CG3226 encodes the Drosophila ortholog of mammalian calcyclin-binding protein / Siah-interacting protein (CacyBP/SIP), a small (230 aa) protein with an N-terminal Siah-interacting domain, a central CS (HSP20-like, SGT1-type) domain and a C-terminal SGS domain. By homology it is expected to act as an adaptor that bridges the RING E3 ubiquitin ligase Sina/Siah to SKP1 and an F-box protein, promoting ubiquitination of targets such as beta-catenin, and to reside in the cytoplasm and nucleus; none of these molecular properties has been tested for the fly protein. Genetic evidence shows that loss of CG3226 causes dilated cardiomyopathy in adult flies, rescued by genomic or heart-specific expression of CG3226. Unlike its vertebrate orthologs, the fly protein has no S100 calcium-binding partners, because the S100 family is restricted to chordates.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005634 nucleus | IBA GO_REF:0000033 | ACCEPT | Summary: Nuclear localization is inferred from the PANTHER node seeded by mammalian CacyBP/SIP, which is experimentally nuclear and cytoplasmic. Not tested in flies, but a reasonable conserved property of this small nucleocytoplasmic adaptor. Reason: The phylogenetic inference is sound (CG3226 is a clear one-to-one member of PTHR13164:SF3) and the human ortholog is experimentally nuclear. Supporting Evidence: file:human/CACYBP/CACYBP-uniprot.txt it localizes in both the nucleus and cytoplasm |
| GO:0005634 nucleus | IEA GO_REF:0000044 | ACCEPT | Summary: Electronic mapping of an ARBA-predicted UniProt subcellular location; agrees with the IBA row and with the experimentally nuclear human ortholog. Reason: Consistent with the IBA nucleus annotation; the same claim is accepted on orthology grounds. Supporting Evidence: file:human/CACYBP/CACYBP-uniprot.txt it localizes in both the nucleus and cytoplasm |
| GO:0005737 cytoplasm | IEA GO_REF:0000044 | ACCEPT | Summary: Electronic mapping of an ARBA-predicted UniProt location; the human ortholog is cytoplasmic at resting calcium, and the predicted E3-adaptor role is cytoplasmic. Reason: Cytoplasmic localization is experimentally established for the human ortholog and is expected to be conserved. Supporting Evidence: file:human/CACYBP/CACYBP-uniprot.txt Cytoplasmic at low calcium |
| GO:0007507 heart development | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: This IBA is seeded in part by CG3226's own IMP annotation (FB:FBgn0029882 in WITH/FROM, which is expected and not circular) plus rat Cacybp. The fly genetic data show a cardiac requirement; the process is a downstream, tissue-level role rather than the protein's molecular activity. Reason: Consistent with the IMP row for the same term. Heart requirement is genetically established in the fly, but the phenotype is adult cardiac function and the mechanism (presumed Sina/Armadillo-related ubiquitination) is untested, so this is a peripheral, non-core process. Supporting Evidence: PMID:22398840 Heart specific expression of CG3226 rescued heart function in Df(1)Exel6240 |
| GO:0007507 heart development | IMP PMID:22398840 Deletion of Siah-interacting protein gene in Drosophila caus... | KEEP AS NON CORE | Summary: Casad et al. identified CG3226 as the gene responsible for dilated cardiomyopathy in Df(1)Exel6240 using smaller deletions and rescue with genomic and heart-specific CG3226 transgenes. The assayed phenotype is adult heart dimensions and fractional shortening; a strictly developmental defect versus a defect in adult heart maintenance or contraction is not distinguished. Reason: Solid genetic evidence (deletion phenocopy plus transgene rescue) that CG3226 is required for normal adult heart structure/function. The FlyBase curator chose heart development from the full text; we defer on term choice, but the term is a pleiotropic tissue-level outcome rather than the protein's core molecular role. Supporting Evidence: PMID:22398840 Drosophila containing the Df(1)Exel6240 deletion and a copy of the CG3226 genomic transgene had a normal heart phenotype, indicating that additional expression of CG3226 rescued the cardiomyopathy PMID:22398840 In our screen, dilated cardiomyopathy is defined as either |
| GO:0015631 tubulin binding | IEA GO_REF:0000002 | MARK AS OVER ANNOTATED | Summary: InterPro2GO transfer from the CacyBP CS-domain entry, reflecting reports of tubulin binding by mammalian CacyBP/SIP. No fly data, and tubulin binding is not part of the established adaptor mechanism. Reason: The tubulin interaction was reported for mammalian CacyBP/SIP in differentiating neuroblastoma cells, and the InterPro2GO mapping carries it to the whole family. Tubulin is conserved in flies, so the term is not impossible, but there is no fly evidence, and tubulin binding is not part of the Siah adaptor mechanism that defines the core function. The human CACYBP review reached the same call. Supporting Evidence: PMID:22398840 was shown to interact with tubulin and actin in differentiating neuroblastoma cells PMID:24078263 Since CacyBP/SIP interacts with tubulin, actin and tropomyosin |
