{"gene":"ZP4","run_date":"2026-04-28T23:00:24","timeline":{"discoveries":[{"year":1994,"finding":"Porcine zona pellucida glycoprotein ZP4 amino acid sequence was determined by peptide mapping and cDNA cloning, revealing two peptide isoforms (128 and 133 amino acids) with two potential N-linked glycosylation sites, five conserved cysteine residues, and sequence similarity to mouse ZP2; ZP4 and ZP2 appear to be derived from a common parent polypeptide by proteolytic cleavage.","method":"Peptide mapping, cDNA cloning, sequence analysis","journal":"Journal of reproduction and fertility","confidence":"Medium","confidence_rationale":"Tier 1 — direct biochemical sequencing and cDNA cloning, single lab","pmids":["8182597"],"is_preprint":false},{"year":1998,"finding":"Porcine ZPB (ortholog of mouse ZP1) and ZPC (ortholog of mouse ZP3) form hetero-oligomeric complexes that bind to boar sperm membrane vesicles with high affinity; neither free ZPB nor free ZPC alone binds sperm membranes, demonstrating that heterocomplex formation is required for sperm-zona binding activity.","method":"Solid phase binding assays, size exclusion chromatography, Western blot, biotin-labeling competition assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 — multiple orthogonal methods (binding assay, chromatography, Western blot) in a single rigorous study","pmids":["9516448"],"is_preprint":false},{"year":2003,"finding":"Porcine ZPB and ZPC C-terminal residues were identified (Ala-462 and Ser-332, respectively) by mass spectrometry, revealing that ZPB is processed at its furin consensus site whereas ZPC is processed N-terminal to the furin consensus site; disulfide bond analysis within the ZP domains suggested the ZP domain consists of two subdomains.","method":"Mass spectrometry of C-terminal polypeptide fragments","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 1 — direct mass spectrometry characterization, single lab","pmids":["12878193"],"is_preprint":false},{"year":2007,"finding":"Recombinant bovine ZP3 and ZP4 co-expressed in Sf9 insect cells form a hetero-complex that inhibits bovine sperm-ZP binding; neither ZP3 nor ZP4 alone shows inhibitory activity. The N-terminal and trefoil domains of ZP4 are dispensable for formation of the sperm-binding active ZP3/ZP4 complex, and the molecular interactions between ZP3 and ZP4 are conserved between bovine and porcine systems.","method":"Baculovirus-Sf9 cell expression, sperm-ZP competitive inhibition assay, immunoprecipitation, deletion mutagenesis","journal":"The FEBS journal","confidence":"High","confidence_rationale":"Tier 1 — reconstitution with deletion mutagenesis, functional assay, and immunoprecipitation in a single study","pmids":["17894824"],"is_preprint":false},{"year":2010,"finding":"Human ZP4 expressed in transgenic mice is incorporated into the zona pellucida matrix with molecular mass similar to native isoforms, but is not sufficient to confer human sperm binding to mouse eggs, indicating that additional zona proteins are required for species-selective human gamete recognition.","method":"Transgenic mouse model, sperm binding assay, molecular mass analysis","journal":"Reproduction (Cambridge, England)","confidence":"High","confidence_rationale":"Tier 2 — clean transgenic KI model with defined functional readout (sperm binding) and protein incorporation confirmed","pmids":["21173071"],"is_preprint":false},{"year":2011,"finding":"In porcine ZP, native ZP4 (nZP4) — but not recombinant ZP4 — is necessary for the sperm-binding activity of the ZP3/ZP4 complex toward porcine sperm, demonstrating that carbohydrate structures on ZP4 are responsible for porcine sperm-binding activity of the complex; ZP3 and ZP4 reciprocally co-precipitate each other via recombinant N-terminal tags.","method":"Competitive sperm-binding inhibition assay, co-immunoprecipitation, baculovirus-Sf9 expression","journal":"Zygote (Cambridge, England)","confidence":"High","confidence_rationale":"Tier 1-2 — functional reconstitution with native vs. recombinant protein comparison and reciprocal co-precipitation","pmids":["22008510"],"is_preprint":false},{"year":2015,"finding":"The hinge region (Arg-32 to Arg-160) of bovine ZP3 is a ZP4-binding site; N-glycosylation at Asn-146 (but not Asn-124) in this region is required for the sperm-binding activity of the ZP3/ZP4 complex, although it does not prevent ZP4 interaction per se.","method":"Baculovirus-Sf9 expression, ZP3 deletion and point mutants, co-expression, sperm-ZP binding inhibition assay","journal":"Biomolecules","confidence":"High","confidence_rationale":"Tier 1 — mutagenesis combined with functional reconstitution assay mapping the ZP4-binding site on ZP3","pmids":["26610590"],"is_preprint":false},{"year":2019,"finding":"ZP4 ablation in rabbits (CRISPR KO) produced a zona pellucida that was significantly thinner, more permeable, and more disorganized/fenestrated; ovulation, fertilization, and in vitro development were unaffected, but in vivo preimplantation development was severely impaired, demonstrating that ZP4 is required for structural integrity of the zona pellucida necessary for embryo protection in vivo.","method":"CRISPR-Cas9 knockout rabbit model, electron microscopy of ZP structure, permeability assays, in vivo/in vitro fertility assays","journal":"eLife","confidence":"High","confidence_rationale":"Tier 2 — clean KO with multiple orthogonal phenotypic readouts (structural, permeability, developmental) and rigorous controls","pmids":["31635692"],"is_preprint":false},{"year":2019,"finding":"ZP4 is expressed in immature human fetal enterocytes and is regulated by Toll-like receptor 2 and 4 signaling; knockdown of ZP4 in fetal intestinal cells altered IL-8 mRNA expression in