ufsp2 encodes a UFM1-specific cysteine protease (peptidase C78 family) that functions as the primary deUFMylase in the UFM1 conjugation pathway. The enzyme cleaves UFM1 from conjugated substrates (deconjugation) and processes UFM1 precursors to expose the C-terminal glycine required for conjugation (maturation). Its principal substrate is UFMylated RPL26 on ER-bound 60S ribosomal subunits, where deUFMylation by ufsp2 is required for ribosome recycling during ER-associated ribosome quality control. All annotations for zebrafish ufsp2 are inferred from mammalian orthologs (human UFSP2 and mouse Ufsp2) by sequence similarity.
Definition: The process in which the ubiquitin-fold modifier UFM1 is cleaved from its target protein, removing one or more covalently attached UFM1 groups from a substrate. This is the deconjugation arm of the UFMylation cycle, catalyzed by UFM1-specific proteases (e.g. UFSP2).
Justification: GO provides a molecular function term for deUFMylase activity (GO:0071567) and a biological process term for the forward reaction, protein ufmylation (GO:0071569), but lacks a corresponding biological process term for the reverse (deconjugation) reaction. Analogous deconjugation processes exist for other ubiquitin-like modifiers as children of GO:0070646 protein modification by small protein removal, namely protein deubiquitination (GO:0016579), protein desumoylation (GO:0016926), and protein deneddylation (GO:0000338). A protein deUFMylation term would complete this set and allow UFSP2 deconjugation activity to be annotated to an appropriately specific BP.
Parent term: protein modification by small protein removal
Supporting Evidence:
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005783 endoplasmic reticulum | IBA GO_REF:0000033 | ACCEPT | Summary: Correct. UFSP2 is recruited to the ER via its N-terminal domain interaction with DDRGK1, and acts on UFMylated RPL26 on ER-bound ribosomes. This is a functionally important localization site. Reason: The ER localization is well supported by structural studies showing the N-terminal domain recruits UFSP2 to the ER via DDRGK1, and functional studies demonstrating its role in deUFMylation of ER-associated ribosomal subunits. Supporting Evidence: PMID:21228277 plays a role in the recognition of its cellular substrate C20orf116 and thus in the recruitment of UfSP2 to the endoplasmic reticulum PMID:30626644 RPL26 UFMylation and de-UFMylation is catalyzed by enzyme complexes tethered to the cytoplasmic surface of the ER file:DANRE/ufsp2/ufsp2-deep-research-falcon.md In the current pathway-centric view, this ER tethering spatially matches UFSP2 to major UFMylation events on **ER-associated ribosomes**, and thus to ER proteostasis/ribosome quality control processes. |
| GO:0071567 deUFMylase activity | IBA GO_REF:0000033 | ACCEPT | Summary: Correct. deUFMylase activity is the core molecular function of UFSP2. The enzyme cleaves UFM1 from conjugated substrates with strict specificity, using a catalytic cysteine-dependent mechanism. Reason: This is the defining molecular function of UFSP2, supported by direct biochemical evidence from the mammalian orthologs showing specific cleavage of UFM1 conjugates, loss of activity upon Cys-to-Ser mutation, and accumulation of UFMylated RPL26 when UFSP2 is disrupted. Supporting Evidence: PMID:17182609 cleaved the C-terminal extension of Ufm1 but not that of ubiquitin PMID:17182609 replacement of the conserved Cys residue by Ser resulted in a complete loss of the UfSP1 and UfSP2 activities PMID:30626644 RPL26, is the principal target of UFM1 conjugation...UFMylated RPL26 is highly enriched on ER membrane-bound ribosomes and polysomes file:DANRE/ufsp2/ufsp2-deep-research-bioreason-sft.md A cytoplasmic cysteine protease that specializes in processing the ubiquitin-like modifier UFM1 file:DANRE/ufsp2/ufsp2-deep-research-falcon.md UFSP2 is described as a UFM1-specific **cysteine protease/isopeptidase**, with activity dependent on a catalytic cysteine and a conserved **Cys–Asp–His** catalytic triad (often discussed alongside an additional conserved Tyr typical of cysteine-based Ub/UBL proteases). UFSP activity can be inhibited by thiol-reactive reagents such as **N-ethylmaleimide (NEM)** or by mutation of the catalytic cysteine. file:DANRE/ufsp2/ufsp2-deep-research-falcon.md - **UFSP2**: a major **de-UFMylase** for ER-associated substrates, particularly efficiently removing UFM1 from the ribosomal protein **RPL26 (uL24)**; UFSP2 knockout causes a **drastic accumulation of UFMylated proteins** in cell systems summarized in reviews. |
