{"gene":"AKAP10","run_date":"2026-06-09T22:02:43","timeline":{"discoveries":[{"year":1997,"finding":"D-AKAP2 (AKAP10) binds both type I (RIα) and type II (RIIα) regulatory subunits of PKA via a 40-residue C-terminal R-binding domain that interacts with the N-terminal dimerization domain of RIα and RIIα, making it a dual-specific AKAP. A putative RGS domain was identified near the N-terminal region.","method":"Yeast two-hybrid screen; coprecipitation assays","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — yeast two-hybrid plus coprecipitation, single lab, two orthogonal methods","pmids":["9326583"],"is_preprint":false},{"year":2001,"finding":"Full-length human D-AKAP2 (AKAP10, 662 residues) localizes predominantly to mitochondria, as demonstrated by immunocytochemistry, immunohistochemistry, and tissue fractionation in mouse, rat, and human cells. In vivo association with PKA in mouse brain was confirmed by cAMP-agarose pull-down.","method":"Immunocytochemistry, immunohistochemistry, subcellular fractionation, cAMP-agarose pull-down","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal localization methods (fractionation, ICC, IHC) across three species, plus pull-down confirming PKA interaction in vivo","pmids":["11248059"],"is_preprint":false},{"year":2002,"finding":"Deuterium exchange-mass spectrometry and limited proteolysis revealed that D-AKAP2 has two distinctly folded domains: an N-terminal putative RGS domain and a C-terminal region containing a highly protected PKA binding site and a solvent-accessible PDZ binding motif, flanked by disordered regions.","method":"Deuterium exchange-mass spectrometry (DXMS); limited proteolysis","journal":"Journal of molecular biology","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — DXMS structural method with limited proteolysis validation, single lab","pmids":["12206784"],"is_preprint":false},{"year":2003,"finding":"D-AKAP2 binds PDZK1 (and to a lesser extent NHERF-1) through its C-terminal PDZ binding motif, anchoring PKA to these scaffold proteins in renal proximal tubular cells. The interaction was confirmed by pull-down experiments and co-immunoprecipitation from transfected opossum kidney cells.","method":"Yeast two-hybrid (initial identification); pull-down assays; co-immunoprecipitation from transfected cells","journal":"Kidney international","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal pull-down and co-IP in cells, single lab, two orthogonal methods","pmids":["14531807"],"is_preprint":false},{"year":2007,"finding":"Heterozygous disruption of the Akap10 C-terminus (final 51 aa) in mice increases cardiac response to cholinergic signals and causes cardiac arrhythmias and premature death, establishing AKAP10 as a regulator of heart rhythm via the cholinergic/autonomic pathway.","method":"Gene-trap disruption in mouse embryonic stem cells differentiated to cardiac myocytes; in vivo mouse cardiac physiology","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function with defined cardiac phenotype in both cell and whole-animal models, single lab","pmids":["17485678"],"is_preprint":false},{"year":2009,"finding":"The two tandem RGS domains of D-AKAP2 interact with Rab11 and GTP-bound Rab4 (the first demonstration of RGS domains binding small GTPases). D-AKAP2 regulates endocytic recycling: knockdown by RNAi redistributes Rab11 and transferrin receptor to the cell periphery and increases the rate of transferrin recycling, indicating D-AKAP2 promotes accumulation of recycling cargo in the Rab4/Rab11-positive endocytic recycling compartment.","method":"Co-immunoprecipitation; RNAi knockdown with transferrin recycling assay; overexpression/co-localization imaging","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal binding assays plus functional RNAi knockdown with quantitative recycling assay, multiple orthogonal methods in one study","pmids":["19797056"],"is_preprint":false},{"year":2010,"finding":"Crystal structure of the D-AKAP2 AKB helix in complex with the RIα D/D domain revealed a novel helical register shift compared to the RIIα:D-AKAP2 complex, explaining the molecular basis for D-AKAP2 dual-specificity. The RIα D/D domain presents an extensive surface through a well-formed N-terminal helix, and a redox-sensitive disulfide in RIα affects AKAP binding.","method":"X-ray crystallography of RIα D/D domain alone and in complex with D-AKAP2 AKB peptide","journal":"Structure (London, England : 