id: F2UDK1 gene_symbol: yorkie aliases: - PTSG_06057 - Yorkie - Yki - srYki - YAP/TAZ homolog product_type: PROTEIN status: DRAFT taxon: id: NCBITaxon:946362 label: Salpingoeca rosetta description: >- yorkie (PTSG_06057) encodes the Yorkie/YAP/TAZ homolog of the colonial choanoflagellate Salpingoeca rosetta, a 714-residue protein with two WW domains (residues 189-222 and 244-277) embedded in long disordered, low-complexity regions. In animals Yorkie/YAP is a transcriptional coactivator: it binds Scalloped/TEAD transcription factors through an N-terminal TEAD-binding domain, its WW domains bind PPxY motifs of partner proteins, and phosphorylation by Warts/LATS on HXRXXS motifs keeps it in the cytoplasm. The S. rosetta protein carries four HXRXXS motifs, one of them N-terminal to the WW domains as in YAP, but its predicted TEAD-binding region is poorly conserved, and whether it binds a TEAD protein or acts as a coactivator is unknown. yorkie knockout cells proliferate at the wild-type rate and form rosettes of normal size, whereas Warts and Yorkie both influence expression of extracellular matrix genes, including couscous. existing_annotations: - term: id: GO:0005737 label: cytoplasm evidence_type: IEA original_reference_id: GO_REF:0000118 qualifier: located_in supporting_entities: - PANTHER:PTN002569196 review: summary: >- TreeGrafter transfer from PANTHER node PTN002569196. TreeGrafter places F2UDK1 in PTHR10316:SF68 (family "MEMBRANE ASSOCIATED GUANYLATE KINASE-RELATED"), and the node is an ecdysozoan speciation node whose members are Drosophila CG42788, Anopheles AGAP003128 and a Pristionchus gene (PANTHER 19 tree), not the YAP1 family (PTHR17616) of human YAP1 and Drosophila Yorkie. Our own analysis finds that F2UDK1 matches YAP1, Yorkie and Capsaspora coYki better than the S. rosetta protein that PANTHER does place in PTHR17616 (F2U5K0, PTSG_03848). The localization of S. rosetta Yorkie has not been examined. action: KEEP_AS_NON_CORE reason: >- The propagation route is wrong: the term reaches a choanoflagellate Yorkie homolog from a node of insect and nematode MAGI-related proteins. The term itself is plausible on independent grounds, since Yorkie/YAP proteins shuttle between cytoplasm and nucleus and are held in the cytoplasm when phosphorylated by Warts/LATS, and F2UDK1 carries four HXRXXS motifs. Kept as a non-core, unverified location, and the nucleus should be tested alongside it. supported_by: - reference_id: file:SALRS/yorkie/yorkie-uniprot.txt record_section: ["DR:PANTHER"] supporting_text: PANTHER; PTHR10316:SF68; AGAP003128-PA; 1. - reference_id: file:SALRS/yorkie/yorkie-bioinformatics/RESULTS.md supporting_text: >- F2UDK1 is the better match to all three Yorkie/YAP proteins. - reference_id: PMID:38729842 supporting_text: srYki has 4 putative Warts phosphorylation sites propagation_review: root_cause: PROPAGATION_BAD failure_modes: - WRONG_ORTHOLOG_OR_PARALOG source_entities: - source_id: PANTHER:PTN002569196 source_label: Ecdysozoa node, PTHR10316:SF68 source_status: SUPPORTS_SOURCE_BUT_NOT_TARGET comment: >- Node of insect and nematode MAGI-related WW/PDZ proteins; F2UDK1 is a Yorkie/YAP homolog misgrafted into this family. The term happens to be plausible for a Yorkie homolog anyway. - term: id: GO:0007165 label: signal transduction evidence_type: IEA original_reference_id: GO_REF:0000118 qualifier: involved_in supporting_entities: - PANTHER:PTN002569196 review: summary: >- TreeGrafter transfer from the same misplaced ecdysozoan node (PTN002569196) in the MAGI-related family. As a Yorkie homolog, F2UDK1 is expected to be the transcriptional effector of hippo signaling. In S. rosetta, Warts and Yorkie both regulate extracellular matrix genes [PMID:41037400], but yorkie knockout has no effect on proliferation or rosette size [DOI:10.1101/2024.07.13.603360], and no experiment shows Yorkie responding to Warts. action: KEEP_AS_NON_CORE reason: >- The term is broad, and is plausible for a Yorkie homolog through its own family. A shared set of extracellular-matrix target genes for Warts and Yorkie fits a Warts-to-Yorkie pathway, but the phenotypes do not follow the animal pattern (warts loss enlarges rosettes, yet yorkie loss has no size effect), so no specific S. rosetta signal or downstream process can be named. Kept as non-core because it rests on a wrong-family propagation and on family-level inference, not on S. rosetta data. Hippo signaling is not proposed as NEW for the same reason, and because the predicted TEAD-binding region needed for the canonical output is not conserved. supported_by: - reference_id: PMID:41037400 supporting_text: >- RNA sequencing revealed that Warts and Yorkie regulated several extracellular