response to IL-1β, implicating ZP4 in innate immune signaling in the developing intestine.","method":"Transcription profiling, ZP4 knockdown in H4 fetal intestinal cell line, IL-8 mRNA measurement","journal":"American journal of physiology. Gastrointestinal and liver physiology","confidence":"Low","confidence_rationale":"Tier 3 — single knockdown experiment in a cell line, limited mechanistic follow-up","pmids":["31314571"],"is_preprint":false},{"year":2021,"finding":"Zp4 knockout female rats (CRISPR-Cas9) showed normal ZP morphology and were completely fertile, with no defects in oogenesis, ovulation, fertilization, or pup production, demonstrating that Zp4 is fully dispensable for female fertility in rats.","method":"CRISPR-Cas9 KO rat model, PAS staining, superovulation, in vitro fertilization, natural mating","journal":"Biology of reproduction","confidence":"High","confidence_rationale":"Tier 2 — clean KO model with multiple orthogonal reproductive phenotype readouts","pmids":["33709118"],"is_preprint":false},{"year":2021,"finding":"A mutant ZP1 protein in rats is intracellularly sequestered and interacts with wild-type ZP3 or ZP4 (shown in 293T cells), impeding incorporation of ZP3 and ZP4 into the zona pellucida, resulting in zona absence and female infertility.","method":"Homologous rat KI model, 293T cell co-expression, co-immunoprecipitation, immunofluorescence","journal":"Biology of reproduction","confidence":"Medium","confidence_rationale":"Tier 2 — reciprocal co-IP in cells, KI animal model; single lab","pmids":["33624742"],"is_preprint":false},{"year":2021,"finding":"The N-terminal region (Lys-25 to Asp-136) and middle region (Ser-290 to Lys-340) of bovine ZP4 contain sperm-binding sites, as demonstrated by binding of sperm to ZP4-coated plates and systematic deletion mutagenesis; ZP4 harbors the major potential sperm-binding sites among bovine ZP glycoproteins, but multivalent complex formation is required for full activity.","method":"ZP4-coated plate solid-phase binding assay, deletion mutagenesis, Sf9-expressed recombinant proteins","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 — systematic deletion mapping with functional readout, single lab","pmids":["34242308"],"is_preprint":false},{"year":2021,"finding":"ZP4 mutations identified in human patients (whole-exome sequencing) reduced ZP4 secretion in vitro when mutant plasmids were transfected into cells, linking ZP4 loss-of-function to abnormal (thin and irregular) zona pellucida and female infertility.","method":"Whole-exome sequencing, in vitro plasmid expression/secretion assay in transfected cells","journal":"Journal of clinical pathology","confidence":"Medium","confidence_rationale":"Tier 2-3 — human genetic evidence combined with in vitro secretion assay; mechanistic follow-up limited to secretion","pmids":["33461974"],"is_preprint":false}],"current_model":"ZP4 is a zona pellucida glycoprotein that forms a functional heterocomplex with ZP3 (requiring ZP4's carbohydrate moieties) to mediate sperm binding to the zona pellucida; it confers structural integrity (thickness, permeability, organization) to the zona matrix essential for in vivo embryo protection, though its requirement for fertility is species-dependent (indispensable in rabbits and humans, dispensable in rats and mice)."},"narrative":{"teleology":[{"year":1994,"claim":"Determining ZP4's primary structure established it as a zona pellucida glycoprotein with conserved cysteine residues and N-glycosylation sites, related to ZP2 via proteolytic processing of a common precursor.","evidence":"Peptide mapping and cDNA cloning of porcine ZP4","pmids":["8182597"],"confidence":"Medium","gaps":["Single-lab biochemical characterization","No functional assay linking ZP4 to sperm binding","Relationship to ZP2 not independently confirmed"]},{"year":1998,"claim":"Demonstration that neither ZPB (ZP4 ortholog) nor ZPC (ZP3 ortholog) alone binds sperm—only the hetero-oligomeric complex does—established that heterocomplex formation is the functional unit for sperm recognition at the zona pellucida.","evidence":"Solid-phase binding assays, size-exclusion chromatography, and Western blot with porcine zona proteins and boar sperm membrane vesicles","pmids":["9516448"],"confidence":"High","gaps":["Which glycan or peptide determinants on the complex mediate sperm binding was unknown","Whether the requirement for heterocomplex is conserved across species was untested"]},{"year":2007,"claim":"Reconstitution of recombinant bovine ZP3/ZP4 complexes with deletion mutants showed that the N-terminal and trefoil domains of ZP4 are dispensable for complex formation and sperm-binding inhibition, narrowing the essential region to the ZP domain, and demonstrating cross-species conservation of ZP3–ZP4 interactions.","evidence":"Baculovirus-Sf9 co-expression, deletion mutagenesis, immunoprecipitation, and competitive sperm-binding inhibition assay","pmids":["17894824"],"confidence":"High","gaps":["Precise ZP4 residues mediating ZP3 interaction not identified","Glycosylation contribution not dissected"]},{"year":2011,"claim":"Comparison of native versus recombinant ZP4 revealed that ZP4's carbohydrate moieties—not merely its polypeptide backbone—are required for sperm-binding activity of the ZP3/ZP4 complex, shifting the model from purely protein-mediated to glycan-dependent recognition.","evidence":"Competitive sperm-binding inhibition with native vs. Sf9-expressed recombinant porcine ZP4; reciprocal co-immunoprecipitation","pmids":["22008510"],"confidence":"High","gaps":["Specific glycan structures responsible not identified","Whether mammalian-type glycosylation fully restores activity was untested"]},{"year":2015,"claim":"Mapping ZP3's