NCU04302 encodes the Neurospora crassa ortholog of Ubc9, the single E2 conjugating enzyme of the SUMO pathway. The 157-residue protein is a full-length UBC fold (UBC core 3-156) with the catalytic cysteine (Cys92) that accepts activated SUMO from the E1 (Aos1/Uba2) as a thioester and transfers it, usually with an E3 ligase, to lysine residues of target proteins. It sits in the fungal/ invertebrate Ubc9 subfamily (PANTHER PTHR24067:SF248) together with budding-yeast UBC9 and fission-yeast Hus5, at the eukaryote-wide Ubc9 node of the E2 family tree. By orthology it is expected to act mainly in the nucleus, where in yeasts Ubc9-dependent sumoylation of repair and recombination proteins supports the DNA damage response and sumoylation of HP1-family and H3K9-methyltransferase proteins supports heterochromatin silencing. No Neurospora-specific biochemical or genetic study of the protein has been reported, so its substrates and roles in this fungus are inferred.
Summary: Heterochromatin association transferred from fission-yeast Hus5 (same PANTHER subfamily SF248), where ChIP shows heterochromatin-dependent enrichment; plausible but peripheral to the core E2 activity.
Reason: Re-review change from UNDECIDED. The source ortholog P40984 (Hus5) is in the same PANTHER subfamily (PTHR24067:SF248) as NCU04302, and Shin et al. 2005 showed by ChIP that Hus5 is enriched at heterochromatin in a heterochromatin-dependent manner through binding Swi6/Chp2 (HP1) and Clr4 (H3K9 methyltransferase). Neurospora retains HP1 and the Clr4 ortholog DIM-5, so the recruitment mechanism is plausibly conserved, but no Neurospora localisation data exist. The association is a site of action for a general nuclear E2, not its defining function, so it is kept as non-core.
Summary: Nuclear localization is supported by the conserved Ubc9 phylogenetic annotation at PTN000629675 and the nuclear SUMO-conjugation pathway.
Reason: Nuclear localization is supported by the conserved Ubc9 phylogenetic annotation at PTN000629675 and the nuclear SUMO-conjugation pathway. This is a family-based localization inference rather than direct imaging of NCU04302.
Summary: Nuclear localization is supported by the conserved Ubc9 phylogenetic annotation at PTN000629675 and the nuclear SUMO-conjugation pathway.
Reason: Nuclear localization is supported by the conserved Ubc9 phylogenetic annotation at PTN000629675 and the nuclear SUMO-conjugation pathway. This is a family-based localization inference rather than direct imaging of NCU04302.
Summary: Chromosome association is consistent with the chromatin enrichment of the same-subfamily ortholog Hus5; kept as a non-core site of action.
Reason: Re-review change from UNDECIDED. Although this row comes from an ARBA rule, the claim is independently supported by the ChIP enrichment of fission-yeast Hus5 (same PANTHER subfamily SF248) on chromatin. Many Ubc9 substrates are chromatin proteins, so association with chromosomes is expected, but it is broader than heterochromatin and is not restricted to one chromatin compartment; chromosome is therefore kept at this level rather than narrowed. It is a site of action rather than the core activity.
Hus5 associates with heterochromatin through interactions with heterochromatin proteins and modifies substrates whose sumoylations are required for heterochromatin stability
Summary: Ubc9-mediated SUMO conjugation can support genome maintenance through modification of repair proteins.
Reason: Ubc9-mediated SUMO conjugation can support genome maintenance through modification of repair proteins. The fission-yeast ortholog transfer is biologically coherent, but DNA repair is one substrate-dependent role of a general SUMO E2 rather than its defining biochemical function. In fission yeast, hus5 mutants are sensitive to UV, ionising radiation and hydroxyurea, and Hus5 conjugates SUMO (Pmt3) to the Rad52 homolog Rad22 and to Rhp51 in vitro, so the E2 itself performs the modification step on repair proteins; the source ortholog is in the same PANTHER subfamily (SF248).
The Schizosaccharomyces pombe rad31 and hus5 genes are required for the DNA damage response, as mutants defective in these genes are sensitive to DNA damaging agents
The Hus5.62 protein is also capable of forming a thioester link with SUMO, but it does not function in sumoylation assays, either in the modification of Rad22 or in SUMO chain formation.
Summary: Ubc9 catalyzes transfer of activated SUMO through a thioester intermediate; ATP-dependent modifier activation belongs to the E1 step.
