UBP3

Summary: Phylogenetic annotation based on conserved catalytic domain (C19 family, IPR001394). Direct experimental evidence confirms cysteine-type deubiquitinating activity (Baker et al. 1992). IBA appropriately reflects conservation across eukaryotes. Core catalytic function of enzyme.

Reason: IBA evidence reflects true catalytic function conserved across eukaryotic orthologs. EC number 3.4.19.12 assigned to this reaction in UniProt. Essential and well-characterized molecular function. Multiple experimental sources confirm specificity as cysteine-type protease cleaving at C-terminus of ubiquitin.

Summary: Automated inference based on InterPro domain IPR001394 (peptidase C19, ubiquitin carboxyl-terminal hydrolase) and EC number 3.4.19.12. Appropriate for enzyme with characterized catalytic domain and known EC classification.

Reason: IEA from InterPro combined with EC classification is appropriate for a well-characterized deubiquitinase. This is a standard annotation rule for ubiquitin-specific proteases. The domain association (IPR001394) is highly specific to deubiquitination catalysis.

Summary: Experimental demonstration that Ubp3 catalytic activity is required for function. Used catalytic activity as mechanistic explanation for observed phenotypes (Ras/cAMP signaling modulation). Direct evidence of catalytic requirement.

Reason: IMP evidence demonstrates that catalytic activity is mechanistically required for biological function in vivo. Not merely showing activity in vitro but proving essential catalytic role in living cells.

Summary: Direct in vitro biochemical demonstration of cysteine-type deubiquitinating activity by Baker et al. (1992). Original cloning and characterization paper for UBP3. Gold-standard evidence for catalytic activity.

Reason: IDA from primary biochemical characterization providing direct evidence of enzyme activity on ubiquitin substrates. Landmark paper establishing Ubp3 as bona fide deubiquitinase.

Summary: High-throughput direct assay from yeast mRNP proteome survey. Identified Ubp3 in mRNP complexes at proteome-wide level. Limited mechanistic detail; may reflect co-localization in stress granules rather than functional RNA binding.

Reason: HDA identifies co-occurrence in mRNP complexes but does not demonstrate direct functional mRNA binding or substrate specificity. Ubp3 presence in stress granules during assembly may be sufficient to detect by biochemical fractionation without direct catalytic role in mRNA recognition. Consider that functional role is protein deubiquitination, not mRNA interaction per se. IDA in stress granule assembly context suggests presence, not functional binding.

Summary: Proteome-wide search for RNA-binding proteins using high-throughput methods. Ubp3 identified in genome-wide survey. Limited specificity regarding substrate mRNA or mechanism.

Reason: Proteome-wide search methodology identifies proteins physically present in RNA-bound complexes but does not distinguish between direct functional binding and indirect association. Ubp3 likely associates with mRNAs through stress granule complex components rather than direct recognition domain. The functional role in stress granule assembly is deubiquitination, not mRNA binding.

Summary: Automated inference from InterPro domain IPR001394 (peptidase C19, ubiquitin carboxyl-terminal hydrolase). Appropriate inference for enzyme family known to catalyze deubiquitination.

Reason: IEA appropriately infers the biological process from conserved deubiquitinase domain. All ubiquitin-specific proteases (USPs) catalyze protein deubiquitination as fundamental function.

Summary: Experimental mutation analysis demonstrating Ubp3 role in Ste7 (MAPKK) deubiquitination. Dynamic ubiquitination of Ste7 determines MAPK specificity. Ubp3 required for proper signaling response.

Reason: IMP evidence demonstrates that Ubp3-mediated deubiquitination is mechanistically required for normal MAPK pathway regulation. Shows substrate-specific function in vivo.

Summary: Experimental demonstration that Ubp3 activity reverses RNA polymerase II ubiquitylation. IMP shows catalytic activity necessary for transcriptional regulation phenotype.

