Spa2 (SPAC3G9.05) is the fission-yeast member of the fungal Spa2 family of cortical scaffold proteins, which are related to metazoan GIT proteins through the N-terminal Spa2 homology domain (SHD). Spa2 localizes as arcs at the cortex of growing cell tips, first at the old end and at both ends after new-end take-off, and relocates during mitosis to the cell division site, where it sits in the intermediate layer of the actomyosin contractile ring and lines the forming septum. Ring and tip recruitment depends partly on a PXXP motif binding the SH3 domains of the F-BAR proteins Cdc15 and Imp2, and largely on the exocyst. Spa2 self-associates through coiled-coil regions and binds the Schizosaccharomyces-specific protein Pos1 to form stable oligomeric complexes. Through its SHD it recruits the Rab GAP Gyp3, which acts on the Sec4 homolog Ypt2, to tips and the division site. Loss of Spa2 broadens the growth zone and the distribution of the exocyst, giving shorter, wider cells, and abolishes invasive growth. Unlike budding-yeast Spa2, it has no detectable connection to formin/Bud6 actin assembly or to MAP kinase modules.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0000165 MAPK cascade | IEA GO_REF:0000108 | REMOVE | Summary: Inter-ontology inference from the MAP kinase scaffold activity IBA (GO:0005078). In S. pombe there is no physical or genetic link between Spa2 and the cell-integrity MAPK module, and the MAPKK Pek1 and the MAPKKK Mkh1 localize independently of Spa2. Reason: This row depends entirely on the GO:0005078 IBA, which is not supported for S. pombe Spa2 (see that row). Two independent Gould-lab studies looked for and failed to find the S. cerevisiae-style Spa2βMAPK connection in fission yeast. Supporting Evidence: PMID:25428987 We did not detect physical or genetic interactions between Spa2 and the S. pombe homologues of Bni1, Bud6, or MAP kinases (Supplemental Table S2 and unpublished data). PMID:40931936 Pek1 (also known as Skh1), the ortholog of S. cerevisiae Mkk1 and Mkk2, was not observed at tips, as previously reported (Madrid et al., 2006), and its localization was unaffected in spa2Ξ cells |
| GO:0005078 MAP kinase scaffold activity | IBA GO_REF:0000033 | REMOVE | Summary: MAP kinase scaffold activity propagated from the PTHR21601 family (GOA cites old node PTN000492368; current PAINT places the IBD on the Eukaryota root node PTN004550576, seeded only by S. cerevisiae Spa2/SGD:S000003944). Reason: This is target-specific evidence of divergence from the budding-yeast function. Ren et al. (2015) detected no physical or genetic interaction between S. pombe Spa2 and MAP kinases. Ren et al. (2025) tested the SHDβMAPK connection directly: Spa2 localizes normally without Mkh1, and the Mkk1/2 ortholog Pek1 is neither at cell tips nor dependent on Spa2. In fission yeast the conserved SHD instead recruits the Rab GAP Gyp3. The budding-yeast MAPKK-binding SHD1 motif interaction (PMID:41527849) has not been shown outside Saccharomyces, so a Eukaryota-root placement seeded by one S. cerevisiae gene over-reaches for this branch. Propagation Review Root cause: PROPAGATION BAD Failure modes: FUNCTIONAL DIVERGENCE Sources checked: PANTHER:PTN000492368 Β· PTN000492368 SOURCE STALE OR MISSING GOA node absent from cached PTHR21601 PAINT; the IBD now sits on Eukaryota root PTN004550576. PANTHER:PTN004550576 Β· PTN004550576 SUPPORTS SOURCE BUT NOT TARGET Sole seed is S. cerevisiae Spa2; S. pombe Spa2 has negative MAPK interaction and localization-dependence data. Supporting Evidence: PMID:25428987 We did not detect physical or genetic interactions between Spa2 and the S. pombe homologues of Bni1, Bud6, or MAP kinases (Supplemental Table S2 and unpublished data). PMID:40931936 Pek1 (also known as Skh1), the ortholog of S. cerevisiae Mkk1 and Mkk2, was not observed at tips, as previously reported (Madrid et al., 2006), and its localization was unaffected in spa2Ξ cells PMID:40931936 We also found that the SHD is used to interface with Gyp3, the Rab GAP for the Sec4 homolog Ypt2 |