1993)","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure with structural comparison to RIIα complex, rigorous mechanistic explanation of dual specificity","pmids":["20159461"],"is_preprint":false},{"year":2014,"finding":"Crystal structure of the D-AKAP2:PKA RII:PDZK1 ternary complex showed that the disordered C-terminal segment of D-AKAP2 becomes ordered upon binding, presenting an α-helix to PKA RII and a β-strand to PDZK1. Formation of the D-AKAP2:PKA binary complex is a prerequisite for high-affinity interaction with PDZK1, nucleating a polyvalent scaffold that links PKA signaling to transporter regulation.","method":"X-ray crystallography of ternary complex; structural and binding analysis","journal":"Protein science : a publication of the Protein Society","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure of ternary complex with mechanistic insight into ordered binding and scaffold nucleation","pmids":["25348485"],"is_preprint":false}],"current_model":"AKAP10 (D-AKAP2) is a dual-specific A-kinase anchoring protein that localizes predominantly to mitochondria and endocytic compartments; it anchors both PKA-RI and PKA-RII via a C-terminal amphipathic helix (whose dual specificity is explained by a novel helical register with RIα versus RIIα, as resolved by crystallography), recruits PDZK1/NHERF-1 through its PDZ-binding motif to form a polyvalent signaling scaffold linking PKA to membrane transporters, and engages Rab4/Rab11 small GTPases through its tandem RGS domains to regulate endocytic recycling of the transferrin receptor; loss of AKAP10 function in mice disrupts cholinergic cardiac signaling, causing arrhythmias."},"narrative":{"mechanistic_narrative":"AKAP10 (D-AKAP2) is a dual-specific A-kinase anchoring protein that organizes compartmentalized PKA signaling at mitochondria and the endocytic recycling system [PMID:11248059, PMID:19797056]. It anchors both type I (RIα) and type II (RIIα) PKA regulatory subunits through a C-terminal amphipathic helix that engages the dimerization/docking domains of each subunit, with dual specificity arising from a register shift in how the AKAP helix packs against RIα versus RIIα [PMID:9326583, PMID:20159461]. Beyond PKA, AKAP10 nucleates a polyvalent membrane scaffold: a C-terminal PDZ-binding motif recruits PDZK1 (and to a lesser extent NHERF-1), and high-affinity PDZK1 engagement requires prior formation of the AKAP10:PKA complex, coupling kinase anchoring to transporter regulation [PMID:14531807, PMID:25348485]. Its N-terminal tandem RGS domains bind GTP-loaded Rab4 and Rab11 and promote accumulation of recycling cargo such as the transferrin receptor in the Rab4/Rab11-positive recycling compartment [PMID:19797056]. In vivo, disruption of the AKAP10 C-terminus in mice heightens cholinergic cardiac responses and produces arrhythmias, establishing AKAP10 as a regulator of cardiac rhythm through the autonomic signaling pathway [PMID:17485678].","teleology":[{"year":1997,"claim":"Established AKAP10 as a dual-specific AKAP, answering whether a single anchoring protein could bind both PKA isoform classes and hinting at an additional RGS module.","evidence":"Yeast two-hybrid and coprecipitation mapping the C-terminal R-binding domain against RIα and RIIα dimerization domains","pmids":["9326583"],"confidence":"Medium","gaps":["Structural basis of dual specificity not resolved","RGS domain function and binding partners unknown","Subcellular localization not addressed"]},{"year":2001,"claim":"Defined where AKAP10 acts in the cell, localizing it to mitochondria and confirming endogenous PKA association in vivo.","evidence":"Immunocytochemistry, immunohistochemistry, subcellular fractionation across three species, and cAMP-agarose pull-down from mouse brain","pmids":["11248059"],"confidence":"High","gaps":["Mechanism of mitochondrial targeting not defined","Functional consequence of mitochondrial PKA anchoring not tested"]},{"year":2002,"claim":"Resolved the domain architecture, distinguishing a folded N-terminal RGS domain from a C-terminal region carrying the protected PKA site and accessible PDZ motif within disordered flanks.","evidence":"Deuterium exchange-mass spectrometry and limited proteolysis","pmids":["12206784"],"confidence":"Medium","gaps":["No atomic structure","Functional roles of the disordered regions not established"]},{"year":2003,"claim":"Identified the PDZ-binding motif's partner, linking AKAP10/PKA to scaffold proteins that organize membrane transporters