Reason: IMP evidence from targeted manipulation demonstrates Ubp3 mediates essential deubiquitination of transcriptional machinery (RNAP II). Key role in transcriptional regulation.

Summary: Direct biochemical analysis of Ubp3 activity on RNAP II ubiquitin conjugates. Experimental characterization of deubiquitination substrate and reaction.

Reason: IDA provides direct biochemical evidence of Ubp3-catalyzed deubiquitination of RNAP II in vitro. Gold standard evidence for substrate-specific deubiquitination.

Summary: Functional analysis showing Ubp3 negatively regulates Ras/cAMP signaling through deubiquitination. Catalytic activity essential for signaling modulation.

Reason: IMP demonstrates mechanistic requirement for deubiquitination in signal transduction pathway regulation. Shows biological importance of Ubp3 deubiquitinase activity.

Summary: Named assertion that ribophagy requires Ubp3/Bre5. Experimental identification of Ubp3 catalytic activity requirement for selective ribosome degradation. Process named/discovered in this study.

Reason: NAS appropriately reflects discovery of ribophagy as biological process dependent on Ubp3 function. The term ribophagy was introduced by authors of referenced work (Kraft et al. 2008).

Summary: Genetic analysis demonstrating Ubp3 catalytic activity is essential for selective ribosome degradation. ubp3Delta cells accumulate ribosomal particles during starvation. Phenotypic evidence for mechanistic requirement.

Reason: IMP evidence from knockout/mutation analysis definitively shows Ubp3 catalytic function required for ribophagy. Strong functional evidence from multiple complementary approaches (genetics, accumulation assays, ubiquitination analysis).

Summary: Experimental characterization demonstrating Ubp3 catalytic activity is specifically required for stress granule assembly. Genetic screen identified Ubp3 as unique requirement among UBP family proteases. IDA from primary functional analysis.

Reason: IDA evidence from targeted genetic and biochemical analysis demonstrates Ubp3 catalytic activity is essential for efficient stress granule formation. The catalytic requirement is demonstrated through active site mutation analysis.

Summary: Experimental analysis showing Ubp3 activity is modulated by Hog1 MAPK during osmotic stress response. Control of Ubp3 ubiquitin protease activity by stress signaling SAPK determines transcriptional response. Ubp3 is effector of osmotic stress pathway.

Reason: IMP evidence demonstrates Ubp3 modulates osmotic stress response through protein deubiquitination. Hog1 MAPK-dependent phosphorylation of Ubp3 couples osmostress signaling to deubiquitinase activity.

Summary: Protein-protein interaction between Ubp3 and Hog1 (osmotic stress-response MAPK). IPI based on direct molecular interaction from biochemical evidence.

Reason: IPI evidence documenting physical interaction between Ubp3 and Hog1 kinase supports mechanistic model of osmotic stress response regulation. Interaction is direct and specific.

Summary: Named assertion based on experimental finding that Ubp3-Bre5 specifically deoubiquitinates SEC23 (COPII subunit). Regulation of SEC23 abundance affects ER-Golgi transport efficiency.

Reason: NAS appropriately reflects established mechanistic understanding that Ubp3 regulates COPII-mediated ER-Golgi transport through SEC23 deubiquitination. This is not speculative but well-characterized substrate specificity.

Summary: Experimental analysis of Ubp3 function in vivo. Mutation of Ubp3 or its cofactor Bre5 impairs SEC23 protein levels and ER-Golgi transport. Direct evidence linking catalytic activity to transport process.

Reason: IMP from functional analysis demonstrates Ubp3 catalytic activity is required for normal ER-Golgi transport. The substrate (SEC23) and mechanism (mono-ubiquitination reversal) are characterized.

Summary: Named assertion about Ubp3 role in retrograde transport regulation through deubiquitination. Experimental work demonstrates Ubp3 affects protein movement from Golgi back to ER.