matrix genes involved in multicellularity (including couscous) - reference_id: DOI:10.1101/2024.07.13.603360 supporting_text: >- Growth curves indicated that yorkiepac1 KO cells proliferated at a similar rate to the wild type - reference_id: DOI:10.1101/2024.07.13.603360 supporting_text: >- The size of hippopac1 and yorkiepac1 rosettes did not significantly differ from wild type (11.6 ± 1.7 cells and 9.8 ± 3.1 cells propagation_review: root_cause: PROPAGATION_BAD failure_modes: - WRONG_ORTHOLOG_OR_PARALOG source_entities: - source_id: PANTHER:PTN002569196 source_label: Ecdysozoa node, PTHR10316:SF68 source_status: SUPPORTS_SOURCE_BUT_NOT_TARGET comment: >- Wrong family for a Yorkie homolog; the generic signalling term survives only because it is also plausible for Yorkie/YAP. core_functions: - description: >- Yorkie/YAP/TAZ-family WW-domain protein, predicted by homology to be the transcriptional effector restrained by Warts in the S. rosetta Hippo pathway. It carries four Warts-consensus HXRXXS motifs, but its predicted TEAD-binding region is poorly conserved, so its partner transcription factor and coactivator activity are unknown. In S. rosetta it shares extracellular-matrix target genes, including couscous, with Warts, but is not required for normal proliferation or rosette size. No molecular function term is asserted until coactivator activity or TEAD binding is shown. supported_by: - reference_id: PMID:38729842 supporting_text: srYki has 4 putative Warts phosphorylation sites - reference_id: PMID:38729842 supporting_text: >- The degree to which choanoflagellate Yorkie orthologs interact with Sd/TEAD proteins is therefore currently unclear - reference_id: PMID:41037400 supporting_text: >- RNA sequencing revealed that Warts and Yorkie regulated several extracellular matrix genes involved in multicellularity (including couscous) suggested_questions: - question: >- Does S. rosetta Yorkie bind the S. rosetta Scalloped/TEAD homolog despite the poorly conserved TEAD-binding region, and does it activate transcription in a reporter assay? - question: >- Is S. rosetta Yorkie phosphorylated by Warts on its four HXRXXS motifs, and does Warts loss shift it to the nucleus? - question: >- Do Warts and Yorkie regulate the shared extracellular matrix genes (e.g. couscous) in opposite directions, as the canonical pathway predicts? - question: >- Is F2UDK1 the Yorkie ortholog, given that PANTHER places it with MAGI-related proteins and puts another S. rosetta WW protein (F2U5K0, PTSG_03848) in the YAP1 family? A phylogeny of holozoan YAP/Yorkie and WW-domain proteins would settle it. - question: >- Why does yorkie knockout not affect rosette size or proliferation? Is there redundancy with another WW-domain coactivator? suggested_experiments: - hypothesis: S. rosetta Yorkie is a Warts-regulated nucleocytoplasmic protein. description: >- Express fluorescently tagged wild-type and four-site S-to-A Yorkie in wild-type and warts knockout cells and score its nuclear-to-cytoplasmic ratio. experiment_type: localization / imaging - hypothesis: S. rosetta Yorkie binds the S. rosetta TEAD homolog. description: >- Co-immunoprecipitation or yeast two-hybrid of Yorkie with the S. rosetta Scalloped/TEAD homolog, with AlphaFold-Multimer modelling to locate any divergent interface. experiment_type: protein interaction assay - hypothesis: The enlarged rosettes of warts mutants require Yorkie. description: >- Construct warts yorkie double knockouts and count cells per rosette and imaging of the ECM core. experiment_type: genetic epistasis references: - id: GO_REF:0000118 title: TreeGrafter-generated GO annotations findings: [] - id: file:SALRS/yorkie/yorkie-uniprot.txt title: UniProt record for yorkie (F2UDK1) findings: - statement: PANTHER places F2UDK1 in a MAGI-related subfamily, not the YAP1 family. record_section: ["DR:PANTHER"] supporting_text: PANTHER; PTHR10316:SF68; AGAP003128-PA; 1. - id: file:SALRS/yorkie/yorkie-bioinformatics/RESULTS.md title: F2UDK1 motif and identity checks (reviewer analysis) reference_review: relevance: HIGH correctness: VERIFIED review_notes: >- Re-read the local motif and identity report; it supports the F2UDK1 Yorkie assignment, Warts/LATS motif count and absent recognizable TEAD-binding motifs while leaving a full phylogeny and divergent TEAD binding unresolved. findings: - statement: F2UDK1 has four HXRXXS motifs. supporting_text: >- F2UDK1 (714 aa) carries four HXRXXS motifs - statement: No recognizable TEAD-binding motif was found in F2UDK1. supporting_text: >- We find **no recognizable TEAD-binding motif** in F2UDK1. - statement: F2UDK1 resembles YAP1, Yorkie and coYki more than F2U5K0 does. supporting_text: F2UDK1 is