| GO:0005515 protein binding | IPI PMID:25428987 The Cdc15 and Imp2 SH3 domains cooperatively scaffold a netw... | MODIFY | Summary: IPI with Cdc15 (SPAC20G8.05c). Spa2 was identified as a binding partner of the Cdc15 SH3 domain: a PXXP motif at residues 188-194 binds Cdc15SH3 in two-hybrid and pull-down assays, and the P191A mutation reduces Spa2 recruitment to tips and the contractile ring. Reason: Generic protein binding is uninformative. The paper defines the interaction as Spa2's proline-rich motif binding the SH3 domains of Cdc15 (and Imp2), which GO:0017124 SH3 domain binding captures. Proposed replacements: SH3 domain binding Supporting Evidence: PMID:25428987 Both full-length spa2 and a fragment encoding residues 1β219 interacted with sequences encoding Cdc15SH3 in the yeast two-hybrid assay PMID:25428987 Thus the interaction with the Cdc15 and Imp2 SH3 domains is a strong determinant of Spa2 intracellular localization. |
| GO:0030010 establishment of cell polarity | IMP PMID:40931936 Oligomerization and exocyst coupling underlie Spa2-mediated ... | ACCEPT | Summary: spa2 deletion cells are shorter and wider, with a broader growth zone in which exocyst distribution is less focused at cell tips (Ren et al. 2025). Reason: Direct mutant phenotype in full text. Spa2 is a cortical scaffold at the growing tip that recruits Gyp3 and focuses exocyst distribution, so it contributes structurally to polarity establishment and is not merely required for it. Supporting Evidence: PMID:40931936 Lastly, we re-examined the role of Spa2 in S. pombe morphology and found that Spa2 is required to focus the zone of polarized vegetative growth. PMID:40931936 Upon careful examination, we found that spa2Ξ cells and pos1Ξ cells, which have less Spa2 at polarization sites, were reproducibly shorter and wider than wild-type cells and spa2βGFP cells PMID:40931936 We found that not only were the Exo70βmNG and Sec3βmNG distributions broader in spa2Ξ cells than in wild-type cells |
| GO:0030036 actin cytoskeleton organization | ISO GO_REF:0000024 | MARK AS OVER ANNOTATED | Summary: 2008 PomBase ortholog transfer (ISO) from S. cerevisiae SPA2, where Spa2 is a polarisome scaffold for Bni1/Bud6-driven actin cable assembly. Reason: Later S. pombe data do not support this transfer. spa2 deletion cells have normal F-actin distribution, Spa2 localization is largely F-actin independent, no interaction with the S. pombe Bni1 or Bud6 homologues was detected, and S. pombe lacks Pea2 and Aip5. Only spa2 overexpression perturbs actin. The ISO was a reasonable 2008 curator call, but it now over-states a budding-yeast-specific polarisome-actin function. MARK_AS_OVER_ANNOTATED is used rather than REMOVE because the case against the transfer rests on negative results (no deletion phenotype, no detected interaction), which redundancy could mask, while overexpression does perturb actin and the 2025 study itself leaves an actin link for non-budding-yeast Spa2 proteins open; the term is not shown to be wrong, only unsupported as a function of S. pombe Spa2. Propagation Review Root cause: PROPAGATION BAD Failure modes: FUNCTIONAL DIVERGENCE Sources checked: SGD:S000003944 Β· SPA2 SUPPORTS SOURCE BUT NOT TARGET Budding-yeast Spa2 actin-organization evidence is sound, but S. pombe Spa2 shows normal F-actin when deleted and no Bni1/Bud6 interaction. Supporting Evidence: PMID:25428987 At 25β36Β°C, spa2β cells were morphologically normal, established a normal bipolar growth pattern after cell division, and exhibited normal patterns of F-actin distribution PMID:25428987 We did not detect physical or genetic interactions between Spa2 and the S. pombe homologues of Bni1, Bud6, or MAP kinases (Supplemental Table S2 and unpublished data). PMID:40931936 Thus, whether Spa2 proteins are physically connected to the actin cytoskeleton in organisms other than budding yeast is unclear. PMID:25428987 Further, spa2 overexpression depolarized the actin cytoskeleton and altered cell growth patterns file:SCHPO/spa2/spa2-deep-research-falcon.md copurification of the customary budding-yeast polarisome components or report interactions with the corresponding fission-yeast formin, Bud6 or MAP kinases |