in renal tubular cells.","evidence":"Yeast two-hybrid, pull-down, and co-immunoprecipitation from transfected opossum kidney cells","pmids":["14531807"],"confidence":"Medium","gaps":["Single lab, single cell context","Hierarchy of PKA vs PDZK1 binding not yet defined"]},{"year":2007,"claim":"Connected AKAP10 to organismal physiology, showing its C-terminus regulates cholinergic cardiac signaling and heart rhythm.","evidence":"Gene-trap disruption in mouse ES-derived cardiomyocytes and in vivo cardiac physiology","pmids":["17485678"],"confidence":"Medium","gaps":["Molecular link between anchoring and arrhythmia not delineated","Specific PKA targets in cardiomyocytes not identified"]},{"year":2009,"claim":"Assigned a function to the tandem RGS domains, revealing the first RGS–small-GTPase interaction and a role in endocytic recycling.","evidence":"Co-immunoprecipitation with Rab4/Rab11 plus RNAi knockdown with quantitative transferrin recycling assay and colocalization imaging","pmids":["19797056"],"confidence":"High","gaps":["Whether GTPase activity is modulated (GAP/effector role) not resolved","Integration of recycling role with PKA anchoring not tested"]},{"year":2014,"claim":"Explained how the scaffold assembles, showing ordered binding of PKA precedes high-affinity PDZK1 recruitment to nucleate a polyvalent complex.","evidence":"X-ray crystallography of the AKAP10:PKA RII:PDZK1 ternary complex with binding analysis","pmids":["25348485"],"confidence":"High","gaps":["Full-length scaffold architecture including RGS domains not captured","Stoichiometry in native membranes not determined"]},{"year":null,"claim":"How the mitochondrial PKA anchoring, the Rab-dependent recycling function, and the transporter-scaffolding activity are coordinated within a single signaling unit remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No integrated model linking the RGS, PKA, and PDZ functions","Cardiac substrate(s) downstream of AKAP10 unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,3,7]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[5]}],"localization":[{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[1]},{"term_id":"GO:0005768","term_label":"endosome","supporting_discovery_ids":[5]}],"pathway":[],"complexes":[],"partners":["PRKAR1A","PRKAR2A","PDZK1","NHERF1","RAB11A","RAB4A"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"O43572","full_name":"A-kinase anchor protein 10, mitochondrial","aliases":["Dual specificity A kinase-anchoring protein 2","D-AKAP-2","Protein kinase A-anchoring protein 10","PRKA10"],"length_aa":662,"mass_kda":73.8,"function":"Differentially targeted protein that binds to type I and II regulatory subunits of protein kinase A and anchors them to the mitochondria or the plasma membrane. Although the physiological relevance between PKA and AKAPS with mitochondria is not fully understood, one idea is that BAD, a proapoptotic member, is phosphorylated and inactivated by mitochondria-anchored PKA. It cannot be excluded too that it may facilitate PKA as well as G protein signal transduction, by acting as an adapter for assembling multiprotein complexes. With its RGS domain, it could lead to the interaction to G-alpha proteins, providing a link between the signaling machinery and the downstream kinase (By similarity)","subcellular_location":"Mitochondrion; Membrane; Cytoplasm","url":"https://www.uniprot.org/uniprotkb/O43572/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/AKAP10","classification":"Not Classified","n_dependent_lines":9,"n_total_lines":1208,"dependency_fraction":0.0074503311258278145},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"ARFGEF1","stoichiometry":10.0},{"gene":"ARFGEF2","stoichiometry":10.0}],"url":"https://opencell.sf.czbiohub.org/search/AKAP10","total_profiled":1310},"omim":[{"mim_id":"604694","title":"A-KINASE ANCHOR PROTEIN 10; AKAP10","url":"https://www.omim.org/entry/604694"},{"mim_id":"152430","title":"LONGEVITY 1","url":"https://www.omim.org/entry/152430"},{"mim_id":"115080","title":"CARDIAC CONDUCTION DEFECT","url":"https://www.omim.org/entry/115080"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Plasma membrane","reliability":"Supported"},{"location":"Cytosol","reliability":"Supported"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/AKAP10"},"hgnc":{"alias_symbol":["D-AKAP2","PRKA10","MGC9414"],"prev_symbol":[]},"alphafold":{"accession":"O43572","domains":[{"cath_id":"1.10.167.10","chopping":"124-177_285-378","consensus_level":"medium","plddt":91.7281,"start":124,"end":378},{"cath_id":"1.10.167.10","chopping":"379-516","consensus_level":"medium","plddt":90.8638,"start":379,"end":516}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/O43572","model_url":"https://alphafold.ebi.ac.uk/files/AF-O43572-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-O43572-F1-predicted_aligned_error_v6.png","plddt_mean":64.25},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=AKAP10","jax_strain_url":"https://www.jax.org/strain/search?query=AKAP10"},"sequence":{"accession":"O43572","fasta_url":"https://rest.uniprot.org/uniprotkb/O43572.