{"gene":"PIH1D2","run_date":"2026-06-10T06:43:35","timeline":{"discoveries":[{"year":2018,"finding":"PIH1D2 forms an R2TP-like complex termed R2SP together with SPAG1 (an RPAP3-like protein), RUVBL1, and RUVBL2. Systematic interaction analyses showed that SPAG1 binds PIH1D2 and RUVBL1/2. R2SP is enriched in testis, functions in quaternary protein folding of specific clients (e.g., liprin-α2), and is required for liprin-α2 expression and assembly of liprin-α2 complexes.","method":"Systematic interaction analyses (co-immunoprecipitation/pulldown), structure determination of RPAP3-C-terminal domain, client identification by mass spectrometry, functional validation by R2SP depletion with client expression and complex assembly readout","journal":"Nature Communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal interaction analyses, structural data, functional depletion experiments with defined client readout, multiple orthogonal methods in one study","pmids":["29844425"],"is_preprint":false},{"year":2018,"finding":"PIH1D2 (zebrafish pih1d2) is specifically required for the assembly of a defined subset of axonemal dynein subtypes in sperm flagella; loss of pih1d2 results in loss of specific dynein arms as visualized by cryo-electron tomography, leading to abnormal sperm motility and defects in Kupffer's vesicle cilia motility.","method":"Zebrafish mutant generation and cryo-electron tomography of axonemal dynein structure in mutant spermatozoa, motility assays","journal":"eLife","confidence":"High","confidence_rationale":"Tier 2 / Strong — loss-of-function mutants with defined ultrastructural phenotype (cryo-ET) and motility readout, replicated across all four PIH family members in same study","pmids":["29741156"],"is_preprint":false},{"year":2019,"finding":"Within the R2SP complex, PIH1D2 partners with SPAG1, RUVBL1, and RUVBL2. SPAG1's TPR domains recruit HSP70 and HSP90 chaperones to the complex, and SPAG1 regulates nucleotide hydrolysis activity of HSP and RUVBL1/2 partners rather than possessing autonomous GTPase activity.","method":"Biochemical assays, isothermal titration calorimetry (ITC), NMR spectroscopy, molecular dynamics simulations, in vitro GTP hydrolysis assay","journal":"The Biochemical Journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal biochemical and structural methods (ITC, NMR, MD), single lab; directly characterizes SPAG1 binding properties within the PIH1D2-containing R2SP complex","pmids":["31118266"],"is_preprint":false},{"year":2025,"finding":"Cryo-EM, NMR, and structural mass spectrometry of the human R2SP complex (RUVBL1, RUVBL2, SPAG1, PIH1D2) reveal a three-dimensional organization similar to the canonical R2TP complex but with distinct differences in the ATPase activity of the RUVBL1/2 core and in the mode of binding of adaptors SPAG1 and PIH1D2.","method":"Cryo-EM, NMR spectroscopy, structural mass spectrometry, biochemical ATPase assays","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — structural and biochemical methods (cryo-EM, NMR, MS) are high-tier but this is a preprint, not yet peer-reviewed, single study","pmids":["bio_10.1101_2025.01.27.635100"],"is_preprint":true}],"current_model":"PIH1D2 is an adaptor subunit of the R2SP quaternary chaperone complex (together with SPAG1, RUVBL1, and RUVBL2) that promotes the folding and assembly of specific protein clients including liprin-α2 and axonemal dynein subtypes, particularly in testis and motile cilia, with SPAG1 recruiting HSP70/HSP90 to the complex and RUVBL1/2 providing ATPase activity; the overall architecture resembles the canonical R2TP complex but with distinct differences in ATPase regulation and adaptor binding modes."},"narrative":{"mechanistic_narrative":"PIH1D2 is an adaptor subunit of the R2SP chaperone complex, a testis-enriched R2TP-like machine that promotes the quaternary folding and assembly of specific protein clients [PMID:29844425]. Within R2SP, PIH1D2 partners with SPAG1 (an RPAP3-like protein) and the AAA+ ATPases RUVBL1 and RUVBL2, with SPAG1 binding both PIH1D2 and RUVBL1/2 and recruiting HSP70/HSP90 chaperones through its TPR domains [PMID:29844425, PMID:31118266]. SPAG1 itself regulates the nucleotide hydrolysis activity of its HSP and RUVBL1/2 partners rather than acting as an autonomous GTPase [PMID:31118266]. The complex is required for the expression and assembly of clients such as liprin-α2 [PMID:29844425], and in vivo loss of PIH1D2 abolishes the assembly of a defined subset of axonemal dynein arms in sperm flagella and Kupffer's vesicle cilia, producing abnormal motility [PMID:29741156]. This links PIH1D2 to the building of motile-ciliary and flagellar dynein machinery through chaperone-assisted protein assembly.","teleology":[{"year":2018,"claim":"Established that PIH1D2 is not a free-standing protein but the adaptor subunit of an R2TP-like assembly chaperone (R2SP), defining its molecular context and a first client.","evidence":"Systematic co-IP/pulldown interaction analyses, RPAP3 C-terminal domain structure, mass-spec client identification, and depletion with liprin-α2 assembly readout","pmids":["29844425"],"confidence":"High","gaps":["Full client repertoire beyond liprin-α2 not defined","Mechanism by which PIH1D2 selects clients not resolved"]},{"year":2018,"claim":"Demonstrated an in vivo function for PIH1D2 in assembling specific axonemal dynein subtypes, connecting the chaperone adaptor to motile cilia/flagellar biology.","evidence":"Zebrafish loss-of-function mutants with cryo-electron tomography of sperm axonemal dynein and motility assays","pmids":["29741156"],"confidence":"High","gaps":["Which dynein subunits are direct R2SP clients not established","Relationship to R2SP-mediated folding in this context not directly shown"]},{"year":2019,"claim":"Clarified how chaperone activity is recruited and regulated within the PIH1D2 complex, showing SPAG1 brings in HSP70/HSP90 and tunes hydrolysis of partner ATPases.","evidence":"ITC, NMR, molecular dynamics, and in vitro GTP hydrolysis assays on SPAG1 within the R2SP complex","pmids":["31118266"],"confidence":"Medium","gaps":["Single-lab biochemical characterization","PIH1D2's own contribution to ATPase regulation not isolated"]},{"year":2025,"claim":"Provided a structural model of the human R2SP complex, showing how PIH1D2 and SPAG1 dock onto the RUVBL1/2 core and how its ATPase regulation differs from canonical R2TP.","evidence":"Cryo-EM, NMR, structural mass spectrometry, and biochemical ATPase assays of human R2SP (preprint)","pmids":["bio_10.1101_2025.01.27.635100"],"confidence":"Medium","gaps":["Preprint, not peer-reviewed","Client-bound structure not determined","Functional consequence of distinct ATPase regulation untested"]},{"year":null,"claim":"How PIH1D2 recognizes and selects its diverse clients (liprin-α2 versus axonemal dyneins) and how client specificity is encoded in the complex remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No client-recognition determinant mapped on PIH1D2","Tissue-specific regulation of R2SP activity uncharacterized"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,2]},{"term_id":"GO:0044183","term_label":"protein folding chaperone","supporting_discovery_ids":[0,2]}],"localization":[],"pathway":[{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0]}],"complexes":["R2SP"],"partners":["SPAG1","RUVBL1","RUVBL2"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q8WWB5","full_name":"PIH1 domain-containing protein 2","aliases":[],"length_aa":315,"mass_kda":36.0,"function":"","subcellular_location":"","url":"https://www.uniprot.org/uniprotkb/Q8WWB5/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/PIH1D2","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/PIH1D2","total_profiled":1310},"omim":[],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Nucleoplasm","reliability":"Approved"},{"location":"Vesicles","reliability":"Additional"},{"location":"Basal body","reliability":"Additional"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Tissue enriched","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"testis","ntpm":62.1}],"url":"https://www.proteinatlas.org/search/PIH1D2"},"hgnc":{"alias_symbol":["DNAAF15"],"prev_symbol":[]},"alphafold":{"accession":"Q8WWB5","domains":[{"cath_id":"-","chopping":"11-46","consensus_level":"medium","plddt":81.0678,"start":11,"end":46},{"cath_id":"-","chopping":"50-165","consensus_level":"high","plddt":91.5414,"start":50,"end":165},{"cath_id":"2.60.40.790","chopping":"236-314","consensus_level":"high","plddt":86.7822,"start":236,"end":314}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8WWB5","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q8WWB5-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q8WWB5-F1-predicted_aligned_error_v6.png","plddt_mean":77.94},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=PIH1D2","jax_strain_url":"https://www.jax.org/strain/search?query=PIH1D2"},"sequence":{"accession":"Q8WWB5","fasta_url":"https://rest.uniprot.org/uniprotkb/Q8WWB5.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q8WWB5/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8WWB5"}},"corpus_meta":[{"pmid":"29844425","id":"PMC_29844425","title":"The RPAP3-Cterminal domain identifies R2TP-like quaternary chaperones.","date":"2018","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/29844425","citation_count":63,"is_preprint":false},{"pmid":"29741156","id":"PMC_29741156","title":"Systematic studies of all PIH proteins in zebrafish reveal their distinct roles in axonemal dynein assembly.","date":"2018","source":"eLife","url":"https://pubmed.ncbi.nlm.nih.gov/29741156","citation_count":54,"is_preprint":false},{"pmid":"35207567","id":"PMC_35207567","title":"Omics and Male Infertility: Highlighting the Application of Transcriptomic Data.","date":"2022","source":"Life (Basel, Switzerland)","url":"https://pubmed.ncbi.nlm.nih.gov/35207567","citation_count":30,"is_preprint":false},{"pmid":"32251302","id":"PMC_32251302","title":"Transcriptomic analysis of female and male gonads in juvenile snakeskin gourami (Trichopodus pectoralis).","date":"2020","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/32251302","citation_count":14,"is_preprint":false},{"pmid":"31118266","id":"PMC_31118266","title":"Binding properties of the quaternary assembly protein SPAG1.","date":"2019","source":"The Biochemical journal","url":"https://pubmed.ncbi.nlm.nih.gov/31118266","citation_count":13,"is_preprint":false},{"pmid":"20310044","id":"PMC_20310044","title":"Novel germline SDHD deletion associated with an unusual sympathetic head and neck paraganglioma.","date":"2010","source":"Head & neck","url":"https://pubmed.ncbi.nlm.nih.gov/20310044","citation_count":4,"is_preprint":false},{"pmid":"39857704","id":"PMC_39857704","title":"Genetic Nurture Effects on Type 2 Diabetes Among Chinese Han Adults: A Family-Based Design.","date":"2025","source":"Biomedicines","url":"https://pubmed.ncbi.nlm.nih.gov/39857704","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.01.27.635100","title":"R2TP-like Quaternary Chaperones: a comprehensive overview to understand the dynamic R2SP complex","date":"2025-01-28","source":"bioRxiv","url":"https://doi.org/10.1101/2025.01.27.635100","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":6501,"output_tokens":1248,"usd":0.019112,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":7822,"output_tokens":1744,"usd":0.041355,"stage2_stop_reason":"end_turn"},"total_usd":0.060467,"stage1_batch_id":"msgbatch_01CNoDbkChHEkgrJnL7BwtGJ","stage2_batch_id":"msgbatch_016Y4x9E9Wg5VQDANj9quSQD","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2018,\n      \"finding\": \"PIH1D2 forms an R2TP-like complex termed R2SP together with SPAG1 (an RPAP3-like protein), RUVBL1, and RUVBL2. Systematic interaction analyses showed that SPAG1 binds PIH1D2 and RUVBL1/2. R2SP is enriched in testis, functions in quaternary protein folding of specific clients (e.g., liprin-α2), and is required for liprin-α2 expression and assembly of liprin-α2 complexes.\",\n      \"method\": \"Systematic interaction analyses (co-immunoprecipitation/pulldown), structure determination of RPAP3-C-terminal domain, client identification by mass spectrometry, functional validation by R2SP depletion with client expression and complex assembly readout\",\n      \"journal\": \"Nature Communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal interaction analyses, structural data, functional depletion experiments with defined client readout, multiple orthogonal methods in one study\",\n      \"pmids\": [\"29844425\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"PIH1D2 (zebrafish pih1d2) is specifically required for the assembly of a defined subset of axonemal dynein subtypes in sperm flagella; loss of pih1d2 results in loss of specific dynein arms as visualized by cryo-electron tomography, leading to abnormal sperm motility and defects in Kupffer's vesicle cilia motility.\",\n      \"method\": \"Zebrafish mutant generation and cryo-electron tomography of axonemal dynein structure in mutant spermatozoa, motility assays\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — loss-of-function mutants with defined ultrastructural phenotype (cryo-ET) and motility readout, replicated across all four PIH family members in same study\",\n      \"pmids\": [\"29741156\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Within the R2SP complex, PIH1D2 partners with SPAG1, RUVBL1, and RUVBL2. SPAG1's TPR domains recruit HSP70 and HSP90 chaperones to the complex, and SPAG1 regulates nucleotide hydrolysis activity of HSP and RUVBL1/2 partners rather than possessing autonomous GTPase activity.\",\n      \"method\": \"Biochemical assays, isothermal titration calorimetry (ITC), NMR spectroscopy, molecular dynamics simulations, in vitro GTP hydrolysis assay\",\n      \"journal\": \"The Biochemical Journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal biochemical and structural methods (ITC, NMR, MD), single lab; directly characterizes SPAG1 binding properties within the PIH1D2-containing R2SP complex\",\n      \"pmids\": [\"31118266\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Cryo-EM, NMR, and structural mass spectrometry of the human R2SP complex (RUVBL1, RUVBL2, SPAG1, PIH1D2) reveal a three-dimensional organization similar to the canonical R2TP complex but with distinct differences in the ATPase activity of the RUVBL1/2 core and in the mode of binding of adaptors SPAG1 and PIH1D2.