hinge region (Arg-32–Arg-160) as the ZP4-binding site and showing that Asn-146 glycosylation is required for functional activity (but not for physical interaction) defined the molecular interface governing the sperm-binding competence of the complex.","evidence":"ZP3 deletion/point mutants co-expressed with ZP4 in Sf9 cells, sperm-binding inhibition assay","pmids":["26610590"],"confidence":"High","gaps":["Reciprocal mapping of ZP3-contact residues on ZP4 not performed","No structural data for the ZP3/ZP4 interface"]},{"year":2019,"claim":"CRISPR-knockout rabbits revealed that ZP4 is essential for zona pellucida thickness, impermeability, and ultrastructural organization, and that while fertilization proceeds normally without ZP4, in vivo preimplantation development fails—establishing ZP4's structural role beyond sperm binding.","evidence":"CRISPR-Cas9 ZP4 KO rabbit, electron microscopy, permeability assays, in vivo and in vitro developmental tracking","pmids":["31635692"],"confidence":"High","gaps":["Molecular basis of how ZP4 cross-links the zona matrix unknown","Whether the structural phenotype is separable from specific ZP4 domains untested"]},{"year":2021,"claim":"ZP4 knockout in rats showed completely normal fertility and zona morphology, while human patient mutations impairing ZP4 secretion caused thin zonae and infertility, establishing that ZP4's requirement for reproduction is species-dependent and confirming its essential role in humans.","evidence":"CRISPR-Cas9 KO rat model with reproductive assays; whole-exome sequencing of infertile patients with in vitro secretion assays","pmids":["33709118","33461974"],"confidence":"High","gaps":["Basis for species-specific dispensability is mechanistically unexplained","Only secretion defect tested for human mutations; effects on ZP3 interaction or zona incorporation not assessed"]},{"year":2021,"claim":"Systematic deletion mapping of bovine ZP4 identified two discrete sperm-binding regions (N-terminal Lys-25–Asp-136 and middle Ser-290–Lys-340), establishing that ZP4 harbors the major sperm-binding determinants among bovine ZP glycoproteins.","evidence":"Solid-phase sperm-binding assay with Sf9-expressed ZP4 deletion mutants","pmids":["34242308"],"confidence":"Medium","gaps":["Single-lab study; awaits independent replication","Whether these regions are sufficient in context of the ZP3/ZP4 complex is untested","Contribution of glycans on these regions not dissected"]},{"year":null,"claim":"Key unresolved questions include the atomic-level structure of the ZP3/ZP4 heterocomplex, the identity of specific glycan structures on ZP4 that mediate sperm binding, and the molecular basis for species-dependent dispensability of ZP4 in zona matrix assembly.","evidence":"","pmids":[],"confidence":"High","gaps":["No crystal or cryo-EM structure of ZP3/ZP4 complex","Specific glycan structures on ZP4 responsible for sperm binding remain uncharacterized","Mechanistic explanation for species differences in ZP4 requirement is lacking"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[7,1,3]}],"localization":[{"term_id":"GO:0005576","term_label":"extracellular region","supporting_discovery_ids":[4,7,12]}],"pathway":[{"term_id":"R-HSA-1474165","term_label":"Reproduction","supporting_discovery_ids":[1,3,5,7,9,12]}],"complexes":["ZP3/ZP4 heterocomplex"],"partners":["ZP3","ZP1"],"other_free_text":[]},"mechanistic_narrative":"ZP4 is a zona pellucida glycoprotein that contributes to both the structural integrity of the zona matrix and sperm–zona recognition through formation of a glycosylation-dependent heterocomplex with ZP3. Native ZP4 glycoforms are required for the sperm-binding activity of the ZP3/ZP4 complex, with the N-terminal (Lys-25–Asp-136) and middle (Ser-290–Lys-340) regions of ZP4 harboring sperm-binding sites, while ZP3's hinge region (including Asn-146 glycosylation) mediates the ZP4 interaction [PMID:9516448, PMID:22008510, PMID:26610590, PMID:34242308]. CRISPR knockout in rabbits demonstrates that ZP4 is essential for zona thickness, impermeability, and ultrastructural organization required for in vivo preimplantation embryo survival, whereas Zp4-null rats retain normal zona morphology and full fertility, revealing species-dependent reliance on ZP4 [PMID:31635692, PMID:33709118]. Loss-of-function mutations in human ZP4, which impair protein secretion, are associated with thin, irregular zonae and female infertility [PMID:33461974]."},"prefetch_data":{"uniprot":{"accession":"Q12836","full_name":"Zona pellucida sperm-binding protein 4","aliases":["Zona pellucida glycoprotein 4","Zp-4","Zona pellucida protein B"],"length_aa":540,"mass_kda":59.4,"function":"Component of the zona pellucida, an extracellular matrix surrounding oocytes which mediates sperm binding, induction of the acrosome reaction and prevents post-fertilization polyspermy. The zona pellucida is composed of 3 to 4 glycoproteins, ZP1, ZP2, ZP3, and ZP4. ZP4 may act as a sperm receptor","subcellular_location":"Cell membrane","url":"https://www.uniprot.org/uniprotkb/Q12836/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ZP4","classification":"Not Classified","n_dependent_lines":1,"n_total_lines":1208,"dependency_fraction":0.0008278145695364238},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/ZP4","total_profiled":1310},"omim":[{"mim_id":"613514","title":"ZONA PELLUCIDA GLYCOPROTEIN 4; ZP4","url":"https://www.omim.org/entry/613514"},{"mim_id":"195000","title":"ZONA PELLUCIDA GLYCOPROTEIN 1; ZP1","url":"https://www.omim.org/entry/195000"},{"mim_id":"182889","title":"ZONA PELLUCIDA GLYCOPROTEIN 3; ZP3","url":"https://www.omim.org/entry/182889"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Tissue enriched","tissue_distribution":"Detected