Q6YYC5

UniProt ID: Q6YYC5
Organism: Oryza sativa subsp. japonica
Review Status: COMPLETE
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Gene Description

OsRGLG4 (Os08g0135400, Q6YYC5, NCBI Gene 4344608) is a RING-type E3 ubiquitin-protein ligase from rice (Oryza sativa subsp. japonica) belonging to the RGLG (RING domain Ligase) family. It is classified as PANTHER subfamily PTHR45751:SF16 (E3 ubiquitin-protein ligase RGLG4) with 65.6% sequence identity to the functionally characterized Arabidopsis ortholog AtRGLG4 (Q9SAL0). The protein contains three conserved domains: an N-terminal von Willebrand factor A (vWA) domain (IPR002035) that mediates protein-protein interactions and substrate recognition, a copine-related C-terminal region (IPR010734/IPR052079) suggesting possible calcium-dependent membrane association, and a C-terminal RING-H2 finger zinc-binding domain (IPR001841) with all eight metal-coordinating residues conserved, confirmed at high confidence by AlphaFold (pLDDT 91.4). Q6YYC5 has a myristoylation-compatible N-terminus (MGGVIG...) unlike AtRGLG4 (MTMGN...), suggesting possible plasma membrane association analogous to AtRGLG1/2. No direct experimental studies have been published on this specific protein. AtRGLG4 has been directly demonstrated to possess E3 ubiquitin ligase activity including in vitro auto-ubiquitination, substrate ubiquitination (GRXS17), and in vivo proteasomal degradation of targets. Within rice, OsRGLG5 and OsRGLG6 have also confirmed E3 ligase activity, reinforcing functional conservation of this protein family.

Existing Annotations Review

GO Term Evidence Action Reason
GO:0004842 ubiquitin-protein transferase activity
IBA
GO_REF:0000033 		MODIFY
Summary: Q6YYC5 contains a canonical RING finger zinc-binding domain (IPR001841, PROSITE PS50089) at its C-terminus (residues 356-389), which is the hallmark catalytic domain of RING-type E3 ubiquitin ligases. The PANTHER classification places it in PTHR45751:SF16 (E3 ubiquitin-protein ligase RGLG4), and the IBA annotation is transferred from four well-characterized Arabidopsis RGLG orthologs (RGLG1-3, RGLG5). The more specific term GO:0061630 (ubiquitin protein ligase activity), which distinguishes E3 ligases from E2 conjugating enzymes, would be more precise since the RING domain specifically mediates E3 (ligase) activity rather than E2 (conjugating enzyme) activity. However, the current annotation is correct and not misleading.
Reason: GO:0004842 covers both E2 and E3 enzymes. Since Q6YYC5 has a RING domain characteristic of E3 ligases (not E2 conjugating enzymes), the more specific child term GO:0061630 (ubiquitin protein ligase activity) is appropriate. This refinement was also independently identified by ProtNLM2 prediction review.
GO:0005634 nucleus
IBA
GO_REF:0000033 		ACCEPT
Summary: Nuclear localization is transferred from Arabidopsis RGLG orthologs via PANTHER phylogenetic inference. Arabidopsis RGLG1 and RGLG2 have been shown to localize to both nucleus and cytoplasm. In rice, the related OsRGLG6 localizes predominantly to cytoplasm and endoplasmic reticulum with minor nuclear presence. The qualifier is_active_in implies the protein carries out its function in the nucleus, which is plausible but uncertain for this uncharacterized protein. Accepted as reasonable phylogenetic inference, though the primary site of activity may be cytoplasmic.
Reason: Consistent with localization data from Arabidopsis RGLG orthologs. Nuclear localization of RGLG family members is established in model species.
GO:0016567 protein ubiquitination
IBA
GO_REF:0000033 		ACCEPT
Summary: Protein ubiquitination is the expected biological process for a RING-type E3 ubiquitin ligase. This annotation is a direct logical consequence of the E3 ligase molecular function (GO:0004842/GO:0061630). Characterized rice RGLG family members such as OsRGLG6 ubiquitinate specific substrates (e.g., OsOTUB1) for proteasomal degradation, and Arabidopsis RGLG1/RGLG2 regulate BIK1 homeostasis through ubiquitination. The term is appropriate at this level of specificity given that no specific substrates are known for Q6YYC5.
Reason: Core biological process for any E3 ubiquitin ligase. Well-supported by domain architecture and phylogenetic inference from characterized RGLG orthologs.
GO:0004842 ubiquitin-protein transferase activity
IEA
GO_REF:0000107 		MODIFY
Summary: This is a duplicate of the IBA annotation for the same GO term, transferred via EnsemblPlants Compara orthology from Arabidopsis RGLG1 (AT1G79380, Q9SAL0). The annotation is correct for the same reasons as the IBA version. As with the IBA annotation, the more specific term GO:0061630 would be preferable.
Reason: Same reasoning as the IBA annotation above. GO:0061630 (ubiquitin protein ligase activity) is more precise for a RING-domain E3 ligase.
GO:0005634 nucleus
IEA
GO_REF:0000107 		ACCEPT
Summary: Orthology-based transfer of nuclear localization from Arabidopsis RGLG1. This annotation uses the qualifier located_in rather than is_active_in (as in the IBA annotation above). Both qualifiers are reasonable. Arabidopsis RGLG family members have been shown to localize to the nucleus. Accepted as consistent with RGLG family localization data.
Reason: Consistent with established localization of Arabidopsis RGLG orthologs. The located_in qualifier is appropriate given that no direct localization data exists for Q6YYC5 in rice.
GO:0005737 cytoplasm
IEA
GO_REF:0000107 		ACCEPT
Summary: Cytoplasmic localization is transferred from Arabidopsis RGLG1 via Ensembl Compara orthology. This is well-supported: in rice, the closely related OsRGLG6 localizes predominantly to the cytoplasm and endoplasmic reticulum. Arabidopsis RGLG family members also show cytoplasmic localization. Cytoplasm is likely the primary compartment where Q6YYC5 functions, consistent with its role as an E3 ligase targeting cytoplasmic substrates for proteasomal degradation.
Reason: Well-supported by localization data from both rice (OsRGLG6) and Arabidopsis (RGLG1) orthologs. Cytoplasm is likely the main site of activity for this E3 ligase.