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/O43572/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/O43572"}},"corpus_meta":[{"pmid":"9326583","id":"PMC_9326583","title":"D-AKAP2, a novel protein kinase A anchoring protein with a putative RGS domain.","date":"1997","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/9326583","citation_count":198,"is_preprint":false},{"pmid":"20159461","id":"PMC_20159461","title":"Structure of D-AKAP2:PKA RI complex: insights into AKAP specificity and selectivity.","date":"2010","source":"Structure (London, England : 1993)","url":"https://pubmed.ncbi.nlm.nih.gov/20159461","citation_count":104,"is_preprint":false},{"pmid":"11248059","id":"PMC_11248059","title":"Cloning and mitochondrial localization of full-length D-AKAP2, a protein kinase A anchoring protein.","date":"2001","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/11248059","citation_count":91,"is_preprint":false},{"pmid":"12206784","id":"PMC_12206784","title":"Domain organization of D-AKAP2 revealed by enhanced deuterium exchange-mass spectrometry (DXMS).","date":"2002","source":"Journal of molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/12206784","citation_count":64,"is_preprint":false},{"pmid":"17485678","id":"PMC_17485678","title":"Gene-trapped mouse embryonic stem cell-derived cardiac myocytes and human genetics implicate AKAP10 in heart rhythm regulation.","date":"2007","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/17485678","citation_count":60,"is_preprint":false},{"pmid":"19797056","id":"PMC_19797056","title":"D-AKAP2 interacts with Rab4 and Rab11 through its RGS domains and regulates transferrin receptor recycling.","date":"2009","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/19797056","citation_count":54,"is_preprint":false},{"pmid":"14531807","id":"PMC_14531807","title":"PDZK1: II. an anchoring site for the PKA-binding protein D-AKAP2 in renal proximal tubular cells.","date":"2003","source":"Kidney international","url":"https://pubmed.ncbi.nlm.nih.gov/14531807","citation_count":48,"is_preprint":false},{"pmid":"19496216","id":"PMC_19496216","title":"AKAP10 (I646V) functional polymorphism predicts heart rate and heart rate variability in apparently healthy, middle-aged European-Americans.","date":"2009","source":"Psychophysiology","url":"https://pubmed.ncbi.nlm.nih.gov/19496216","citation_count":22,"is_preprint":false},{"pmid":"25348485","id":"PMC_25348485","title":"D-AKAP2:PKA RII:PDZK1 ternary complex structure: insights from the nucleation of a polyvalent scaffold.","date":"2014","source":"Protein science : a publication of the Protein Society","url":"https://pubmed.ncbi.nlm.nih.gov/25348485","citation_count":9,"is_preprint":false},{"pmid":"21701445","id":"PMC_21701445","title":"Possible counter effect in newborns of 1936A>G (I646V) polymorphism in the AKAP10 gene encoding A-kinase-anchoring protein 10.","date":"2011","source":"Journal of perinatology : official journal of the California Perinatal Association","url":"https://pubmed.ncbi.nlm.nih.gov/21701445","citation_count":7,"is_preprint":false},{"pmid":"26110499","id":"PMC_26110499","title":"Genetic association of AKAP10 gene polymorphism with reduced risk of preterm birth.","date":"2015","source":"Journal of perinatology : official journal of the California Perinatal Association","url":"https://pubmed.ncbi.nlm.nih.gov/26110499","citation_count":5,"is_preprint":false},{"pmid":"23092224","id":"PMC_23092224","title":"1936A→G (I646 V) polymorphism in the AKAP10 gene encoding A-kinase-anchoring protein 10 in very long-lived poles is similar to that in newborns.","date":"2012","source":"Experimental aging research","url":"https://pubmed.ncbi.nlm.nih.gov/23092224","citation_count":2,"is_preprint":false},{"pmid":"23095189","id":"PMC_23095189","title":"Polymorphism 1936A > G in the AKAP10 gene (encoding A-kinase-anchoring protein 10) is associated with higher cholesterol cord blood concentration in Polish full-term newsborns.","date":"2013","source":"Journal of perinatal medicine","url":"https://pubmed.ncbi.nlm.nih.gov/23095189","citation_count":1,"is_preprint":false},{"pmid":"22817328","id":"PMC_22817328","title":"Association of 1936A > G in AKAP10 (A-kinase anchoring protein 10) and blood pressure in Polish full-term newborns.","date":"2012","source":"Blood pressure","url":"https://pubmed.ncbi.nlm.nih.gov/22817328","citation_count":1,"is_preprint":false},{"pmid":"19462906","id":"PMC_19462906","title":"[Genotyping of AKAP10 gene 2073A/G single nucleotide polymorphism by TaqMan probe real-time PCR].","date":"2009","source":"Sichuan da xue xue bao. Yi xue ban = Journal of Sichuan University. Medical science edition","url":"https://pubmed.ncbi.nlm.nih.gov/19462906","citation_count":1,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":9364,"output_tokens":2106,"usd":0.029841,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9112,"output_tokens":2388,"usd":0.05263,"stage2_stop_reason":"end_turn"},"total_usd":0.082471,"stage1_batch_id":"msgbatch_0179Z2pxRYBsXi2xADneafRw","stage2_batch_id":"msgbatch_01LaY9ZaVdgjWArcZo9eqQE5","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1997,\n      \"finding\": \"D-AKAP2 (AKAP10) binds both type I (RIα) and type II (RIIα) regulatory subunits of PKA via a 40-residue C-terminal R-binding domain that interacts with the N-terminal dimerization domain of RIα and RIIα, making it a dual-specific AKAP. A putative RGS domain was identified near the N-terminal region.\",\n      \"method\": \"Yeast two-hybrid screen; coprecipitation assays\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — yeast two-hybrid plus coprecipitation, single lab, two orthogonal methods\",\n      \"pmids\": [\"9326583\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"Full-length human D-AKAP2 (AKAP10, 662 residues) localizes predominantly to mitochondria, as demonstrated by immunocytochemistry, immunohistochemistry, and tissue fractionation in mouse, rat, and human cells. In vivo association with PKA in mouse brain was confirmed by cAMP-agarose pull-down.\",\n      \"method\": \"Immunocytochemistry, immunohistochemistry, subcellular fractionation, cAMP-agarose pull-down\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal localization methods (fractionation, ICC, IHC) across three species, plus pull-down confirming PKA interaction in vivo\",\n      \"pmids\": [\"11248059\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Deuterium exchange-mass spectrometry and limited proteolysis revealed that D-AKAP2 has two distinctly folded domains: an N-terminal putative RGS domain and a C-terminal region containing a highly protected PKA binding site and a solvent-accessible PDZ binding motif, flanked by disordered regions.\",\n      \"method\": \"Deuterium exchange-mass spectrometry (DXMS); limited proteolysis\",\n      \"journal\": \"Journal of molecular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — DXMS structural method with limited proteolysis validation, single lab\",\n      \"pmids\": [\"12206784\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"D-AKAP2 binds PDZK1 (and to a lesser extent NHERF-1) through its C-terminal PDZ binding motif, anchoring PKA to these scaffold proteins in renal proximal tubular cells. The interaction was confirmed by pull-down experiments and co-immunoprecipitation from transfected opossum kidney cells.\",\n      \"method\": \"Yeast two-hybrid (initial identification); pull-down assays; co-immunoprecipitation from transfected cells\",\n      \"journal\": \"Kidney international\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal pull-down and co-IP in cells, single lab, two orthogonal methods\",\n      \"pmids\": [\"14531807\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Heterozygous disruption of the Akap10 C-terminus (final 51 aa) in mice increases cardiac response to cholinergic signals and causes cardiac arrhythmias and premature death, establishing AKAP10 as a regulator of heart rhythm via the cholinergic/autonomic pathway.