in single","driving_tissues":[{"tissue":"ovary","ntpm":3.6}],"url":"https://www.proteinatlas.org/search/ZP4"},"hgnc":{"alias_symbol":["ZPB","ZPB2","ZP1B"],"prev_symbol":[]},"alphafold":{"accession":"Q12836","domains":[{"cath_id":"2.60.40,2.60.40","chopping":"31-130","consensus_level":"high","plddt":75.3041,"start":31,"end":130},{"cath_id":"2.60.40.3210","chopping":"183-266_276-303","consensus_level":"high","plddt":82.9827,"start":183,"end":303},{"cath_id":"2.60.40.4100","chopping":"317-459_476-489","consensus_level":"high","plddt":86.8611,"start":317,"end":489}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q12836","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q12836-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q12836-F1-predicted_aligned_error_v6.png","plddt_mean":74.62},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ZP4","jax_strain_url":"https://www.jax.org/strain/search?query=ZP4"},"sequence":{"accession":"Q12836","fasta_url":"https://rest.uniprot.org/uniprotkb/Q12836.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q12836/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q12836"}},"corpus_meta":[{"pmid":"7841460","id":"PMC_7841460","title":"Cloning and characterization of zona pellucida genes and cDNAs from a variety of mammalian species: the ZPA, ZPB and ZPC gene families.","date":"1994","source":"DNA sequence : the journal of DNA sequencing and mapping","url":"https://pubmed.ncbi.nlm.nih.gov/7841460","citation_count":273,"is_preprint":false},{"pmid":"12580603","id":"PMC_12580603","title":"ZP4, an improved neuronal Zn2+ sensor of the Zinpyr family.","date":"2003","source":"Journal of the American Chemical Society","url":"https://pubmed.ncbi.nlm.nih.gov/12580603","citation_count":229,"is_preprint":false},{"pmid":"9516448","id":"PMC_9516448","title":"Hetero-oligomerization-dependent binding of pig oocyte zona pellucida glycoproteins ZPB and ZPC to boar sperm membrane vesicles.","date":"1998","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/9516448","citation_count":126,"is_preprint":false},{"pmid":"10979984","id":"PMC_10979984","title":"The major chicken egg envelope protein ZP1 is different from ZPB and is synthesized in the liver.","date":"2000","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/10979984","citation_count":102,"is_preprint":false},{"pmid":"10456835","id":"PMC_10456835","title":"Infertility in female rabbits (Oryctolagus cuniculus) alloimmunized with the rabbit zona pellucida protein ZPB either as a purified recombinant protein or expressed by recombinant myxoma virus.","date":"1999","source":"Biology of reproduction","url":"https://pubmed.ncbi.nlm.nih.gov/10456835","citation_count":46,"is_preprint":false},{"pmid":"12968942","id":"PMC_12968942","title":"Analysis of fish ZP1/ZPB homologous genes--evidence for both genome duplication and species-specific amplification models of evolution.","date":"2003","source":"Reproduction (Cambridge, England)","url":"https://pubmed.ncbi.nlm.nih.gov/12968942","citation_count":45,"is_preprint":false},{"pmid":"8182597","id":"PMC_8182597","title":"Amino acid sequence of a porcine zona pellucida glycoprotein ZP4 determined by peptide mapping and cDNA cloning.","date":"1994","source":"Journal of reproduction and fertility","url":"https://pubmed.ncbi.nlm.nih.gov/8182597","citation_count":45,"is_preprint":false},{"pmid":"31635692","id":"PMC_31635692","title":"ZP4 confers structural properties to the zona pellucida essential for embryo development.","date":"2019","source":"eLife","url":"https://pubmed.ncbi.nlm.nih.gov/31635692","citation_count":39,"is_preprint":false},{"pmid":"11006142","id":"PMC_11006142","title":"Effectiveness of zona pellucida protein ZPB as an immunocontraceptive antigen.","date":"2000","source":"Journal of reproduction and fertility","url":"https://pubmed.ncbi.nlm.nih.gov/11006142","citation_count":34,"is_preprint":false},{"pmid":"1501206","id":"PMC_1501206","title":"Antifertility effect of active immunization with ZP4 glycoprotein family of porcine zona pellucida in hamsters.","date":"1992","source":"Journal of reproductive immunology","url":"https://pubmed.ncbi.nlm.nih.gov/1501206","citation_count":30,"is_preprint":false},{"pmid":"12878193","id":"PMC_12878193","title":"Identification of the carboxyl termini of porcine zona pellucida glycoproteins ZPB and ZPC.","date":"2003","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/12878193","citation_count":29,"is_preprint":false},{"pmid":"11291774","id":"PMC_11291774","title":"Immunohistochemical localization of zona pellucida proteins ZPA, ZPB and ZPC in human, cynomolgus monkey and mouse ovaries.","date":"2001","source":"Cell and tissue research","url":"https://pubmed.ncbi.nlm.nih.gov/11291774","citation_count":23,"is_preprint":false},{"pmid":"21173071","id":"PMC_21173071","title":"Human ZP4 is not sufficient for taxon-specific sperm recognition of the zona pellucida in transgenic mice.","date":"2010","source":"Reproduction (Cambridge, England)","url":"https://pubmed.ncbi.nlm.nih.gov/21173071","citation_count":23,"is_preprint":false},{"pmid":"33461974","id":"PMC_33461974","title":"Mutations in ZP4 are associated with abnormal zona pellucida and female infertility.","date":"2021","source":"Journal of clinical pathology","url":"https://pubmed.ncbi.nlm.nih.gov/33461974","citation_count":21,"is_preprint":false},{"pmid":"17894824","id":"PMC_17894824","title":"Recombinant bovine zona pellucida glycoproteins ZP3 and ZP4 coexpressed in Sf9 cells form a sperm-binding active hetero-complex.","date":"2007","source":"The FEBS journal","url":"https://pubmed.ncbi.nlm.nih.gov/17894824","citation_count":21,"is_preprint":false},{"pmid":"21729200","id":"PMC_21729200","title":"Oocytic expression