Reason: Ubc9 catalyzes transfer of activated SUMO through a thioester intermediate; ATP-dependent modifier activation belongs to the E1 step. The more specific SUMO conjugating enzyme activity identifies the E2 chemistry and avoids the generic ATP-coupled ligase assignment.
Summary: The UBC catalytic domain and PAINT placement at PTN000629675 support the Ubc9 SUMO-E2 subfamily.
Reason: The UBC catalytic domain and PAINT placement at PTN000629675 support the Ubc9 SUMO-E2 subfamily. Biochemical work on yeast and mammalian Ubc9 establishes SUMO thioester formation, making SUMO conjugation a well-grounded conserved function. The target has modifier-specific CDD UBCc_UBE2I and FunFam Ubc9 assignments, corroborated by 75% full-length identity to characterized fission-yeast Ubc9.
The full-length target sequence is 75.0% identical to fission-yeast Ubc9 P40984 across 156 aligned residue pairs (99.36% target coverage), and 62.42% identical to human Ubc9 P63279 across 157 residue pairs (100% target coverage).
The Hus5.62 protein is also capable of forming a thioester link with SUMO, but it does not function in sumoylation assays, either in the modification of Rad22 or in SUMO chain formation.
Summary: The UBC catalytic domain and PAINT placement at PTN000629675 support the Ubc9 SUMO-E2 subfamily.
Reason: The UBC catalytic domain and PAINT placement at PTN000629675 support the Ubc9 SUMO-E2 subfamily. Biochemical work on yeast and mammalian Ubc9 establishes SUMO thioester formation, making SUMO conjugation a well-grounded conserved function. The target has modifier-specific CDD UBCc_UBE2I and FunFam Ubc9 assignments, corroborated by 75% full-length identity to characterized fission-yeast Ubc9.
The full-length target sequence is 75.0% identical to fission-yeast Ubc9 P40984 across 156 aligned residue pairs (99.36% target coverage), and 62.42% identical to human Ubc9 P63279 across 157 residue pairs (100% target coverage).
The Hus5.62 protein is also capable of forming a thioester link with SUMO, but it does not function in sumoylation assays, either in the modification of Rad22 or in SUMO chain formation.
Summary: SUMO transferase activity is true but general; the E2-specific child term SUMO conjugating enzyme activity, which the gene already carries by IBA, is the precise function.
Reason: Re-review change from ACCEPT. GO:0019789 SUMO transferase activity is an is_a ancestor of GO:0061656 SUMO conjugating enzyme activity (checked in QuickGO) and also covers E3 SUMO ligase activities. The evidence (Ubc9-subfamily placement at PTN000629675 and PTHR24067:SF248, catalytic Cys92, ortholog biochemistry) supports the specific E2 activity, which this gene already carries from the IBA and Compara rows, so the general term is refined to that child rather than accepted.
The Pmt3 modification that we detect requires the presence of added Pmt3-GG (the mature form of Pmt3), the activator (Rad31 + Fub2) and the conjugator, Hus5.
Summary: This EC:2.3.2.23 mapping specifies transfer of ubiquitin, whereas PAINT places NCU04302 in the SUMO-conjugating Ubc9 lineage.
Reason: This EC:2.3.2.23 mapping specifies transfer of ubiquitin, whereas PAINT places NCU04302 in the SUMO-conjugating Ubc9 lineage. The substrate distinction is mechanistic: the characterized Ubc9 homologs form SUMO thioesters. Replace the ubiquitin-specific activity with SUMO conjugating enzyme activity. The target has modifier-specific CDD UBCc_UBE2I and FunFam Ubc9 assignments, corroborated by 75% full-length identity to characterized fission-yeast Ubc9.
The full-length target sequence is 75.0% identical to fission-yeast Ubc9 P40984 across 156 aligned residue pairs (99.36% target coverage), and 62.42% identical to human Ubc9 P63279 across 157 residue pairs (100% target coverage).
The Hus5.62 protein is also capable of forming a thioester link with SUMO, but it does not function in sumoylation assays, either in the modification of Rad22 or in SUMO chain formation.
Summary: The UBC catalytic domain and PAINT placement at PTN000629675 support the Ubc9 SUMO-E2 subfamily.
Reason: The UBC catalytic domain and PAINT placement at PTN000629675 support the Ubc9 SUMO-E2 subfamily. Biochemical work on yeast and mammalian Ubc9 establishes SUMO thioester formation, making SUMO conjugation a well-grounded conserved function. The target has modifier-specific CDD UBCc_UBE2I and FunFam Ubc9 assignments, corroborated by 75% full-length identity to characterized fission-yeast Ubc9.