| GO:0019005 SCF ubiquitin ligase complex | IBA GO_REF:0000033 | MARK AS OVER ANNOTATED | Summary: The complex that CacyBP/SIP joins is a Siah RING E3 complex containing SKP1 and the F-box protein Ebi but no cullin, so it is not an SCF (SKP1-cullin-F-box) complex as defined by GO:0019005. Reason: Same rationale as in the human CACYBP review; sharing SKP1 and an F-box protein does not make the Siah-SIP complex an SCF. Not tested in flies. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: COMPARTMENT OR COMPLEX MISMATCH Sources checked: MGI:MGI:1270839 · mouse Cacybp SOURCE BAD Seed annotation equates the cullin-free Siah1-SIP-Skp1-Ebi E3 with an SCF complex. Supporting Evidence: PMID:16085652 The E3 complex comprises, in addition to Siah1, Siah-interacting protein (SIP), the adaptor protein Skp1, and the F-box protein Ebi |
| GO:0031625 ubiquitin protein ligase binding | IBA GO_REF:0000033 | ACCEPT | Summary: Mammalian SIP binds the RING E3 Siah1 directly through its N-terminal region; CG3226 retains the Siah-interacting N-terminal domain, and flies have Sina/Siah orthologs, so the inference is reasonable though untested in flies. Reason: Binding the E3 ligase Siah is the conserved defining interaction of the family, and the fly protein carries the Siah-interacting domain. Supporting Evidence: PMID:16085652 SIP engages Siah1 by means of two elements file:DROME/CG3226/CG3226-uniprot.txt Siah-Interact_N |
| GO:0031625 ubiquitin protein ligase binding | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO version of the same claim as the IBA row; accepted on the same grounds. Reason: Consistent with the IBA row; Siah binding is the family-defining interaction. Supporting Evidence: PMID:16085652 SIP engages Siah1 by means of two elements |
| GO:0044548 S100 protein binding | IEA GO_REF:0000002 | REMOVE | Summary: InterPro2GO transfer of the calcium-dependent S100A6 (calcyclin) interaction of vertebrate CacyBP/SIP to all family members. The S100 family is restricted to chordates (vertebrates plus a few tunicate sequences); D. melanogaster encodes no S100 protein (0 PF01023 proteins in UniProt), so CG3226 has no S100 partner in vivo. This is the annotation flagged by Wei et al. 2020 (PMID:32470107), and the flag holds. Reason: Taxon-inappropriate electronic transfer. The partner family does not exist in insects, so the binding function cannot be exercised in the fly. The fix belongs upstream, as a taxon constraint on GO:0044548 (Chordata) or a restriction of the IPR037893 InterPro2GO mapping. Propagation Review Root cause: PROPAGATION BAD Failure modes: LINEAGE OR TAXON MISMATCH Sources checked: InterPro:IPR037893 · CS domain, CacyBP-type (CS_CacyBP) SUPPORTS SOURCE BUT NOT TARGET The InterPro2GO mapping is valid for vertebrate members, whose S100 partners exist, but is applied family-wide including insects, which have no S100 proteins. Supporting Evidence: PMID:18275604 There is no evidence for S100 proteins in invertebrates PMID:27764152 All S100 hits were within vertebrates, with the exception of four hits from the tunicate Oikopleura dioica PMID:31837246 The calcium-binding, vertebrate-specific S100 protein family file:DROME/CG3226/CG3226-bioinformatics/RESULTS.md CG3226 has no S100 protein to bind in D. melanogaster |
| GO:0060090 molecular adaptor activity | IBA GO_REF:0000033 | ACCEPT | Summary: Mammalian SIP bridges Siah to SKP1/Ebi, scaffolding substrate and E2 in the E3 complex; this adaptor role is the family's core molecular function and is reasonably inherited by CG3226, which conserves the N-terminal Siah-binding and C-terminal SGS/CS regions. Reason: Core conserved function of the CacyBP/SIP family; fly-specific biochemical confirmation is lacking. Supporting Evidence: PMID:16085652 this surface provides the scaffold for bringing substrate and the E2 enzyme into apposition in the functional complex PMID:11389839 Siah is physically linked to Ebi by association with a novel Sgt1 homolog SIP that binds Skp1 |
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Download this section (compressed HTML)Q: Does CG3226 bind Sina and/or Sinah and SkpA in vivo, and does it promote ubiquitination of Armadillo or other substrates such as Tramtrack?
Q: Is the cardiomyopathy of CG3226 loss a developmental defect or a failure of adult cardiac maintenance/contractility, and is it mediated through Armadillo?
Q: Should GO:0044548 S100 protein binding carry a taxon constraint (only in Chordata), or the IPR037893 InterPro2GO mapping be restricted, so that it is not propagated to invertebrate CacyBP/SIP orthologs?
Q: In the absence of S100 proteins, is the C-terminal SGS region of fly CacyBP/SIP regulated by another calcium sensor, or is calcium regulation a vertebrate innovation?
Experiment: Co-immunoprecipitation or yeast two-hybrid of CG3226 with Sina, Sinah, SkpA and Ebi, and in vivo ubiquitination assays of Armadillo in CG3226 mutant tissue.
Experiment: Temporally restricted (adult-only versus developmental) heart-specific RNAi or rescue of CG3226 with OCT phenotyping, to separate developmental from maintenance roles.
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