| GO:0005634 nucleus | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: Likely correct but non-core. Nuclear localization of UFSP2 has been reported for the mammalian ortholog, and UFSP2 has been observed to be recruited to DNA double-strand breaks. However, the primary functional site is the ER. Reason: Nuclear localization is reported for the mammalian ortholog and there is evidence for UFSP2 function at DNA damage sites, but the core function of UFSP2 is at the ER where it deUFMylates RPL26 on ribosomal subunits. Falcon deep research notes that reviews list nucleus/cytoplasm/ER localization, but ER tethering is the strongest mechanistic conclusion, consistent with retaining nucleus as non-core. Supporting Evidence: file:DANRE/ufsp2/ufsp2-deep-research-falcon.md Some reviews also list nucleus/cytoplasm/ER localization, but ER tethering is the strongest mechanistic conclusion. |
| GO:0006508 proteolysis | IBA GO_REF:0000033 | MODIFY | Summary: Correct but too general. UFSP2 performs a highly specific form of proteolysis -- it cleaves the isopeptide bond between UFM1 and its conjugated substrates. The term proteolysis does not capture this specificity. Reason: While proteolysis is technically accurate, it is overly broad for this specialized isopeptidase. The specific process is removal of the ubiquitin-like modifier UFM1 from substrate proteins (deUFMylation). No dedicated GO term for protein deUFMylation exists, so the closest correct existing BP term is GO:0070646 protein modification by small protein removal, whose definition explicitly covers removal of ubiquitin-like proteins (a sibling of protein deneddylation and protein desumoylation). A dedicated protein deUFMylation term is requested in proposed_new_terms. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: PANTHER:PTN008489070 · UFSP UFM1-specific protease ancestral node SUPPORTS TRANSFER The node correctly transfers proteolytic activity to UFSP2, so the propagation is sound. The problem is granularity: GO:0006508 (proteolysis) is far broader than the specific UFM1 deconjugation (deUFMylation) that UFSP2 catalyzes. UniProtKB:Q9NUQ7 · Human UFSP2 SUPPORTS TRANSFER Experimentally characterized ortholog donor establishing UFM1-specific isopeptidase activity. Proposed replacements: protein modification by small protein removal Supporting Evidence: file:DANRE/ufsp2/ufsp2-deep-research-falcon.md Cross-species evidence supports that UFSP2 catalyzes: - **Proteolytic maturation of pro-UFM1** to mature UFM1 required for UFMylation, and - **Hydrolysis of isopeptide bonds** linking UFM1 to substrate proteins (de-UFMylation). |
| GO:0005634 nucleus | IEA GO_REF:0000044 | KEEP AS NON CORE | Summary: Redundant with the IBA annotation for nucleus. Consistent with subcellular localization data from the mammalian ortholog. Reason: Redundant with the IBA-supported nucleus annotation. Retained as non-core since the primary functional site is the ER. |
| GO:0005737 cytoplasm | IEA GO_REF:0000044 | KEEP AS NON CORE | Summary: Correct. UFSP2 is a soluble enzyme that resides in the cytoplasm and is recruited to the ER membrane surface where it acts on UFMylated substrates. Reason: Cytoplasm is listed in UniProt subcellular location data and is consistent with UFSP2 being a soluble protease. However, the mechanistic picture points to constitutive ER tethering via the N-terminal domain interaction with DDRGK1/ODR4 rather than dynamic recruitment, so cytoplasm is retained as a non-core localization. Supporting Evidence: file:DANRE/ufsp2/ufsp2-deep-research-falcon.md Some reviews also list nucleus/cytoplasm/ER localization, but ER tethering is the strongest mechanistic conclusion. |