Reason: NAS reflects mechanistic understanding that Ubp3-mediated deubiquitination regulates protein retention/recycling between Golgi and ER. Functionally opposite to anterograde transport but same molecular mechanism (SEC23-related COPII coatomer effects).

Summary: Experimental analysis showing Ubp3-Bre5 complex is required for retention of Golgi membrane proteins. A novel mechanism mediated by deubiquitination. Demonstrated through genetic manipulation.

Reason: IMP evidence from targeted genetic and cellular analysis demonstrates Ubp3-Bre5 complex catalytic activity is required for Golgi protein retention mechanism. This is novel mechanistic insight into how deubiquitination affects protein trafficking.

Summary: Additional experimental evidence (replicate study/condition) demonstrating Ubp3-Bre5 requirement for Golgi protein retention in vivo.

Reason: IMP from same study, different experimental condition or replicate analysis. Supports robustness of Golgi retention phenotype.

Summary: Third experimental evidence supporting Golgi protein retention role. Likely represents different mutant analysis or cellular condition variant from same study.

Reason: IMP from multiple experimental approaches within same research validates Golgi retention function. Demonstrates robustness across conditions.

Summary: Fourth IMP annotation from detailed genetic/cellular analysis of Ubp3-Bre5 in Golgi retention.

Reason: Accumulation of evidence from same study demonstrates comprehensive characterization of Golgi retention phenotype.

Summary: Inferred from genetic interaction analysis. Ubp3 genetically interacts with another component of Golgi retention pathway. IGI indicates functional relationship demonstrated through double mutant or interaction study.

Reason: IGI evidence (genetic interaction with PMID:32673164, SGD:S000002780 likely a Golgi component) supports that Ubp3 functions in the same biological pathway as interacting protein. Demonstrates pathway context.

Summary: Second IGI annotation from genetic interaction analysis in same study.

Reason: IGI confirms pathway membership through additional genetic interaction data point.

Summary: Experimental identification showing Ubp3-Bre5 complex INHIBITS mitophagy while PROMOTING other autophagy pathways (ribophagy). Not all autophagy pathways are equally regulated by Ubp3. This is selective regulatory function.

Reason: While well-supported by evidence, negative regulation of mitophagy represents a specialized regulatory function, not a core catalytic or primary biological role. Ubp3 is positively required for ribophagy and stress granule assembly (core functions), but negatively regulates mitophagy. Kept as non-core annotation because the primary substrate functions do not directly involve mitochondrial proteins, but rather the complex has broader effects on multiple autophagy pathways.

Summary: Direct biochemical evidence that Ubp3 forms heterotetrameric complex with cofactor BRE5 (2 molecules each). Crystal structure available (PDB:2QIY). IPI based on confirmed direct protein-protein interaction.

Reason: IPI evidence from primary biochemical/structural characterization demonstrates stable Ubp3-Bre5 complex. Complex localization and function are well-characterized. This is not generic binding but a named complex with specific catalytic function.

Summary: Phylogenetic annotation based on ortholog function in nucleus. Ubp3 has nuclear localization activity suggested by substrate specificity (RNAP II deubiquitination, histone H2B deubiquitination).

Reason: IBA appropriately reflects nuclear functions documented in experimental literature (RNAP II and histone deubiquitination). Nuclear activity is well-supported by substrate-specific functional analysis.

Summary: Direct experimental detection of Ubp3 in cytoplasm. Consistent with stress granule assembly function, which is cytoplasmic compartment. IDA from cellular localization study.

Reason: IDA from direct cellular fractionation/localization analysis confirms cytoplasmic distribution. Consistent with multiple functional roles (stress granules are cytoplasmic).

Summary: Phylogenetic annotation for cytosolic distribution. Many deubiquitinase substrates localize to cytosol (ribosomal proteins, Ste7 kinase). Appropriate for conserved cytosolic function.

Reason: IBA reflects conservation of cytosolic deubiquitinase activity across eukaryotic orthologs. Substrate specificity supports cytosolic localization (ribophagy targets are cytoplasmic ribosomal proteins).