| GO:0030427 site of polarized growth | IBA GO_REF:0000033 | ACCEPT | Summary: Site of polarized growth propagated from PTHR21601 node PTN001091459 (seeded by Spa2 from S. cerevisiae, C. albicans, S. japonicus and S. pombe itself). Reason: Accurate for S. pombe Spa2, which marks growing cell tips; the gene's own IDA is among the seeds, which shows the localization is inherited across fungi. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN001091459 Β· PTN001091459 SUPPORTS TRANSFER Fungal Spa2 node; S. pombe Spa2 has its own direct evidence at growing tips. Supporting Evidence: PMID:25428987 Spa2-GFP was detected in bright arcs at one or both cell tips during interphase; it localized first to the βoldβ cell ends (the cell ends opposite of the CR) after cytokinesis, and then, after new end take off (NETO), it localized to both cell tips |
| GO:0030427 site of polarized growth | IDA PMID:40931936 Oligomerization and exocyst coupling underlie Spa2-mediated ... | ACCEPT | Summary: Spa2 localizes to sites of polarized growth in an exocyst-dependent manner (Ren et al. 2025). Reason: Direct localization evidence in full text. Supporting Evidence: PMID:40931936 we conclude that the exocyst complex plays a major role in targeting Spa2 to sites of polarized growth. |
| GO:0030427 site of polarized growth | IEA GO_REF:0000117 | ACCEPT | Summary: ARBA machine-learning assignment of site of polarized growth. Reason: Consistent with direct evidence. Supporting Evidence: PMID:40931936 we conclude that the exocyst complex plays a major role in targeting Spa2 to sites of polarized growth. |
| GO:0030674 protein-macromolecule adaptor activity | EXP PMID:40931936 Oligomerization and exocyst coupling underlie Spa2-mediated ... | ACCEPT | Summary: PomBase EXP annotation (Ren et al. 2025). Spa2 binds Pos1 through its CC2 region to form oligomeric complexes, and through its SHD binds the Rab GAP Gyp3, which it is absolutely required to localize at cell tips and the division site. Reason: This adaptor activity is the best-supported molecular function of S. pombe Spa2. It brings Gyp3 (and Pos1) to the cortex at growth sites, so that Gyp3 can act on the Sec4-homolog Ypt2 and focus exocytosis. Supporting Evidence: PMID:40931936 At cell tips, stable complexes of oligomerized Spa2 contribute to constraining the growth zone, in part by delivering the Rab GTPase-activating protein for the Sec4 homolog Ypt2. PMID:40931936 Gyp3βmNG localization to cell tips and division sites was lost in spa2Ξ cells PMID:40931936 We also found that the SHD is used to interface with Gyp3, the Rab GAP for the Sec4 homolog Ypt2 PMID:40931936 These results indicate that Pos1 binds the Spa2 CC2. |
| GO:0032153 cell division site | HDA PMID:16823372 ORFeome cloning and global analysis of protein localization ... | ACCEPT | Summary: Genome-wide ORFeome GFP localization (Matsuyama et al. 2006) placed Spa2 at the cell division site. Reason: HDA abstract-only in cache; confirmed by later dedicated studies (Ren et al. 2015, 2025). Supporting Evidence: PMID:16823372 Next, we determined the localization of 4,431 proteins, corresponding to approximately 90% of the fission yeast proteome, by tagging each ORF with the yellow fluorescent protein. PMID:25428987 The previously uncharacterized protein identified here as an SH3 interactor, Spa2, localizes to arc-like structures at cell tips during interphase and the cell division site during cytokinesis and septation. |
