id: A0A9L0RQI4 gene_symbol: AFAP1L2 product_type: PROTEIN status: IN_PROGRESS taxon: id: NCBITaxon:9796 label: Equus caballus description: AFAP1L2 (XB130, PI3KAP) is a cytoplasmic adaptor and scaffold protein of the AFAP family, built from an N-terminal region carrying SH2- and SH3-binding motifs, two pleckstrin homology domains, a coiled-coil region and a short C-terminal actin-binding segment. It binds the Src tyrosine kinase and enhances its activity, and once tyrosine-phosphorylated by Src or RET it docks the SH2 domains of the p85 regulatory subunit of phosphatidylinositol 3-kinase, coupling tyrosine kinase signals to PI3K-Akt signalling, cell-cycle progression and survival. Through its C-terminal segment it binds and crosslinks actin filaments and is recruited to lamellipodial F-actin, where it supports cell spreading and motility. In mammals it is most abundant in thyroid epithelium. The horse product is an Ensembl-predicted transcript model of the AFAP1L2 gene that retains every characterized human domain and motif. existing_annotations: - term: id: GO:0005737 label: cytoplasm evidence_type: IEA original_reference_id: GO_REF:0000044 qualifier: located_in supporting_entities: - UniProtKB-SubCell:SL-0086 review: action: ACCEPT summary: AFAP1L2 is a cytoplasmic protein, partitioning between the cytosol and cortical/lamellipodial F-actin; cytoplasm is the right level to cover both pools. reason: Human XB130 is cytoplasmic by immunofluorescence (PMID:17412687, the experimental basis of the human UniProt subcellular-location statement) and is described as a cytosolic adaptor that translocates to lamellipodial F-actin on stimulation (PMID:21084565); the mouse ortholog localizes to actin stress fibres and is diffuse in the cytosol when the actin-binding region is deleted (PMID:27462298). The horse model sits in the same PANTHER subfamily (PTHR14338:SF4) as human and mouse AFAP1L2 and retains all characterized regions, including the C-terminal actin-binding segment (file:HORSE/AFAP1L2/AFAP1L2-bioinformatics/RESULTS-rereview.md). The ARBA-derived horse UniProt statement is not itself used as evidence. The more specific cytosol term from the PAINT node is carried in GOA by another horse AFAP1L2 product (A0A3Q2HBW2) and is recorded in core_functions; cytoplasm remains accurate for the actin-associated pool as well, so it is kept rather than refined. supported_by: - reference_id: file:human/AFAP1L2/AFAP1L2-uniprot.txt record_section: ["CC:SUBCELLULAR LOCATION"] supporting_text: 'CC -!- SUBCELLULAR LOCATION: Cytoplasm {ECO:0000269|PubMed:17412687}.' - reference_id: PMID:21084565 supporting_text: XB130 is a newly described cytosolic adaptor protein and tyrosine kinase substrate - reference_id: PMID:21084565 supporting_text: In stimulated cells, XB130 localized to the lamellipodial F-actin meshwork. - reference_id: file:HORSE/AFAP1L2/AFAP1L2-uniprot.txt record_section: ["DR:PANTHER"] supporting_text: DR PANTHER; PTHR14338:SF4; ACTIN FILAMENT-ASSOCIATED PROTEIN 1-LIKE 2; 1. - term: id: GO:0036312 label: phosphatidylinositol 3-kinase regulatory subunit binding evidence_type: ISS original_reference_id: PMID:19060924 qualifier: enables review: action: NEW summary: Tyrosine-phosphorylated AFAP1L2 docks the SH2 domains of the PI3K p85 regulatory subunit, linking Src/RET activity to PI3K-Akt signalling. reason: In human cells RET/PTC phosphorylates XB130 Tyr54, which is a critical binding site for the p85alpha SH2 domains, and XB130 knockdown lowers Akt activity (PMID:19060924). The rat ortholog PI3KAP associates with p85 and with PI3K activity in a Src-dependent manner, and a Y72F mutant that cannot bind p85 fails to enhance IGF-I-induced DNA synthesis (PMID:22496359). Both proteins are in the AFAP1L2 subfamily (PTHR14338:SF4) with the horse model; horse Tyr65 is the conserved equivalent of human Tyr54. This is the adaptor's informative binding activity; no GO annotation currently records it in any species, so this is a curation gap rather than a PAINT omission of an existing experimental term. Evidence is ISS from the human and rat experimental work. supported_by: - reference_id: PMID:19060924 supporting_text: We identified tyrosine 54 of XB130 as the major target of RET/PTC-mediated phosphorylation and a critical binding site for the SH2 domains of p85alpha. - reference_id: PMID:22496359 supporting_text: a mutant Y72F incapable of binding to p85 PI3K did not show this response - reference_id: file:HORSE/AFAP1L2/AFAP1L2-bioinformatics/RESULTS-rereview.md supporting_text: '| Tyr54 (p85 SH2-binding phosphosite) | 65-65 | 1/1 | 1/1 |' - term: id: GO:0051015 label: actin filament binding evidence_type: ISS original_reference_id: PMID:27462298 qualifier: enables review: action: NEW summary: The C-terminal segment of AFAP1L2 binds actin filaments directly, and the full-length multimeric protein crosslinks F-actin in vitro. reason: Recombinant mouse