidence that ASNA-1 is required for C. elegans' defense against As(III) and Sb(III) toxicity. file:worm/asna-1/asna-1-deep-research-manual.md consequences of loss of TA protein biogenesis or of redox-sensor function. |
| GO:0071816 tail-anchored membrane protein insertion into ER membrane | IBA GO_REF:0000033 | ACCEPT | Summary: Conserved core process of the eukaryotic GET3/TRC40 clade, grounded on yeast Get3, human ASNA1 and rat Asna1. Worm-specific evidence agrees. Reason: Core process. In worms ASNA-1 is required for ER targeting of the TA protein SEC-61beta, binds the substrate and the ER receptor WRB-1, and the targeting defect is shared with wrb-1 mutants. Supporting Evidence: PMID:33883621 This localization required ASNA-1 since, in the worm asna-1(ok938) protein null mutant8, the ER localization of GFP::SEC-61Ξ² was significantly decreased and the protein was instead detected in cytoplasmic foci PMID:33883621 Indeed, using Co-IP/MS/MS analysis SEC-61Ξ² was detected as an ASNA-1::GFP-interacting partner |
| GO:0071816 tail-anchored membrane protein insertion into ER membrane | ISS PMID:17289575 ASNA-1 positively regulates insulin secretion in C. elegans ... | ACCEPT | Summary: Sequence-similarity transfer from yeast Get3 (Q12154). Later confirmed in worms. Reason: Consistent with the IBA row and with direct worm evidence (PMID:33883621) that ASNA-1 is required for ER targeting of the TA protein SEC-61beta. Supporting Evidence: PMID:33883621 Caenorhabditis elegans ASNA-1 is required for the ER targeting of the TAP SEC-61Ξ². |
| GO:0140597 protein carrier chaperone | ISS PMID:33883621 Alternative redox forms of ASNA-1 separate insulin signaling... | NEW | Summary: GET3/TRC40 family members are soluble carriers that shield the TA transmembrane domain and deliver it to the ER receptor. Worm ASNA-1 binds the TA substrate SEC-61beta and the ER receptor WRB-1, and is required for SEC-61beta ER targeting. Reason: This is the molecular activity, beyond ATP hydrolysis, by which ASNA-1 does the work of TA protein targeting. It is annotated to human GET3 by IDA and fits the worm data (substrate and receptor binding, substrate targeting defect in nulls). It names ASNA-1 as the entity that carries the substrate, so it passes the participation test, unlike a phenotype-based process term. Supporting Evidence: file:worm/asna-1/asna-1-uniprot.txt Recognizes and selectively binds the transmembrane domain of TA proteins in the cytosol. PMID:33883621 Indeed, using Co-IP/MS/MS analysis SEC-61Ξ² was detected as an ASNA-1::GFP-interacting partner |
| GO:0005794 Golgi apparatus | IDA PMID:36939052 A proinsulin-dependent interaction between ENPL-1 and ASNA-1... | NEW | Summary: Endogenously tagged ASNA-1::mNeonGreen forms puncta in neurons that colocalize with the Golgi marker AMAN-2::wrmScarlet. The localization depends on ENPL-1. Reason: Direct localization evidence (colocalization with a Golgi marker) for an endogenously tagged allele. It is relevant to the separate insulin-secretion role. This is a root organelle term from an assay that cannot resolve a sub-compartment, so no sub-compartment is proposed. Supporting Evidence: PMID:36939052 The punctate distribution of ASNA-1 in neurons was found to be in the Golgi, as ASNA-1:mNeonGreen colocalized with AMAN-2::wrmScarlet expressed in head neurons from the rawEx90 transgene PMID:36939052 We conclude that ASNA-1 is found in the Golgi and its localization there depends on ENPL-1. |
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Download this section (compressed HTML)Q: By what mechanism does ASNA-1 promote DAF-28/insulin secretion in neurons, given that this role is separable from TA protein targeting and does not need WRB-1? Does it depend on the oxidized chaperone form and its interaction with ENPL-1?
Q: Is the arsenite and antimonite hypersensitivity of asna-1 mutants caused by a direct metalloid-handling activity of ASNA-1, or indirectly by loss of TA protein biogenesis or redox-sensor function? Is the detoxification process annotation appropriate for a eukaryotic GET3 ortholog?
Q: Which ASNA-1-dependent TA protein substrates account for cisplatin resistance and for the L1 growth arrest?
Experiment: Reconstitute TA protein capture and release with purified reduced and oxidized ASNA-1, a model TA substrate (SEC-61beta) and the WRB-1 cytosolic domain, and measure ATPase stimulation and substrate transfer for wild type, dHis164 and A63V.
Experiment: Use proximity labelling or quantitative ER-membrane proteomics in asna-1 and wrb-1 mutants to define the in vivo set of ASNA-1-dependent TA substrates in neurons and intestine.
Experiment: Test whether ASNA-1 lacking ATPase activity, or locked in the oxidized state, still binds ENPL-1 and rescues DAF-28 secretion when expressed only in neurons.
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