Core Functions

E3 ubiquitin-protein ligase activity mediated by the C-terminal RING-H2 finger domain with all eight metal-coordinating residues conserved (AlphaFold pLDDT 91.4). The vWA domain provides substrate recognition and protein-protein interaction capability. AtRGLG4 (Q9SAL0, 65.6% identity) has been directly demonstrated to possess E3 ligase activity including in vitro auto-ubiquitination, substrate ubiquitination of GRXS17 via E2 UBC30, and in vivo proteasomal degradation. Rice family members OsRGLG5 and OsRGLG6 also have confirmed E3 ligase activity. The copine-related domain suggests possible calcium-dependent membrane association, and Q6YYC5's myristoylation-compatible N-terminus (MGGVIG...) may indicate plasma membrane association analogous to AtRGLG1/2. OpenScientist investigation strongly confirmed GO:0061630 as the appropriate E3-specific MF term.

Molecular Function:
ubiquitin protein ligase activity
Directly Involved In:
protein ubiquitination
Cellular Locations:
cytoplasm nucleus

References

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Suggested Questions for Experts

Q: What are the specific ubiquitination substrates of Os08g0135400/Q6YYC5 in rice? Identification of substrates would clarify which signaling pathways this RGLG family member regulates.

Q: Does Q6YYC5 form K63-linked polyubiquitin chains like the Arabidopsis ortholog RGLG2, and if so, which E2 conjugating enzyme(s) does it partner with in rice?

Q: Does the copine-related domain in Q6YYC5 confer calcium-dependent membrane association, and does this influence its E3 ligase activity or localization in response to stress or developmental signals?

Suggested Experiments

Experiment: Express and purify recombinant Q6YYC5 and test for E3 ubiquitin ligase activity using in vitro ubiquitination assays with E1, E2 (UBC35/UBC36 or rice orthologs), ubiquitin, and ATP. Test for auto-ubiquitination and substrate ubiquitination with candidate substrates identified by co-IP/mass spectrometry.

Hypothesis: Q6YYC5 has E3 ubiquitin ligase activity in vitro, mediated by its RING domain.

Type: biochemical assay

Experiment: Generate GFP-Q6YYC5 fusion constructs and express in rice protoplasts or stable transgenic lines. Examine subcellular localization by confocal microscopy under normal conditions and following stress treatments (drought, pathogen challenge, ABA application).

Hypothesis: Q6YYC5 localizes to cytoplasm and nucleus in rice cells, with possible stimulus-dependent relocalization.

Type: subcellular localization

External Prediction Reviews

These computational predictions are reviewed separately from the GOA annotation set used for this review. The assessments below are from this project and do not constitute official GO annotations or endorsement by GO/UniProt. They are not included in the existing annotation review above.

ProtNLM2 External predictions

View prediction review YAML · Q6YYC5-protnlm-predictions-review.yaml · Review status: COMPLETE

Q6YYC5 is an RGLG4-like RING-domain E3 ubiquitin ligase with support for generic protein ubiquitination. Its Lys63-linkage refinement is refuted as a paralog overannotation from the K63-demonstrated RGLG1/RGLG2 clade; Q6YYC5 instead clusters with the degradative RGLG3/RGLG4 clade.