| GO:0005783 endoplasmic reticulum | IEA GO_REF:0000044 | ACCEPT | Summary: Redundant with the IBA annotation for ER. Correct and consistent with the role of UFSP2 at the ER membrane surface. Reason: Redundant with the IBA-supported ER annotation, but independently correct based on UniProt subcellular location data from the mammalian ortholog. Supporting Evidence: PMID:21228277 plays a role in the recognition of its cellular substrate C20orf116 and thus in the recruitment of UfSP2 to the endoplasmic reticulum file:DANRE/ufsp2/ufsp2-deep-research-falcon.md The UniProt accession **Q7T347** corresponds to **Danio rerio ufsp2**, annotated as **Ufm1-specific protease 2 (UfSP2)**, a cysteine protease in the UFMylation (UFM1 conjugation) system. In the retrieved literature corpus, “UFSP2/Ufsp2” consistently refers to the **UFM1-specific protease 2** (not an unrelated gene), functioning as a UFM1-directed deconjugating enzyme and pro-UFM1 maturase, typically positioned at the **endoplasmic reticulum (ER)** through interaction with **ODR4**. |
| GO:0071567 deUFMylase activity | IEA GO_REF:0000117 | ACCEPT | Summary: Redundant with the IBA annotation for deUFMylase activity. Correct annotation from ARBA machine learning models. Reason: Redundant with the IBA-supported annotation but independently correct. deUFMylase activity is the core molecular function. Supporting Evidence: PMID:17182609 cleaved the C-terminal extension of Ufm1 but not that of ubiquitin file:DANRE/ufsp2/ufsp2-deep-research-falcon.md - **UFSP2**: a major **de-UFMylase** for ER-associated substrates, particularly efficiently removing UFM1 from the ribosomal protein **RPL26 (uL24)**; UFSP2 knockout causes a **drastic accumulation of UFMylated proteins** in cell systems summarized in reviews. |
| GO:0032790 ribosome disassembly | ISS GO_REF:0000024 | ACCEPT | Summary: Correct. UFSP2-mediated deUFMylation of RPL26 is a required step in the recycling of 60S ribosomal subunits from the ER. UFMylation marks stalled ribosomes for quality control, and UFSP2 removes UFM1 to enable ribosome recycling. Reason: Supported by studies showing that RPL26 UFMylation at the ER triggers ribosome quality control and that UFSP2-mediated deUFMylation is needed for 60S subunit recycling. Supporting Evidence: PMID:30626644 RPL26, is the principal target of UFM1 conjugation...UFMylated RPL26 is highly enriched on ER membrane-bound ribosomes and polysomes PMID:37036982 UFMylation of translocon-bound 60S subunits modulates the RTJ to promote access of proteasomes and RQC machinery to ER-APs PMID:38383785 UFM1 conjugation (UFMylation) facilitates the rescue of 60S ribosomal subunits (60S) that are released after ribosome-associated quality-control-mediated splitting of ribosomes that stall during co-translational translocation of secretory proteins into the ER. file:DANRE/ufsp2/ufsp2-deep-research-falcon.md A 2024 Nature study (“UFM1 E3 ligase promotes recycling of 60S ribosomal subunits from the ER”) provides primary, high-authority support for a model in which UFMylation machinery promotes **recycling of 60S ribosomal subunits** from ER-associated states, and it explicitly frames UFSP2 as an **ER-tethered UFM1-specific hydrolase** in this functional context. |
| GO:0006508 proteolysis | ISS GO_REF:0000024 | MODIFY | Summary: Correct but too general. Same issue as the IBA proteolysis annotation. UFSP2 performs specific isopeptide bond cleavage of UFM1 conjugates, not general proteolysis. Reason: Overly broad for this specialized isopeptidase. Should be replaced with a more specific term capturing UFM1 removal. As for the IBA proteolysis annotation, no dedicated protein deUFMylation BP term exists, so the closest correct existing term is GO:0070646 protein modification by small protein removal (a sibling of protein desumoylation and protein deneddylation). A dedicated protein deUFMylation term is requested in proposed_new_terms. Proposed replacements: protein modification by small protein removal Supporting Evidence: file:DANRE/ufsp2/ufsp2-deep-research-falcon.md - **Hydrolysis of isopeptide bonds** linking UFM1 to substrate proteins (de-UFMylation). |