of zona pellucida protein ZP4 in Japanese quail (Coturnix japonica).","date":"2011","source":"Animal science journal = Nihon chikusan Gakkaiho","url":"https://pubmed.ncbi.nlm.nih.gov/21729200","citation_count":20,"is_preprint":false},{"pmid":"22008510","id":"PMC_22008510","title":"Porcine zona pellucida glycoprotein ZP4 is responsible for the sperm-binding activity of the ZP3/ZP4 complex.","date":"2011","source":"Zygote (Cambridge, England)","url":"https://pubmed.ncbi.nlm.nih.gov/22008510","citation_count":20,"is_preprint":false},{"pmid":"9890748","id":"PMC_9890748","title":"Isolation and characterisation of a cDNA encoding a zona pellucida protein (ZPB) from the marsupial Trichosurus vulpecula (brushtail possum).","date":"1999","source":"Molecular reproduction and development","url":"https://pubmed.ncbi.nlm.nih.gov/9890748","citation_count":19,"is_preprint":false},{"pmid":"26610590","id":"PMC_26610590","title":"The Hinge Region of Bovine Zona Pellucida Glycoprotein ZP3 Is Involved in the Formation of the Sperm-Binding Active ZP3/ZP4 Complex.","date":"2015","source":"Biomolecules","url":"https://pubmed.ncbi.nlm.nih.gov/26610590","citation_count":19,"is_preprint":false},{"pmid":"25740067","id":"PMC_25740067","title":"Study on the zona pellucida 4 (ZP4) gene sequence and its expression in the ovaries of patients with polycystic ovary syndrome.","date":"2015","source":"Journal of endocrinological investigation","url":"https://pubmed.ncbi.nlm.nih.gov/25740067","citation_count":17,"is_preprint":false},{"pmid":"25229478","id":"PMC_25229478","title":"Structural studies of \"aggregation-prone\" peptide-analogues of teleostean egg chorion ZPB proteins.","date":"2014","source":"Biopolymers","url":"https://pubmed.ncbi.nlm.nih.gov/25229478","citation_count":16,"is_preprint":false},{"pmid":"9488904","id":"PMC_9488904","title":"Localization of the zona glycoproteins ZPB (ZP3 alpha) and ZPC (ZP3 beta) in the bovine ovary during pre- and postnatal development.","date":"1998","source":"Annals of anatomy = Anatomischer Anzeiger : official organ of the Anatomische Gesellschaft","url":"https://pubmed.ncbi.nlm.nih.gov/9488904","citation_count":16,"is_preprint":false},{"pmid":"31314571","id":"PMC_31314571","title":"The developmentally regulated fetal enterocyte gene, ZP4, mediates anti-inflammation by the symbiotic bacterial surface factor polysaccharide A on Bacteroides fragilis.","date":"2019","source":"American journal of physiology. Gastrointestinal and liver physiology","url":"https://pubmed.ncbi.nlm.nih.gov/31314571","citation_count":12,"is_preprint":false},{"pmid":"30611916","id":"PMC_30611916","title":"Mutation analysis of ZP1, ZP2, ZP3 and ZP4 genes in 152 Han Chinese samples with ovarian endometriosis.","date":"2018","source":"Mutation research","url":"https://pubmed.ncbi.nlm.nih.gov/30611916","citation_count":10,"is_preprint":false},{"pmid":"33869165","id":"PMC_33869165","title":"Transcriptional Repression of MFG-E8 Causes Disturbance in the Homeostasis of Cell Cycle Through DOCK/ZP4/STAT Signaling in Buffalo Mammary Epithelial Cells.","date":"2021","source":"Frontiers in cell and developmental biology","url":"https://pubmed.ncbi.nlm.nih.gov/33869165","citation_count":10,"is_preprint":false},{"pmid":"33709118","id":"PMC_33709118","title":"Zp4 is completely dispensable for fertility in female rats†.","date":"2021","source":"Biology of reproduction","url":"https://pubmed.ncbi.nlm.nih.gov/33709118","citation_count":10,"is_preprint":false},{"pmid":"33624742","id":"PMC_33624742","title":"Mutant Zp1 impedes incorporation of ZP3 and ZP4 in the zona pellucida, resulting in zona absence and female infertility in rats†.","date":"2021","source":"Biology of reproduction","url":"https://pubmed.ncbi.nlm.nih.gov/33624742","citation_count":9,"is_preprint":false},{"pmid":"19004505","id":"PMC_19004505","title":"DNA vaccine encoding chimeric protein encompassing epitopes of human ZP3 and ZP4: immunogenicity and characterization of antibodies.","date":"2008","source":"Journal of reproductive immunology","url":"https://pubmed.ncbi.nlm.nih.gov/19004505","citation_count":9,"is_preprint":false},{"pmid":"34242308","id":"PMC_34242308","title":"Sperm-binding regions on bovine egg zona pellucida glycoprotein ZP4 studied in a solid supported form on plastic plate.","date":"2021","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/34242308","citation_count":6,"is_preprint":false},{"pmid":"39980195","id":"PMC_39980195","title":"ZP4: A novel target for CAR-T cell therapy in triple negative breast cancer.","date":"2025","source":"Molecular therapy : the journal of the American Society of Gene Therapy","url":"https://pubmed.ncbi.nlm.nih.gov/39980195","citation_count":3,"is_preprint":false},{"pmid":"30298508","id":"PMC_30298508","title":"[Association of ZP4 gene polymorphism with primary open-angle glaucoma in an ethnic Chinese population from Sichuan province].","date":"2018","source":"Zhonghua yi xue yi chuan xue za zhi = Zhonghua yixue yichuanxue zazhi = Chinese journal of medical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/30298508","citation_count":1,"is_preprint":false},{"pmid":"35804570","id":"PMC_35804570","title":"Quantitative Protein Analysis of ZPB2, ZPB1 and ZPC in the Germinal Disc and a Non-Germinal Disc Region of the Inner Perivitelline Layer in Two Genetic Lines of Turkey Hens That Differ in Fertility.","date":"2022","source":"Animals : an open access journal from MDPI","url":"https://pubmed.ncbi.nlm.nih.gov/35804570","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":15884,"output_tokens":3004,"usd":0.046356},"stage2":{"model":"claude-opus-4-6","input_tokens":6376,"output_tokens":2234,"usd":0.131595},"total_usd":0.177951,"stage1_batch_id":"msgbatch_01BparvgRZ9S2LMidvaoo86K","stage2_batch_id":"msgbatch_01NkBfv9SVZWthhS7j6mU6G3","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1994,\n      \"finding\": \"Porcine zona pellucida glycoprotein ZP4 amino acid sequence was determined by peptide mapping and cDNA cloning, revealing two peptide isoforms (128 and 133 amino acids) with two potential N-linked glycosylation sites, five conserved cysteine residues, and sequence similarity to mouse ZP2; ZP4 and ZP2 appear to be derived from a common parent polypeptide by proteolytic cleavage.