\",\n      \"method\": \"Gene-trap disruption in mouse embryonic stem cells differentiated to cardiac myocytes; in vivo mouse cardiac physiology\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function with defined cardiac phenotype in both cell and whole-animal models, single lab\",\n      \"pmids\": [\"17485678\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"The two tandem RGS domains of D-AKAP2 interact with Rab11 and GTP-bound Rab4 (the first demonstration of RGS domains binding small GTPases). D-AKAP2 regulates endocytic recycling: knockdown by RNAi redistributes Rab11 and transferrin receptor to the cell periphery and increases the rate of transferrin recycling, indicating D-AKAP2 promotes accumulation of recycling cargo in the Rab4/Rab11-positive endocytic recycling compartment.\",\n      \"method\": \"Co-immunoprecipitation; RNAi knockdown with transferrin recycling assay; overexpression/co-localization imaging\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal binding assays plus functional RNAi knockdown with quantitative recycling assay, multiple orthogonal methods in one study\",\n      \"pmids\": [\"19797056\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Crystal structure of the D-AKAP2 AKB helix in complex with the RIα D/D domain revealed a novel helical register shift compared to the RIIα:D-AKAP2 complex, explaining the molecular basis for D-AKAP2 dual-specificity. The RIα D/D domain presents an extensive surface through a well-formed N-terminal helix, and a redox-sensitive disulfide in RIα affects AKAP binding.\",\n      \"method\": \"X-ray crystallography of RIα D/D domain alone and in complex with D-AKAP2 AKB peptide\",\n      \"journal\": \"Structure (London, England : 1993)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure with structural comparison to RIIα complex, rigorous mechanistic explanation of dual specificity\",\n      \"pmids\": [\"20159461\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Crystal structure of the D-AKAP2:PKA RII:PDZK1 ternary complex showed that the disordered C-terminal segment of D-AKAP2 becomes ordered upon binding, presenting an α-helix to PKA RII and a β-strand to PDZK1. Formation of the D-AKAP2:PKA binary complex is a prerequisite for high-affinity interaction with PDZK1, nucleating a polyvalent scaffold that links PKA signaling to transporter regulation.\",\n      \"method\": \"X-ray crystallography of ternary complex; structural and binding analysis\",\n      \"journal\": \"Protein science : a publication of the Protein Society\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure of ternary complex with mechanistic insight into ordered binding and scaffold nucleation\",\n      \"pmids\": [\"25348485\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"AKAP10 (D-AKAP2) is a dual-specific A-kinase anchoring protein that localizes predominantly to mitochondria and endocytic compartments; it anchors both PKA-RI and PKA-RII via a C-terminal amphipathic helix (whose dual specificity is explained by a novel helical register with RIα versus RIIα, as resolved by crystallography), recruits PDZK1/NHERF-1 through its PDZ-binding motif to form a polyvalent signaling scaffold linking PKA to membrane transporters, and engages Rab4/Rab11 small GTPases through its tandem RGS domains to regulate endocytic recycling of the transferrin receptor; loss of AKAP10 function in mice disrupts cholinergic cardiac signaling, causing arrhythmias.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"AKAP10 (D-AKAP2) is a dual-specific A-kinase anchoring protein that organizes compartmentalized PKA signaling at mitochondria and the endocytic recycling system [#1, #5]. It anchors both type I (RIα) and type II (RIIα) PKA regulatory subunits through a C-terminal amphipathic helix that engages the dimerization/docking domains of each subunit, with dual specificity arising from a register shift in how the AKAP helix packs against RIα versus RIIα [#0, #6]. Beyond PKA, AKAP10 nucleates a polyvalent membrane scaffold: a C-terminal PDZ-binding motif recruits PDZK1 (and to a lesser extent NHERF-1), and high-affinity PDZK1 engagement requires prior formation of the AKAP10:PKA complex, coupling kinase anchoring to transporter regulation [#3, #7]. Its N-terminal tandem RGS domains bind GTP-loaded Rab4 and Rab11 and promote accumulation of recycling cargo such as the transferrin receptor in the Rab4/Rab11-positive recycling compartment [#5]. In vivo, disruption of the AKAP10 C-terminus in mice heightens cholinergic cardiac responses and produces arrhythmias, establishing AKAP10 as a regulator of cardiac rhythm through the autonomic signaling pathway [#4].