the better match to all three Yorkie/YAP proteins. - id: DOI:10.1101/2024.07.13.603360 title: A fast and robust gene knockout method for Salpingoeca rosetta informs the genetics of choanoflagellate multicellular development reference_review: relevance: HIGH correctness: VERIFIED review_notes: >- bioRxiv preprint (July 2024), full text cached from the PDF. Reports CRISPR knockout of the S. rosetta Hippo, Warts and Yorkie homologs (two clones each). The cached version does not give locus IDs; PTSG_06057 as the yorkie locus was supplied from the preprint's gRNA-design sentence, which is not in this cached version. F2UDK1's two WW domains and four HXRXXS motifs match the published description of srYki, which supports the mapping. findings: - statement: yorkie knockout cells proliferate at a wild-type rate. supporting_text: >- Growth curves indicated that yorkiepac1 KO cells proliferated at a similar rate to the wild type - statement: yorkie knockout rosettes are of normal size. supporting_text: >- The size of hippopac1 and yorkiepac1 rosettes did not significantly differ from wild type (11.6 ± 1.7 cells and 9.8 ± 3.1 cells - statement: All knockout clones still develop into rosettes. supporting_text: >- all six KO clones reliably developed into rosettes, as did wild-type cells - id: PMID:41037400 title: A selection-based knockout approach for a choanoflagellate reveals regulation of multicellular development by Hippo signaling. full_text_unavailable: true reference_review: relevance: HIGH correctness: VERIFIED review_notes: >- Published version of the preprint; abstract only cached. Adds RNA-seq showing that Warts and Yorkie regulate extracellular matrix genes including couscous. findings: - statement: Warts and Yorkie regulate extracellular matrix genes, including couscous. supporting_text: >- RNA sequencing revealed that Warts and Yorkie regulated several extracellular matrix genes involved in multicellularity (including couscous) - id: PMID:38729842 title: 'Assembling a Hippo: the evolutionary emergence of an animal developmental signaling pathway.' reference_review: relevance: HIGH correctness: VERIFIED review_notes: >- Review with the only published description of S. rosetta Yorkie (srYki) domain architecture, Warts motifs and TEAD-binding-domain conservation. findings: - statement: S. rosetta Yorkie has two WW domains and four putative Warts sites. supporting_text: >- like most animals, srYki has 2 WW domains, whereas mbYki has 1, and srYki has 4 putative Warts phosphorylation sites - statement: The srYki TEAD-binding domain lacks conserved alpha2-helix residues. supporting_text: >- the srYki TBD shows less conservation with Yorkie/YAP/TAZ than coYki, with srYki lacking conserved residues within the α2 helix that are critical for YAP-TEAD interaction in mammals - statement: Whether choanoflagellate Yorkie homologs bind Sd/TEAD is unclear. supporting_text: >- The degree to which choanoflagellate Yorkie orthologs interact with Sd/TEAD proteins is therefore currently unclear - id: PMID:22832104 title: Premetazoan origin of the hippo signaling pathway. reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: >- Identifies Yki homologs in S. rosetta and M. brevicollis that cluster with metazoan Yki. Its claim that all non-metazoan Yki homologs keep the N-terminal TEAD-interaction region is not borne out for F2UDK1 by the later review (PMID:38729842) or by our motif search; the 2012 analysis may have used a different gene model. findings: - statement: S. rosetta has a Yorkie homolog that clusters with metazoan Yorkie. supporting_text: >- We have identified clear Yki homologs in two independent non-metazoan lineages, the filastereans (Capsaspora owczarzaki, from herein called “Capsaspora”) and the choanoflagellates (Monosiga brevicollis and Salpingoeca rosetta) (Figure 1B). - statement: >- All non-metazoan Yki homologs, including S. rosetta's, keep the N-terminal TEAD-interaction region. supporting_text: >- Importantly, all these non-metazoan Yki homologs contain highly conserved functional sites like the Hippo pathway responsive phosphorylation site S168/127 and the N-terminal homology region that is critical for interaction with Sd/TEAD transcription factor (Figure 2A). finding_review: finding_status: DISPUTED review_notes: >- The later review reports that the srYki TEAD-binding domain lacks conserved alpha2-helix residues, and our motif search finds no YAP/Yki TEAD-interface motif in F2UDK1. supported_by: - reference_id: PMID:38729842 supporting_text: >- the srYki TBD shows less conservation with Yorkie/YAP/TAZ than coYki, with srYki lacking conserved residues within the α2 helix that are critical for YAP-TEAD interaction in mammals