\",\n      \"method\": \"Peptide mapping, cDNA cloning, sequence analysis\",\n      \"journal\": \"Journal of reproduction and fertility\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 — direct biochemical sequencing and cDNA cloning, single lab\",\n      \"pmids\": [\"8182597\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"Porcine ZPB (ortholog of mouse ZP1) and ZPC (ortholog of mouse ZP3) form hetero-oligomeric complexes that bind to boar sperm membrane vesicles with high affinity; neither free ZPB nor free ZPC alone binds sperm membranes, demonstrating that heterocomplex formation is required for sperm-zona binding activity.\",\n      \"method\": \"Solid phase binding assays, size exclusion chromatography, Western blot, biotin-labeling competition assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal methods (binding assay, chromatography, Western blot) in a single rigorous study\",\n      \"pmids\": [\"9516448\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Porcine ZPB and ZPC C-terminal residues were identified (Ala-462 and Ser-332, respectively) by mass spectrometry, revealing that ZPB is processed at its furin consensus site whereas ZPC is processed N-terminal to the furin consensus site; disulfide bond analysis within the ZP domains suggested the ZP domain consists of two subdomains.\",\n      \"method\": \"Mass spectrometry of C-terminal polypeptide fragments\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 — direct mass spectrometry characterization, single lab\",\n      \"pmids\": [\"12878193\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Recombinant bovine ZP3 and ZP4 co-expressed in Sf9 insect cells form a hetero-complex that inhibits bovine sperm-ZP binding; neither ZP3 nor ZP4 alone shows inhibitory activity. The N-terminal and trefoil domains of ZP4 are dispensable for formation of the sperm-binding active ZP3/ZP4 complex, and the molecular interactions between ZP3 and ZP4 are conserved between bovine and porcine systems.\",\n      \"method\": \"Baculovirus-Sf9 cell expression, sperm-ZP competitive inhibition assay, immunoprecipitation, deletion mutagenesis\",\n      \"journal\": \"The FEBS journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — reconstitution with deletion mutagenesis, functional assay, and immunoprecipitation in a single study\",\n      \"pmids\": [\"17894824\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Human ZP4 expressed in transgenic mice is incorporated into the zona pellucida matrix with molecular mass similar to native isoforms, but is not sufficient to confer human sperm binding to mouse eggs, indicating that additional zona proteins are required for species-selective human gamete recognition.\",\n      \"method\": \"Transgenic mouse model, sperm binding assay, molecular mass analysis\",\n      \"journal\": \"Reproduction (Cambridge, England)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean transgenic KI model with defined functional readout (sperm binding) and protein incorporation confirmed\",\n      \"pmids\": [\"21173071\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"In porcine ZP, native ZP4 (nZP4) — but not recombinant ZP4 — is necessary for the sperm-binding activity of the ZP3/ZP4 complex toward porcine sperm, demonstrating that carbohydrate structures on ZP4 are responsible for porcine sperm-binding activity of the complex; ZP3 and ZP4 reciprocally co-precipitate each other via recombinant N-terminal tags.\",\n      \"method\": \"Competitive sperm-binding inhibition assay, co-immunoprecipitation, baculovirus-Sf9 expression\",\n      \"journal\": \"Zygote (Cambridge, England)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — functional reconstitution with native vs. recombinant protein comparison and reciprocal co-precipitation\",\n      \"pmids\": [\"22008510\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"The hinge region (Arg-32 to Arg-160) of bovine ZP3 is a ZP4-binding site; N-glycosylation at Asn-146 (but not Asn-124) in this region is required for the sperm-binding activity of the ZP3/ZP4 complex, although it does not prevent ZP4 interaction per se.\",\n      \"method\": \"Baculovirus-Sf9 expression, ZP3 deletion and point mutants, co-expression, sperm-ZP binding inhibition assay\",\n      \"journal\": \"Biomolecules\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — mutagenesis combined with functional reconstitution assay mapping the ZP4-binding site on ZP3\",\n      \"pmids\": [\"26610590\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"ZP4 ablation in rabbits (CRISPR KO) produced a zona pellucida that was significantly thinner, more permeable, and more disorganized/fenestrated; ovulation, fertilization, and in vitro development were unaffected, but in vivo preimplantation development was severely impaired, demonstrating that ZP4 is required for structural integrity of the zona pellucida necessary for embryo protection in vivo.