The full-length target sequence is 75.0% identical to fission-yeast Ubc9 P40984 across 156 aligned residue pairs (99.36% target coverage), and 62.42% identical to human Ubc9 P63279 across 157 residue pairs (100% target coverage).
The Hus5.62 protein is also capable of forming a thioester link with SUMO, but it does not function in sumoylation assays, either in the modification of Rad22 or in SUMO chain formation.
Summary: The UBC catalytic domain and PAINT placement at PTN000629675 support the Ubc9 SUMO-E2 subfamily.
Reason: The UBC catalytic domain and PAINT placement at PTN000629675 support the Ubc9 SUMO-E2 subfamily. Biochemical work on yeast and mammalian Ubc9 establishes SUMO thioester formation, making SUMO conjugation a well-grounded conserved function. The target has modifier-specific CDD UBCc_UBE2I and FunFam Ubc9 assignments, corroborated by 75% full-length identity to characterized fission-yeast Ubc9.
The full-length target sequence is 75.0% identical to fission-yeast Ubc9 P40984 across 156 aligned residue pairs (99.36% target coverage), and 62.42% identical to human Ubc9 P63279 across 157 residue pairs (100% target coverage).
The Hus5.62 protein is also capable of forming a thioester link with SUMO, but it does not function in sumoylation assays, either in the modification of Rad22 or in SUMO chain formation.
Summary: Ubc9 engages SUMO E3 ligases and substrates during conjugation, which fits the broad SUMO ligase complex definition; kept as non-core because the assemblies are transient and E3-defined.
Reason: Re-review change from ACCEPT. GO:0106068 is defined as a protein ligase complex consisting of a SUMO-protein transferase and other proteins that may confer substrate specificity (QuickGO). Ubc9 binds E3 ligases and substrates during conjugation, and the fission-yeast source ortholog is in the same PANTHER subfamily (SF248), so the transfer is reasonable. The complexes are transient and defined by their E3 components rather than by Ubc9, no Neurospora E3 partner has been characterised, and PAINT curators did not propagate this complex term at the Ubc9 node (PTN000629675 carries only nucleus, SUMO conjugating enzyme activity and protein sumoylation), so it is kept as a non-core component annotation.
The SUMO E2 Ubc9 serves as a lynchpin in the SUMO conjugation pathway, interacting with the SUMO E1 during activation, with thioester linked SUMO after E1 transfer and with the substrate and SUMO E3 ligases during conjugation.
Core Functions
SUMO E2 conjugating enzyme (Ubc9 ortholog) that accepts SUMO from the E1 as a Cys92 thioester and transfers it to lysines of nuclear target proteins, by orthology to same-subfamily yeast Ubc9/Hus5.
The full-length target sequence is 75.0% identical to fission-yeast Ubc9 P40984 across 156 aligned residue pairs (99.36% target coverage), and 62.42% identical to human Ubc9 P63279 across 157 residue pairs (100% target coverage).
The Hus5.62 protein is also capable of forming a thioester link with SUMO, but it does not function in sumoylation assays, either in the modification of Rad22 or in SUMO chain formation.
The Pmt3 modification that we detect requires the presence of added Pmt3-GG (the mature form of Pmt3), the activator (Rad31 + Fub2) and the conjugator, Hus5.
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.
NCU04302 is the Neurospora crassa Ubc9 ortholog, a SUMO E2 conjugating enzyme. The single ProtNLM2 GO prediction, transferase activity, is correct but less precise than the existing SUMO conjugating enzyme activity (GO:0061656), which is an is_a descendant of it.
Review rationale: The target is a Ubc9-family SUMO E2, supported by modifier-specific CDD UBCc_UBE2I and FunFam Ubc9 assignments, full-length similarity to characterized Ubc9, and the curated SUMO-specific PAINT inference. Characterized Ubc9 homologs form thioester intermediates with SUMO. Transferase activity is therefore correct but less precise than the existing SUMO conjugating enzyme activity. This broad GO result does not validate the separate ubiquitin-specific paragraph.
file:NEUCR/NCU04302/NCU04302-bioinformatics/RESULTS.md: "The full-length target sequence is 75.0% identical to fission-yeast Ubc9 P40984 across 156 aligned residue pairs (99.36% target coverage), and 62.42% identical to human Ubc9 P63279 across 157 residue pairs (100% target coverage)."
PMID:12597774: "The Hus5.62 protein is also capable of forming a thioester link with SUMO, but it does not function in sumoylation assays, either in the modification of Rad22 or in SUMO chain formation."