| GO:0071567 deUFMylase activity | ISS GO_REF:0000024 | ACCEPT | Summary: Redundant with IBA and IEA annotations for deUFMylase activity. Correct and well-supported by direct experimental data from the human ortholog. Reason: Core molecular function, well supported by ISS from human UFSP2 (Q9NUQ7). Consistent with all other evidence lines. Supporting Evidence: PMID:17182609 replacement of the conserved Cys residue by Ser resulted in a complete loss of the UfSP1 and UfSP2 activities file:DANRE/ufsp2/ufsp2-deep-research-falcon.md UFSP2 can also reverse UFMylation on model substrates such as **DDRGK1/UFBP1** and **ASC1** in vitro/cell studies. |
| GO:0005634 nucleus | ISS GO_REF:0000024 | KEEP AS NON CORE | Summary: Redundant with IBA and IEA nucleus annotations. Non-core localization. Reason: Nucleus is a secondary localization site. The primary functional site is the ER. |
| GO:0005737 cytoplasm | ISS GO_REF:0000024 | KEEP AS NON CORE | Summary: Redundant with IEA cytoplasm annotation. Correct default localization for this soluble protease. Reason: Cytoplasm is listed in UniProt subcellular location data for this soluble enzyme, but the mechanistic picture points to constitutive ER tethering via DDRGK1/ODR4 rather than dynamic recruitment. Non-core localization. Supporting Evidence: file:DANRE/ufsp2/ufsp2-deep-research-falcon.md Some reviews also list nucleus/cytoplasm/ER localization, but ER tethering is the strongest mechanistic conclusion. |
| GO:0005783 endoplasmic reticulum | ISS GO_REF:0000024 | ACCEPT | Summary: Redundant with IBA and IEA ER annotations. Correct and core localization for UFSP2 function. Reason: ER is the primary functional site where UFSP2 deUFMylates RPL26 on ribosomal subunits. Well supported by structural and functional studies. Supporting Evidence: PMID:21228277 plays a role in the recognition of its cellular substrate C20orf116 and thus in the recruitment of UfSP2 to the endoplasmic reticulum file:DANRE/ufsp2/ufsp2-deep-research-falcon.md In the current pathway-centric view, this ER tethering spatially matches UFSP2 to major UFMylation events on **ER-associated ribosomes**, and thus to ER proteostasis/ribosome quality control processes. |
Loading supporting content…
Download this section (compressed HTML)Q: Has ufsp2 function been directly tested in zebrafish using morpholino knockdown or CRISPR knockout? Are there any zebrafish-specific phenotypes?
Q: Does zebrafish ufsp2 play a role in erythropoiesis or hematopoietic development, as suggested by the UFMylation pathway's role in erythroid differentiation in mammals?
Experiment: Generate ufsp2 CRISPR knockout zebrafish and characterize phenotypes including skeletal development, neurological function, and ER stress markers to determine if the mammalian disease phenotypes are conserved.
Hypothesis: Loss of ufsp2 in zebrafish will cause ER stress-related phenotypes and potentially skeletal or neurological defects analogous to human UFSP2 mutations.
Experiment: Perform immunofluorescence microscopy in zebrafish embryos to confirm subcellular localization of ufsp2 at the ER, cytoplasm, and nucleus.
Hypothesis: Zebrafish ufsp2 will show predominantly ER and cytoplasmic localization, consistent with mammalian orthologs.
Loading supporting content…
Download this section (compressed HTML)Loading supporting content…
Download this section (compressed HTML)Loading supporting content…
Download this section (compressed HTML)Loading supporting content…
Download this section (compressed HTML)Loading supporting content…
Download this section (compressed HTML)