\",\n      \"method\": \"CRISPR-Cas9 knockout rabbit model, electron microscopy of ZP structure, permeability assays, in vivo/in vitro fertility assays\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean KO with multiple orthogonal phenotypic readouts (structural, permeability, developmental) and rigorous controls\",\n      \"pmids\": [\"31635692\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"ZP4 is expressed in immature human fetal enterocytes and is regulated by Toll-like receptor 2 and 4 signaling; knockdown of ZP4 in fetal intestinal cells altered IL-8 mRNA expression in response to IL-1β, implicating ZP4 in innate immune signaling in the developing intestine.\",\n      \"method\": \"Transcription profiling, ZP4 knockdown in H4 fetal intestinal cell line, IL-8 mRNA measurement\",\n      \"journal\": \"American journal of physiology. Gastrointestinal and liver physiology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 — single knockdown experiment in a cell line, limited mechanistic follow-up\",\n      \"pmids\": [\"31314571\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Zp4 knockout female rats (CRISPR-Cas9) showed normal ZP morphology and were completely fertile, with no defects in oogenesis, ovulation, fertilization, or pup production, demonstrating that Zp4 is fully dispensable for female fertility in rats.\",\n      \"method\": \"CRISPR-Cas9 KO rat model, PAS staining, superovulation, in vitro fertilization, natural mating\",\n      \"journal\": \"Biology of reproduction\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean KO model with multiple orthogonal reproductive phenotype readouts\",\n      \"pmids\": [\"33709118\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"A mutant ZP1 protein in rats is intracellularly sequestered and interacts with wild-type ZP3 or ZP4 (shown in 293T cells), impeding incorporation of ZP3 and ZP4 into the zona pellucida, resulting in zona absence and female infertility.\",\n      \"method\": \"Homologous rat KI model, 293T cell co-expression, co-immunoprecipitation, immunofluorescence\",\n      \"journal\": \"Biology of reproduction\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — reciprocal co-IP in cells, KI animal model; single lab\",\n      \"pmids\": [\"33624742\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"The N-terminal region (Lys-25 to Asp-136) and middle region (Ser-290 to Lys-340) of bovine ZP4 contain sperm-binding sites, as demonstrated by binding of sperm to ZP4-coated plates and systematic deletion mutagenesis; ZP4 harbors the major potential sperm-binding sites among bovine ZP glycoproteins, but multivalent complex formation is required for full activity.\",\n      \"method\": \"ZP4-coated plate solid-phase binding assay, deletion mutagenesis, Sf9-expressed recombinant proteins\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — systematic deletion mapping with functional readout, single lab\",\n      \"pmids\": [\"34242308\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ZP4 mutations identified in human patients (whole-exome sequencing) reduced ZP4 secretion in vitro when mutant plasmids were transfected into cells, linking ZP4 loss-of-function to abnormal (thin and irregular) zona pellucida and female infertility.\",\n      \"method\": \"Whole-exome sequencing, in vitro plasmid expression/secretion assay in transfected cells\",\n      \"journal\": \"Journal of clinical pathology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — human genetic evidence combined with in vitro secretion assay; mechanistic follow-up limited to secretion\",\n      \"pmids\": [\"33461974\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ZP4 is a zona pellucida glycoprotein that forms a functional heterocomplex with ZP3 (requiring ZP4's carbohydrate moieties) to mediate sperm binding to the zona pellucida; it confers structural integrity (thickness, permeability, organization) to the zona matrix essential for in vivo embryo protection, though its requirement for fertility is species-dependent (indispensable in rabbits and humans, dispensable in rats and mice).\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"ZP4 is a zona pellucida glycoprotein that contributes to both the structural integrity of the zona matrix and sperm–zona recognition through formation of a glycosylation-dependent heterocomplex with ZP3. Native ZP4 glycoforms are required for the sperm-binding activity of the ZP3/ZP4 complex, with the N-terminal (Lys-25–Asp-136) and middle (Ser-290–Lys-340) regions of ZP4 harboring sperm-binding sites, while ZP3's hinge region (including Asn-146 glycosylation) mediates the ZP4 interaction [PMID:9516448, PMID:22008510, PMID:26610590, PMID:34242308]. CRISPR knockout in rabbits demonstrates that ZP4 is essential for zona thickness, impermeability, and ultrastructural organization required for in vivo preimplantation embryo survival, whereas Zp4-null rats retain normal zona morphology and full fertility, revealing species-dependent reliance on ZP4 [PMID:31635692, PMID:33709118]. Loss-of-function mutations in human ZP4, which impair protein secretion, are associated with thin, irregular zonae and female infertility [PMID:33461974].\",\n  \"teleology\": [\n    {\n      \"year\": 1994,\n      \"claim\": \"Determining ZP4's primary structure established it as a zona pellucida glycoprotein with conserved cysteine residues and N-glycosylation sites, related to ZP2 via proteolytic processing of a common precursor.\",\n      \"evidence\": \"Peptide mapping and cDNA cloning of porcine ZP4\",\n      \"pmids\": [\"8182597\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab biochemical characterization\", \"No functional assay linking ZP4 to sperm binding\", \"Relationship to ZP2 not independently confirmed\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Demonstration that neither ZPB (ZP4 ortholog) nor ZPC (ZP3 ortholog) alone binds sperm—only the hetero-oligomeric complex does—established that heterocomplex formation is the functional unit for sperm recognition at the zona pellucida.