PI3KAP/XB130 fragments containing residues 830-840 co-sedimented with actin filaments, and full-length protein crosslinked F-actin in a manner requiring the N-terminal multimerizing region and this C-terminal actin-binding region (PMID:27462298). The authors note the segment is conserved across vertebrate orthologs. Human XB130 likewise requires its C-terminal 63 residues to reach lamellipodial F-actin (PMID:21084565). Mouse Q5DTU0 and horse A0A9L0RQI4 are both in PTHR14338:SF4; the horse C-terminal 63-residue segment pairs fully with human (57/63 identical) and ends in the same terminal residues. This activity is direct biochemistry of the ortholog, transferred as ISS. supported_by: - reference_id: PMID:27462298 supporting_text: These results indicated that PI3KAP/XB130 bound to actin filaments through amino acid residues 830–840 and that this region needed to be intact. - reference_id: PMID:27462298 supporting_text: These results indicated that PI3KAP/XB130 could crosslink actin filaments in vitro. - reference_id: file:HORSE/AFAP1L2/AFAP1L2-bioinformatics/RESULTS-rereview.md supporting_text: '| C-terminal 63 aa (756-818) | 788-850 | 63/63 | 57/63 |' - term: id: GO:0030027 label: lamellipodium evidence_type: ISS original_reference_id: PMID:21084565 qualifier: located_in review: action: NEW summary: On stimuli that induce lamellipodia, AFAP1L2 translocates to the branched F-actin meshwork of the lamellipodium. reason: Human XB130 (same PANTHER subfamily PTHR14338:SF4 as the horse model) moves to the cell periphery after EGF, wounding or active Rac and localizes to lamellipodial F-actin; the N-terminal 167 residues and C-terminal 63 residues are required, while the PH domains are dispensable (PMID:21084565). Both required regions are fully present in the horse model. This is a stimulus-dependent location of the protein itself, distinct from the cytosolic pool, and is not a downstream phenotype. supported_by: - reference_id: PMID:21084565 supporting_text: In stimulated cells, XB130 localized to the lamellipodial F-actin meshwork. - reference_id: PMID:21084565 supporting_text: both the XB130 N-terminus (167 amino acids) and C-terminus (63 amino acids) harbor crucial regions for its translocation to lamellipodia - reference_id: file:HORSE/AFAP1L2/AFAP1L2-bioinformatics/RESULTS-rereview.md supporting_text: '| C-terminal 63 aa (756-818) | 788-850 | 63/63 | 57/63 |' references: - id: GO_REF:0000044 title: Gene Ontology annotation based on UniProtKB/Swiss-Prot Subcellular Location vocabulary mapping, accompanied by conservative changes to GO terms applied by UniProt findings: [] - id: GO_REF:0000033 title: Annotation inferences using phylogenetic trees findings: [] - id: file:human/AFAP1L2/AFAP1L2-uniprot.txt title: human/AFAP1L2/AFAP1L2-uniprot.txt findings: [] reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: Reviewed human Q8N4X5 record; its subcellular location, function and subunit statements carry ECO:0000269 evidence from PMID:17412687 and PMID:18844995. - id: file:HORSE/AFAP1L2/AFAP1L2-uniprot.txt title: HORSE/AFAP1L2/AFAP1L2-uniprot.txt findings: [] reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: Unreviewed TrEMBL record (Ensembl ENSECAT00000130227.1). Used only for family/domain classification (PANTHER SF4, InterPro AFAP, two PH domains); its ARBA function, subunit and location text is not used as evidence. - id: file:HORSE/AFAP1L2/AFAP1L2-bioinformatics/RESULTS.md title: HORSE/AFAP1L2/AFAP1L2-bioinformatics/RESULTS.md findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: >- Local human-horse sequence comparison inspected; supports conserved AFAP1L2 architecture because the horse record aligns across the full human sequence at 88.6% identity, while explicitly leaving orthology, gene-model completeness and transfer of each biological claim to the primary review. 2026-10-20 re-review: no longer cited as support; superseded by RESULTS-rereview.md, which adds the per-feature mapping and the comparison with the other nine horse gene models. Retained for provenance. - id: file:HORSE/AFAP1L2/AFAP1L2-bioinformatics/RESULTS-rereview.md title: HORSE/AFAP1L2/AFAP1L2-bioinformatics/RESULTS-rereview.md findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: Generated by compare_sequences_rereview.py from local UniProt records; maps human features (Tyr54, PH1, PH2, coiled coil, C-terminal 63 aa) onto the horse model and compares all ten horse AFAP1L2 transcript models. - id: file:interpro/panther/PTHR14338/PTHR14338-paint.tsv title: interpro/panther/PTHR14338/PTHR14338-paint.tsv findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: PAINT IBD assertions for PTHR14338; PTN002682769 (AFAP1L2 clade, taxon 117571) carries GO:0030296 and four process terms, PTN001023816 (taxon 33511) carries GO:0005829, all seeded by human Q8N4X5. - id: PMID:17412687 title: XB130, a novel adaptor protein for signal transduction. full_text_unavailable: true findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: Primary human XB130 characterization (abstract cached); Src interaction and activation, siRNA reduction of Src activity, IL-8 and EGF-induced Akt phosphorylation. - id: PMID:19060924 title: XB130, a tissue-specific adaptor protein that couples the RET/PTC oncogenic kinase to PI 3-kinase pathway. findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: Abstract-only cache; identifies Tyr54 as the p85alpha SH2-binding phosphosite and shows Akt dependence in thyroid cancer cells. - id: PMID:22496359 title: Phosphatidylinositol 3-kinase-binding protein, PI3KAP/XB130, is required for cAMP-induced amplification of IGF mitogenic activity in FRTL-5 thyroid cells. findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: Rat ortholog (PI3KAP); abstract-only cache; p85 binding, Src association and Y72F p85-binding mutant. - id: PMID:21084565 title: Adaptor protein XB130 is a Rac-controlled component of lamellipodia that regulates cell motility and invasion. findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: Abstract-only cache; human XB130 translocation to lamellipodial F-actin requiring N- and C-terminal regions, and knockdown effects on motility. - id: PMID:27462298 title: Phosphatidylinositol 3-Kinase-Associated Protein (PI3KAP)/XB130 Crosslinks Actin Filaments through Its Actin Binding and Multimerization Properties In Vitro and Enhances Endocytosis in HEK293 Cells. findings: [] reference_review: relevance: HIGH correctness: VERIFIED review_notes: Full text cached; mouse construct; F-actin co-sedimentation mapping to residues 830-840, in vitro crosslinking, N-terminal multimerization. - id: PMID:34470464 title: XB130 Deficiency Causes Congenital Hypothyroidism in Mice due to Disorganized Apical Membrane Structure and Function of Thyrocytes. findings: [] reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: Knockout phenotype (thyroid); organismal phenotype used as context only, not as process annotation. - id: PMID:26446840 title: XB130/Tks5 scaffold protein interaction regulates Src-mediated cell proliferation and survival. findings: [] reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: Human XB130 scaffold interaction with Tks5 in Src signalling; context for adaptor role, not used for a specific annotation. core_functions: - description: Binds c-Src via its N-terminal SH2/SH3-binding motifs and enhances Src kinase activity in the cytosol, promoting Src-dependent tyrosine phosphorylation and downstream transcriptional responses. molecular_function: id: GO:0030296 label: protein tyrosine kinase activator activity locations: - id: GO:0005829 label: cytosol supported_by: - reference_id: PMID:17412687 supporting_text: Their co-expression in COS-7 cells resulted in activation of c-Src and elevated tyrosine phosphorylation of multiple proteins, including XB130 itself. - reference_id: file:interpro/panther/PTHR14338/PTHR14338-paint.tsv supporting_text: PTN002682769 - description: Phosphotyrosine-dependent adaptor that, after phosphorylation by Src or RET, recruits the p85 regulatory subunit of PI3K and so couples tyrosine kinase signalling to PI3K-Akt activation. molecular_function: id: GO:0036312 label: phosphatidylinositol 3-kinase regulatory subunit binding locations: - id: GO:0005829 label: cytosol supported_by: - reference_id: PMID:19060924 supporting_text: We identified tyrosine 54 of XB130 as the major target of RET/PTC-mediated phosphorylation and a critical binding site for the SH2 domains of p85alpha. - reference_id: PMID:22496359 supporting_text: a mutant Y72F incapable of binding to p85 PI3K did not show this response - description: Binds and, as a multimer, crosslinks actin filaments through a C-terminal actin-binding segment, associating with lamellipodial F-actin. molecular_function: id: GO:0051015 label: actin filament binding locations: - id: GO:0030027 label: lamellipodium supported_by: - reference_id: PMID:27462298 supporting_text: These results indicated that PI3KAP/XB130 could crosslink actin filaments in vitro. - reference_id: PMID:21084565 supporting_text: In stimulated cells, XB130 localized to the lamellipodial F-actin meshwork. suggested_questions: - question: Which of the ten horse AFAP1L2 TrEMBL transcript models is the principal product, and is the 20-residue coiled-coil insertion and alternative N-terminal exon of A0A9L0RQI4 expressed in horse thyroid? - question: Should the GO record for AFAP1L2 orthologs capture the p85-binding adaptor activity (PI3K regulatory subunit binding) and actin filament binding, which are experimentally shown in human and rodent but currently unannotated in any species? suggested_experiments: - description: RT-PCR or long-read isoform sequencing of horse thyroid and lung to establish which AFAP1L2 5' exon and whether the coiled-coil cassette exon are used. experiment_type: transcript isoform profiling