| GO:0032153 cell division site | IDA PMID:26702831 Oligomerization but Not Membrane Bending Underlies the Funct... | ACCEPT | Summary: McDonald et al. 2015 (Cdc15 F-BAR oligomerization) used Spa2 as a contractile-ring component whose division-site localization was assayed in Cdc15 F-BAR mutants. Reason: Only the abstract is cached and it does not name Spa2. Spa2 division-site localization is independently established (PMID:25428987, PMID:40931936), so we defer to the PomBase curator, who read the full text. Supporting Evidence: PMID:26702831 Nevertheless, tip-to-tip interactions between F-BAR dimers are critical for Cdc15 oligomerization and high-avidity membrane binding, stabilization of contractile ring components at the medial cortex, and the fidelity of cytokinesis. PMID:25428987 The previously uncharacterized protein identified here as an SH3 interactor, Spa2, localizes to arc-like structures at cell tips during interphase and the cell division site during cytokinesis and septation. |
| GO:0051286 cell tip | HDA PMID:16823372 ORFeome cloning and global analysis of protein localization ... | ACCEPT | Summary: ORFeome GFP localization: Spa2 at cell tips. Reason: Confirmed by dedicated studies. Supporting Evidence: PMID:16823372 Next, we determined the localization of 4,431 proteins, corresponding to approximately 90% of the fission yeast proteome, by tagging each ORF with the yellow fluorescent protein. PMID:25428987 Spa2-GFP was detected in bright arcs at one or both cell tips during interphase; it localized first to the βoldβ cell ends (the cell ends opposite of the CR) after cytokinesis, and then, after new end take off (NETO), it localized to both cell tips |
| GO:0051286 cell tip | IDA PMID:40931936 Oligomerization and exocyst coupling underlie Spa2-mediated ... | ACCEPT | Summary: Spa2 localizes to cell tips (Ren et al. 2025). Reason: Direct evidence. Supporting Evidence: PMID:40931936 we conclude that the exocyst complex plays a major role in targeting Spa2 to sites of polarized growth. PMID:25428987 Spa2-GFP was detected in bright arcs at one or both cell tips during interphase; it localized first to the βoldβ cell ends (the cell ends opposite of the CR) after cytokinesis, and then, after new end take off (NETO), it localized to both cell tips |
| GO:0051286 cell tip | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt subcellular-location mapping (Cell tip). Reason: Consistent with direct evidence. Supporting Evidence: PMID:25428987 Spa2-GFP was detected in bright arcs at one or both cell tips during interphase; it localized first to the βoldβ cell ends (the cell ends opposite of the CR) after cytokinesis, and then, after new end take off (NETO), it localized to both cell tips |
| GO:0110085 mitotic actomyosin contractile ring | IDA PMID:25428987 The Cdc15 and Imp2 SH3 domains cooperatively scaffold a netw... | ACCEPT | Summary: Spa2-GFP colocalizes with Cdc15 at the contractile ring during mitosis and cytokinesis; most of it forms a ring around the Cdc15 ring and lines the forming septum. Reason: Direct live-cell localization; recruitment depends on the Cdc15/Imp2 SH3 domains. Supporting Evidence: PMID:25428987 During mitosis, Spa2 relocalized to the cell division site, where it colocalized with Cdc15-mCherry (Figure 3A), and was especially apparent in prometaphase-arrested cells that organize CRs during the cell cycle block PMID:25428987 Only some Spa2 appeared to constrict with the CR, accumulating at a constriction point, but most appeared to form a ring around the Cdc15 ring and lined the septum as it formed |
| GO:0120105 mitotic actomyosin contractile ring, intermediate layer | IBA GO_REF:0000033 | ACCEPT | Summary: Intermediate ring layer propagated from PTHR21601 (GOA cites PTN001091459; current PAINT puts it on Eukaryota root PTN004550576). The gene's own PomBase IDA is the sole seed. Reason: For S. pombe Spa2 itself the IBA is correct; its presence among its own sources reflects the experimental grounding. The over-broad root placement is a problem for budding-yeast SPA2 and SPH1, not for this gene. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN001091459 Β· PTN001091459 SUPPORTS TRANSFER GOA node; same IBD now on PTN004550576 in cached PAINT. Supported for S. pombe by its own IDA. Supporting Evidence: PMID:28914606 Multiple binding partners of Cdc15's and Imp2βs SH3 domains including Pxl1, Fic1, Spa2, Rgf3, and Cyk3 (Roberts-Galbraith et al., 2010; Ren et al., 2015; Roberts-Galbraith et al., 2009) concentrate in this intermediate zone. |