\",\n      \"evidence\": \"Solid-phase binding assays, size-exclusion chromatography, and Western blot with porcine zona proteins and boar sperm membrane vesicles\",\n      \"pmids\": [\"9516448\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Which glycan or peptide determinants on the complex mediate sperm binding was unknown\", \"Whether the requirement for heterocomplex is conserved across species was untested\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Reconstitution of recombinant bovine ZP3/ZP4 complexes with deletion mutants showed that the N-terminal and trefoil domains of ZP4 are dispensable for complex formation and sperm-binding inhibition, narrowing the essential region to the ZP domain, and demonstrating cross-species conservation of ZP3–ZP4 interactions.\",\n      \"evidence\": \"Baculovirus-Sf9 co-expression, deletion mutagenesis, immunoprecipitation, and competitive sperm-binding inhibition assay\",\n      \"pmids\": [\"17894824\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Precise ZP4 residues mediating ZP3 interaction not identified\", \"Glycosylation contribution not dissected\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Comparison of native versus recombinant ZP4 revealed that ZP4's carbohydrate moieties—not merely its polypeptide backbone—are required for sperm-binding activity of the ZP3/ZP4 complex, shifting the model from purely protein-mediated to glycan-dependent recognition.\",\n      \"evidence\": \"Competitive sperm-binding inhibition with native vs. Sf9-expressed recombinant porcine ZP4; reciprocal co-immunoprecipitation\",\n      \"pmids\": [\"22008510\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Specific glycan structures responsible not identified\", \"Whether mammalian-type glycosylation fully restores activity was untested\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Mapping ZP3's hinge region (Arg-32–Arg-160) as the ZP4-binding site and showing that Asn-146 glycosylation is required for functional activity (but not for physical interaction) defined the molecular interface governing the sperm-binding competence of the complex.\",\n      \"evidence\": \"ZP3 deletion/point mutants co-expressed with ZP4 in Sf9 cells, sperm-binding inhibition assay\",\n      \"pmids\": [\"26610590\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Reciprocal mapping of ZP3-contact residues on ZP4 not performed\", \"No structural data for the ZP3/ZP4 interface\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"CRISPR-knockout rabbits revealed that ZP4 is essential for zona pellucida thickness, impermeability, and ultrastructural organization, and that while fertilization proceeds normally without ZP4, in vivo preimplantation development fails—establishing ZP4's structural role beyond sperm binding.\",\n      \"evidence\": \"CRISPR-Cas9 ZP4 KO rabbit, electron microscopy, permeability assays, in vivo and in vitro developmental tracking\",\n      \"pmids\": [\"31635692\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular basis of how ZP4 cross-links the zona matrix unknown\", \"Whether the structural phenotype is separable from specific ZP4 domains untested\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"ZP4 knockout in rats showed completely normal fertility and zona morphology, while human patient mutations impairing ZP4 secretion caused thin zonae and infertility, establishing that ZP4's requirement for reproduction is species-dependent and confirming its essential role in humans.\",\n      \"evidence\": \"CRISPR-Cas9 KO rat model with reproductive assays; whole-exome sequencing of infertile patients with in vitro secretion assays\",\n      \"pmids\": [\"33709118\", \"33461974\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Basis for species-specific dispensability is mechanistically unexplained\", \"Only secretion defect tested for human mutations; effects on ZP3 interaction or zona incorporation not assessed\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Systematic deletion mapping of bovine ZP4 identified two discrete sperm-binding regions (N-terminal Lys-25–Asp-136 and middle Ser-290–Lys-340), establishing that ZP4 harbors the major sperm-binding determinants among bovine ZP glycoproteins.\",\n      \"evidence\": \"Solid-phase sperm-binding assay with Sf9-expressed ZP4 deletion mutants\",\n      \"pmids\": [\"34242308\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab study; awaits independent replication\", \"Whether these regions are sufficient in context of the ZP3/ZP4 complex is untested\", \"Contribution of glycans on these regions not dissected\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Key unresolved questions include the atomic-level structure of the ZP3/ZP4 heterocomplex, the identity of specific glycan structures on ZP4 that mediate sperm binding, and the molecular basis for species-dependent dispensability of ZP4 in zona matrix assembly.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No crystal or cryo-EM structure of ZP3/ZP4 complex\", \"Specific glycan structures on ZP4 responsible for sperm binding remain uncharacterized\", \"Mechanistic explanation for species differences in ZP4 requirement is lacking\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [7, 1, 3]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005576\", \"supporting_discovery_ids\": [4, 7, 12]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1474165\", \"supporting_discovery_ids\": [1, 3, 5, 7, 9, 12]}\n    ],\n    \"complexes\": [\n      \"ZP3/ZP4 heterocomplex\"\n    ],\n    \"partners\": [\n      \"ZP3\",\n      \"ZP1\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}