\",\n      \"method\": \"Cryo-EM, NMR spectroscopy, structural mass spectrometry, biochemical ATPase assays\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — structural and biochemical methods (cryo-EM, NMR, MS) are high-tier but this is a preprint, not yet peer-reviewed, single study\",\n      \"pmids\": [\"bio_10.1101_2025.01.27.635100\"],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"PIH1D2 is an adaptor subunit of the R2SP quaternary chaperone complex (together with SPAG1, RUVBL1, and RUVBL2) that promotes the folding and assembly of specific protein clients including liprin-α2 and axonemal dynein subtypes, particularly in testis and motile cilia, with SPAG1 recruiting HSP70/HSP90 to the complex and RUVBL1/2 providing ATPase activity; the overall architecture resembles the canonical R2TP complex but with distinct differences in ATPase regulation and adaptor binding modes.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"PIH1D2 is an adaptor subunit of the R2SP chaperone complex, a testis-enriched R2TP-like machine that promotes the quaternary folding and assembly of specific protein clients [#0]. Within R2SP, PIH1D2 partners with SPAG1 (an RPAP3-like protein) and the AAA+ ATPases RUVBL1 and RUVBL2, with SPAG1 binding both PIH1D2 and RUVBL1/2 and recruiting HSP70/HSP90 chaperones through its TPR domains [#0, #2]. SPAG1 itself regulates the nucleotide hydrolysis activity of its HSP and RUVBL1/2 partners rather than acting as an autonomous GTPase [#2]. The complex is required for the expression and assembly of clients such as liprin-\\u03b12 [#0], and in vivo loss of PIH1D2 abolishes the assembly of a defined subset of axonemal dynein arms in sperm flagella and Kupffer's vesicle cilia, producing abnormal motility [#1]. This links PIH1D2 to the building of motile-ciliary and flagellar dynein machinery through chaperone-assisted protein assembly.\"\n  ,\n  \"teleology\": [\n    {\n      \"year\": 2018,\n      \"claim\": \"Established that PIH1D2 is not a free-standing protein but the adaptor subunit of an R2TP-like assembly chaperone (R2SP), defining its molecular context and a first client.\",\n      \"evidence\": \"Systematic co-IP/pulldown interaction analyses, RPAP3 C-terminal domain structure, mass-spec client identification, and depletion with liprin-\\u03b12 assembly readout\",\n      \"pmids\": [\"29844425\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full client repertoire beyond liprin-\\u03b12 not defined\", \"Mechanism by which PIH1D2 selects clients not resolved\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Demonstrated an in vivo function for PIH1D2 in assembling specific axonemal dynein subtypes, connecting the chaperone adaptor to motile cilia/flagellar biology.\",\n      \"evidence\": \"Zebrafish loss-of-function mutants with cryo-electron tomography of sperm axonemal dynein and motility assays\",\n      \"pmids\": [\"29741156\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Which dynein subunits are direct R2SP clients not established\", \"Relationship to R2SP-mediated folding in this context not directly shown\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Clarified how chaperone activity is recruited and regulated within the PIH1D2 complex, showing SPAG1 brings in HSP70/HSP90 and tunes hydrolysis of partner ATPases.\",\n      \"evidence\": \"ITC, NMR, molecular dynamics, and in vitro GTP hydrolysis assays on SPAG1 within the R2SP complex\",\n      \"pmids\": [\"31118266\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab biochemical characterization\", \"PIH1D2's own contribution to ATPase regulation not isolated\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Provided a structural model of the human R2SP complex, showing how PIH1D2 and SPAG1 dock onto the RUVBL1/2 core and how its ATPase regulation differs from canonical R2TP.\",\n      \"evidence\": \"Cryo-EM, NMR, structural mass spectrometry, and biochemical ATPase assays of human R2SP (preprint)\",\n      \"pmids\": [\"bio_10.1101_2025.01.27.635100\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Preprint, not peer-reviewed\", \"Client-bound structure not determined\", \"Functional consequence of distinct ATPase regulation untested\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How PIH1D2 recognizes and selects its diverse clients (liprin-\\u03b12 versus axonemal dyneins) and how client specificity is encoded in the complex remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No client-recognition determinant mapped on PIH1D2\", \"Tissue-specific regulation of R2SP activity uncharacterized\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 2]},\n      {\"term_id\": \"GO:0044183\", \"supporting_discovery_ids\": [0, 2]}\n    ],\n    \"localization\": [],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"complexes\": [\"R2SP\"],\n    \"partners\": [\"SPAG1\", \"RUVBL1\", \"RUVBL2\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"faith_supported":4,"faith_total":4,"faith_pct":100.0}}