Source documents: genes/ORYSJ/Q6YYC5/Q6YYC5-uniprot.txt · genes/ORYSJ/Q6YYC5/Q6YYC5-goa.tsv · publications/PMID_22898498.md · publications/PMID_27497447.md · genes/ORYSJ/Q6YYC5/Q6YYC5-hypotheses/prediction-k63-linked-ubiquitination/openscientist.md

Review score: 2 = concordant with evidence; 1 = uncertain; 0 = discordant with evidence. This is an assessment score, not a model probability.

GO:0070534 protein K63-linked ubiquitination GO_BP
PLI — Paralog incorrect Review score: 0/2
Prediction method: ProtNLM2 · Version: UniProt 2024_06 pilot
PARALOG OVERANNOTATION
Review rationale: The target has a RING domain and an RGLG4 subfamily assignment. Arabidopsis RGLG3 and RGLG4 have experimentally demonstrated ubiquitin ligase activity (PMID:22898498), supporting general ubiquitination but not specifying a Lys63-linked chain for this rice protein. The focused report found that Q6YYC5 clusters with RGLG4, in the RGLG3/RGLG4 degradative clade, rather than the RGLG1/RGLG2 clade tied to K63-linked ubiquitination. Chain linkage also depends on the cognate E2 and cannot be established by the RING domain alone. The linkage-specific process is absent from the cached annotations and appears to be a paralog overannotation from the K63-demonstrated RGLG1/RGLG2 branch.
Supporting Evidence:
  • file:ORYSJ/Q6YYC5/Q6YYC5-uniprot.txt: "ID Q6YYC5_ORYSJ Unreviewed; 401 AA. ... DR GO; GO:0005634; C:nucleus; IBA:GO_Central. ... DR GO; GO:0004842; F:ubiquitin-protein transferase activity; IBA:GO_Central. ... DR GO; GO:0016567; P:protein ubiquitination; IBA:GO_Central. ... DR InterPro; IPR001841; Znf_RING. ... DR PANTHER; PTHR45751:SF16; E3 UBIQUITIN-PROTEIN LIGASE RGLG4; 1. ... FT DOMAIN 356..389 ... FT /note="RING-type""
  • PMID:22898498: "Both RGLG3 and RGLG4 possessed ubiquitin ligase activities"
  • PMID:27497447: "pinpoint UBC30 as a cognate E2 UBC capable of interacting with RGLG3 and RGLG4 and mediating auto-ubiquitination of RGLG3 and ubiquitination of GRXS17 in vitro. Accordingly, GRXS17 is ubiquitinated and degraded in an RGLG3- and RGLG4-dependent manner in planta."
  • file:ORYSJ/Q6YYC5/Q6YYC5-hypotheses/prediction-k63-linked-ubiquitination/openscientist.md: "the orthology used to justify it points to RGLG4, the degradative clade (F002, F004), not the K63-demonstrated RGLG1/2"
GO:0061630 ubiquitin protein ligase activity GO_MF
COR — Correct novel prediction Review score: 2/2
Prediction method: ProtNLM2 · Version: UniProt 2024_06 pilot
Review rationale: The target contains the RING domain and accompanying RGLG-family architecture, with a specific RGLG4 subfamily assignment. Ubiquitin ligase assays on Arabidopsis RGLG3/RGLG4 provide biological grounding for transfer of E3 activity (PMID:22898498). The target already has the broader curated IBA ubiquitin-protein transferase activity, but the exact E3 ligase term is absent from the cached annotations. The prediction is a supported refinement without a claim about substrate or ubiquitin-chain linkage.
Supporting Evidence:
  • file:ORYSJ/Q6YYC5/Q6YYC5-uniprot.txt: "ID Q6YYC5_ORYSJ Unreviewed; 401 AA. ... DR GO; GO:0005634; C:nucleus; IBA:GO_Central. ... DR GO; GO:0004842; F:ubiquitin-protein transferase activity; IBA:GO_Central. ... DR GO; GO:0016567; P:protein ubiquitination; IBA:GO_Central. ... DR InterPro; IPR001841; Znf_RING. ... DR PANTHER; PTHR45751:SF16; E3 UBIQUITIN-PROTEIN LIGASE RGLG4; 1. ... FT DOMAIN 356..389 ... FT /note="RING-type""
  • PMID:22898498: "Both RGLG3 and RGLG4 possessed ubiquitin ligase activities"

Deep Research

Falcon

(Q6YYC5-deep-research-falcon.md)

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OpenScientist

(Q6YYC5-hypotheses/core-function-1-go-0061630/openscientist.md)

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OpenScientist

(Q6YYC5-hypotheses/prediction-k63-linked-ubiquitination/openscientist.md)

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