| GO:0120105 mitotic actomyosin contractile ring, intermediate layer | IDA PMID:28914606 Nanoscale architecture of the Schizosaccharomyces pombe cont... | ACCEPT | Summary: Super-resolution (fPALM/FRET) mapping of the S. pombe contractile ring places Spa2 among the Cdc15/Imp2 SH3-domain partners concentrated in the intermediate zone. Reason: Direct nanoscale localization in full text. Supporting Evidence: PMID:28914606 Multiple binding partners of Cdc15's and Imp2βs SH3 domains including Pxl1, Fic1, Spa2, Rgf3, and Cyk3 (Roberts-Galbraith et al., 2010; Ren et al., 2015; Roberts-Galbraith et al., 2009) concentrate in this intermediate zone. |
| GO:0120207 endocytosis, site selection | IMP PMID:40931936 Oligomerization and exocyst coupling underlie Spa2-mediated ... | UNDECIDED | Summary: PomBase IMP annotation from Ren et al. 2025 to 'endocytosis, site selection'. The cached full text describes exocytosis focusing: spa2 deletion broadens the exocyst (Exo70, Sec3) distribution at cell tips and abolishes Gyp3 tip localization. The main text has no endocytosis assay. Reason: The phenotype in the main text is about where exocytosis occurs, not endocytosis, so the term looks like a possible exo/endo slip. However, the supplementary data are not cached, and under the policy of not overruling curators from incomplete evidence this experimental PomBase annotation is left undecided rather than removed. Query PomBase. Supporting Evidence: PMID:40931936 We found that not only were the Exo70βmNG and Sec3βmNG distributions broader in spa2Ξ cells than in wild-type cells PMID:40931936 Lastly, we re-examined the role of Spa2 in S. pombe morphology and found that Spa2 is required to focus the zone of polarized vegetative growth. |
| GO:1902716 cell cortex of growing cell tip | IDA PMID:25428987 The Cdc15 and Imp2 SH3 domains cooperatively scaffold a netw... | ACCEPT | Summary: Spa2 is in arcs at the cortex of growing cell tips: the old end first, then both ends after NETO. Reason: Direct live-cell localization matching this composite term well. Supporting Evidence: PMID:25428987 Spa2-GFP was detected in bright arcs at one or both cell tips during interphase; it localized first to the βoldβ cell ends (the cell ends opposite of the CR) after cytokinesis, and then, after new end take off (NETO), it localized to both cell tips |
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Download this section (compressed HTML)Q: PomBase annotates spa2 to GO:0120207 'endocytosis, site selection' (IMP, PMID:40931936), but the paper's main-text phenotype is a broadened exocyst/exocytosis zone. Was an exocytosis-site term intended, or do the supplementary data contain an endocytic-patch assay?
Q: Should the S. cerevisiae-seeded GO:0005078 MAP kinase scaffold activity IBD on PTHR21601 root node PTN004550576 be moved down to the Saccharomycetaceae Spa2/Sph1 clade, given the negative MAPK data for S. pombe Spa2?
Q: spa2 deletion abolishes invasive (pseudohyphal) growth in S. pombe (PMID:25428987); is the PTN001091461 (Saccharomycetales) placement of invasive and pseudohyphal growth too shallow for the family?
Experiment: Test whether the Gyp3-binding SHD interface and the Pos1-binding CC2 region of Spa2 are separately required for focusing Exo70/Sec3 at tips, using point mutants rather than deletions; in parallel, image endocytic patch markers (e.g. Fim1) in spa2 deletion cells to resolve the endocytosis site-selection annotation.
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