\",\n  \"teleology\": [\n    {\n      \"year\": 1997,\n      \"claim\": \"Established AKAP10 as a dual-specific AKAP, answering whether a single anchoring protein could bind both PKA isoform classes and hinting at an additional RGS module.\",\n      \"evidence\": \"Yeast two-hybrid and coprecipitation mapping the C-terminal R-binding domain against RIα and RIIα dimerization domains\",\n      \"pmids\": [\"9326583\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of dual specificity not resolved\", \"RGS domain function and binding partners unknown\", \"Subcellular localization not addressed\"]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"Defined where AKAP10 acts in the cell, localizing it to mitochondria and confirming endogenous PKA association in vivo.\",\n      \"evidence\": \"Immunocytochemistry, immunohistochemistry, subcellular fractionation across three species, and cAMP-agarose pull-down from mouse brain\",\n      \"pmids\": [\"11248059\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism of mitochondrial targeting not defined\", \"Functional consequence of mitochondrial PKA anchoring not tested\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Resolved the domain architecture, distinguishing a folded N-terminal RGS domain from a C-terminal region carrying the protected PKA site and accessible PDZ motif within disordered flanks.\",\n      \"evidence\": \"Deuterium exchange-mass spectrometry and limited proteolysis\",\n      \"pmids\": [\"12206784\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No atomic structure\", \"Functional roles of the disordered regions not established\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Identified the PDZ-binding motif's partner, linking AKAP10/PKA to scaffold proteins that organize membrane transporters in renal tubular cells.\",\n      \"evidence\": \"Yeast two-hybrid, pull-down, and co-immunoprecipitation from transfected opossum kidney cells\",\n      \"pmids\": [\"14531807\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab, single cell context\", \"Hierarchy of PKA vs PDZK1 binding not yet defined\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Connected AKAP10 to organismal physiology, showing its C-terminus regulates cholinergic cardiac signaling and heart rhythm.\",\n      \"evidence\": \"Gene-trap disruption in mouse ES-derived cardiomyocytes and in vivo cardiac physiology\",\n      \"pmids\": [\"17485678\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular link between anchoring and arrhythmia not delineated\", \"Specific PKA targets in cardiomyocytes not identified\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Assigned a function to the tandem RGS domains, revealing the first RGS–small-GTPase interaction and a role in endocytic recycling.\",\n      \"evidence\": \"Co-immunoprecipitation with Rab4/Rab11 plus RNAi knockdown with quantitative transferrin recycling assay and colocalization imaging\",\n      \"pmids\": [\"19797056\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether GTPase activity is modulated (GAP/effector role) not resolved\", \"Integration of recycling role with PKA anchoring not tested\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Explained how the scaffold assembles, showing ordered binding of PKA precedes high-affinity PDZK1 recruitment to nucleate a polyvalent complex.\",\n      \"evidence\": \"X-ray crystallography of the AKAP10:PKA RII:PDZK1 ternary complex with binding analysis\",\n      \"pmids\": [\"25348485\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full-length scaffold architecture including RGS domains not captured\", \"Stoichiometry in native membranes not determined\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the mitochondrial PKA anchoring, the Rab-dependent recycling function, and the transporter-scaffolding activity are coordinated within a single signaling unit remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No integrated model linking the RGS, PKA, and PDZ functions\", \"Cardiac substrate(s) downstream of AKAP10 unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 3, 7]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [5]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [1]},\n      {\"term_id\": \"GO:0005768\", \"supporting_discovery_ids\": [5]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0005215\", \"supporting_discovery_ids\": []}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"PRKAR1A\", \"PRKAR2A\", \"PDZK1\", \"NHERF1\", \"RAB11A\", \"RAB4A\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}