Summary: Direct experimental evidence from complex portal database indicating Ubp3 localization to cytosol. Consistent with ribophagy and stress granule functions.

Reason: IDA from cellular fractionation/biochemical detection confirms cytosolic presence. Essential for substrate access (cytoplasmic ribosomal proteins, mRNA-binding complexes).

Summary: High-throughput direct assay identifying Ubp3 in cytosolic fraction. Part of systematic subcellular localization study. HDA from large-scale localization screening.

Reason: HDA from systematic proteome localization screening confirms cytosolic distribution. While less specific than targeted IDA studies, provides additional confirmation from independent methodology.

Summary: Direct experimental detection of Ubp3-Bre5 complex at mitochondria during mitophagy induction. Complex translocates dynamically to mitochondria. IDA from cellular biochemistry of dynamic localization.

Reason: IDA demonstrates Ubp3 dynamically translocates to mitochondria specifically during mitophagy response. While not constitutively mitochondrial, transient localization is functionally significant for negative regulation of mitophagy.

Summary: Automated inference from UniProtKB keyword mapping (KW-0378 Hydrolase). Parent term for specific deubiquitinase activity. IEA from standard keyword inference.

Reason: IEA is appropriate for parent/generalized term. Cysteine-type deubiquitinase activity (GO:0004843) is a specialized form of hydrolase activity. Parent term is informative in hierarchy.

Summary: Automated inference from UniProtKB keyword mapping (KW-0645 Protease). Appropriate parent term; Ubp3 is specifically a cysteine-type peptidase.

Reason: IEA appropriately infers parent category peptidase activity from protease keyword. Ubp3 is indeed a peptidase/protease. More specific children (cysteine-type deubiquitinase) are also annotated.

Summary: Automated inference from UniProtKB keyword mapping (KW-0788 Thiol protease). Indicates cysteine-type catalytic mechanism. Appropriate categorization.

Reason: IEA from thiol protease keyword categorization is appropriate. Ubp3 uses cysteine nucleophile (Cys469) in catalytic mechanism. Parent to more specific deubiquitinase annotation.

Summary: Proteolysis is chemically defensible (Ubp3 is a cysteine-type peptidase that hydrolyzes the isopeptide bond linking ubiquitin to substrates) but too broad for UBP3 and obscures its specific deubiquitination-focused biology.

Reason: Not entirely wrong — Ubp3 does hydrolyze an isopeptide bond via a cysteine-protease mechanism, and the corresponding MF term GO:0004843 is indeed classified under peptidase activity — but generic proteolysis is an over-annotation for a deubiquitinase. Note that GO deliberately does not classify GO:0016579 protein deubiquitination under GO:0006508; the BP is under GO:0070646 protein modification by small protein removal, so the broad term is not a parent of the specific process already annotated here. This matches the treatment of the same IEA/TAS proteolysis term on the USP-family DUB USP25.

Summary: Generic protein binding annotation from interaction database (IntAct). IPI from binary interaction data (Bre5 interaction, based on PMID:16429126 - proteome survey). Non-specific and uninformative.

Reason: GO:0005515 protein binding is a genuine, correctly evidenced interaction, not an incorrect one, so REMOVE (defined for annotations unlikely to be correct) is the wrong action; it is overly generic and uninformative rather than wrong. Ubp3 is an enzyme that catalyzes a specific reaction on specific substrates. Catalytic activity (GO:0004843) is already annotated and is far more informative. Protein binding does not distinguish between substrate recognition and non-specific protein aggregation. The catalytic function completely supersedes generic binding annotation. Interaction with BRE5 is better captured by GO:1990861 (Ubp3-Bre5 complex membership).

Summary: Generic protein binding from binary interaction database. Multiple proteins detected binding to Ubp3 (O94742, P39015, P53141, P53741) from high-throughput interaction study.

Reason: The physical interactions are genuinely evidenced (IPI), so REMOVE, which is defined for annotations unlikely to be correct, is not the right action; the issue is that these binary interactions do not indicate catalytic substrates or functional complexes, making the generic term over-annotation rather than error. The meaningful interaction (Bre5) is captured by complex membership term GO:1990861. Following GO curation guidelines that recommend avoiding protein binding in favor of more specific molecular function terms.

Summary: Interaction database annotation for O94742 interaction with Ubp3.

Reason: The interaction is genuinely evidenced (IPI), so REMOVE is not warranted; no mechanistic role is known and generic protein binding is uninformative given the specific catalytic annotation already present, so it is marked as over-annotated rather than incorrect.

Summary: Interaction database annotation for P53141 interaction with Ubp3.

Reason: The interaction is genuinely evidenced (IPI), so REMOVE is not warranted; generic protein binding is uninformative given the specific catalytic annotation already present, so it is marked as over-annotated rather than incorrect.

Summary: Protein binding with Bre5 from structural study (PMID:17632125 - Ubp3-Bre5 crystal structure).

Reason: This interaction (Ubp3-Bre5) is functionally important, well-characterized, and genuinely evidenced, so REMOVE is the wrong action; it should instead be treated as over-annotation, since GO:1990861 (Ubp3-Bre5 complex membership) already captures it far more informatively than generic protein binding.

Summary: Interaction database annotation for Bre5 from large-scale interaction screen.

Reason: The interaction is genuinely evidenced (IPI), so REMOVE is not warranted; complex membership (GO:1990861) already captures this functionally significant interaction more informatively than generic protein binding.

Summary: Protein binding with CDC48 and/or Ufd3 and/or DOA1 from ribophagy complex identification study. IPI indicates these proteins interact with Ubp3.

Reason: These are functionally meaningful, genuinely evidenced interactions in the ribophagy pathway, so REMOVE is not appropriate; the generic protein binding term is uninformative given the more specific process term already annotated (GO:0034517 ribophagy), so it is marked as over-annotation rather than incorrect.

Summary: Protein binding with Bre5 from chromatin-associated protein interaction study.

Reason: The interaction is genuinely evidenced (IPI), so REMOVE is not warranted; it is part of the Ubp3-Bre5 complex (already captured by GO:1990861), so the generic term is over-annotation, not error.

Summary: Protein binding with Hog1 from osmotic stress signaling study. IPI demonstrates direct interaction between Ubp3 and osmotic stress-response kinase.

Reason: This interaction is genuinely evidenced and functionally important for osmotic stress response (regulation of Ubp3 activity by Hog1 phosphorylation), so REMOVE is the wrong action; the relationship is already captured through the more informative process annotation (GO:0047484 regulation of response to osmotic stress) and the corresponding IPI row for that term, so generic protein binding is over-annotation here rather than incorrect.

Summary: Generic protein binding from recent large-scale interaction study (2023). Interactions with P39015 and/or P53741.

Reason: The interaction is genuinely evidenced (IPI), so REMOVE is not warranted; no functional characterization is given and it is redundant with the more informative annotations already present, so it is marked as over-annotation rather than incorrect.

Summary: Phylogenetic annotation suggesting Ubp3 regulates protein stability. This is an indirect consequence of deubiquitination (removing ubiquitin signals targeting proteins for degradation), not a direct catalytic function.

Reason: This is an IBA (phylogenetic) call, and no target-specific divergence or loss evidence for UBP3 was found, so REMOVE is not warranted. The claim itself is also true, not merely generic - Ubp3 demonstrably reverses degradative ubiquitination of specific substrates (Sec23, RNAP II), which is a form of protein-stability regulation. The deubiquitinase activity (GO:0004843) is the direct molecular function, and the multiple substrate-specific deubiquitination process annotations (SEC23 transport, RNAP II regulation, Ste7/MAPK signaling, ribophagy) are more informative than this generic term, so it is retained as non-core rather than removed.