{"gene":"AKAP6","run_date":"2026-06-09T22:02:43","timeline":{"discoveries":[{"year":1995,"finding":"AKAP100 (AKAP6) was identified as a PKA type II regulatory subunit (RII)-binding protein that targets PKA type II to the sarcoplasmic reticulum in cardiac and skeletal muscle cells, demonstrated by RII overlay assay and co-purification with RII from cell extracts.","method":"Interaction cloning with RII probe, solid-phase overlay assay, immunohistochemistry, cAMP-agarose affinity co-purification","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (interaction cloning, overlay assay, affinity purification, immunohistochemistry) in foundational study, independently replicated in subsequent work","pmids":["7721854"],"is_preprint":false},{"year":1998,"finding":"AKAP100 (AKAP6) localizes to multiple subcellular compartments in adult rat cardiomyocytes including the nucleus, sarcolemma, intercalated disc, Z-line, and transverse tubule/junctional SR; RII (but not RI) co-localizes with AKAP100 at these sites.","method":"Immunofluorescence, confocal microscopy, double immunostaining with alpha-actinin and ryanodine receptor antibodies","journal":"The Journal of cell biology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — localization established by immunofluorescence/confocal across multiple structures, single lab but multiple markers","pmids":["9679148"],"is_preprint":false},{"year":1999,"finding":"mAKAP (AKAP6) is targeted to the nuclear membrane of differentiated myocytes; nuclear membrane targeting is conferred by two regions (residues 772–915 and 915–1065) containing spectrin-like repeat sequences, as shown by GFP fusion constructs and displacement experiments.","method":"GFP fusion construct expression in myocytes, heterologous overexpression of targeting domains to displace endogenous mAKAP, immunolocalization","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 2 / Strong — domain mapping with GFP fusions and competitive displacement, replicated in subsequent studies confirming nuclear envelope localization","pmids":["10413680"],"is_preprint":false},{"year":2001,"finding":"mAKAP (AKAP6) assembles a cAMP signaling module containing PKA and PDE4D3 at the nuclear envelope of cardiac myocytes; tonic PDE4D3 activity reduces anchored PKA activity, while PKA activation stimulates mAKAP-associated PDE4D3, forming a negative feedback loop; disruption of PKA–mAKAP interaction prevents PKA-mediated enhancement of PDE4D3 activity.","method":"Co-immunoprecipitation from heart tissue, functional kinase and phosphodiesterase assays, PKA-mAKAP interaction disruption experiments","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — biochemical reconstitution of complex in native tissue, functional assays with disruption mutants, multiple orthogonal approaches","pmids":["11296225"],"is_preprint":false},{"year":2001,"finding":"The mAKAP (AKAP6) complex at the cardiac nuclear envelope also includes ryanodine receptors and protein phosphatase 2A; a subset of cardiac ryanodine receptor binds to mAKAP at the nuclear envelope, potentially enabling PKA-mediated phosphorylation of ryanodine receptor.","method":"Co-immunoprecipitation, immunohistochemistry, tissue fractionation","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — co-IP and fractionation from cardiac tissue, single lab, two orthogonal methods","pmids":["11590243"],"is_preprint":false},{"year":2003,"finding":"Anchoring of PKA by mAKAP (AKAP6) at the sarcoplasmic reticulum co-localizes with RyR1 and increases PKA-dependent phosphorylation of RyR1 and Ca2+ efflux through RyR1; a PKA-binding-deficient mAKAP mutant (mAKAP-P) fails to enhance RyR1 phosphorylation or Ca2+ transient amplitude.","method":"Immunoelectron microscopy, Ca2+ transient measurements, phosphorylation assays in CHO cells stably expressing RyR1, expression of wild-type vs. PKA-binding-deficient mAKAP mutant","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — structure-function analysis with loss-of-function mutant, multiple readouts (phosphorylation + Ca2+ flux), replicated conceptually in cardiac studies","pmids":["12709444"],"is_preprint":false},{"year":2004,"finding":"PKA phosphorylation of PDE4D3 on Ser-13 increases the affinity of PDE4D3 for mAKAP (AKAP6), facilitating recruitment of PDE4D3 to the mAKAP signaling complex for faster cAMP signal termination.","method":"In vitro phosphorylation assays, co-immunoprecipitation, site-directed mutagenesis of PDE4D3 Ser-13, cellular experiments","journal":"The Biochemical journal","confidence":"High","confidence_rationale":"Tier 1 / Strong — site-directed mutagenesis identifying specific phosphorylation site combined with in vitro and cellular binding assays","pmids":["15182229"],"is_preprint":false},{"year":2005,"finding":"mAKAP (AKAP6) coordinates two integrated cAMP effector pathways: anchored PKA stimulates PDE4D3 to reduce local cAMP, while an mAKAP-associated ERK5 module suppresses PDE4D3; PDE4D3 also recruits Epac1 to enable cAMP-dependent attenuation of ERK5; anchored ERK5 can induce cardiomyocyte hypertrophy.","method":"Co-immunoprecipitation, pharmacological inhibitors, RNA interference, dominant-negative constructs, cardiomyocyte hypertrophy assays","journal":"Nature","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (co-IP, RNAi, pharmacology, dominant-negative), replicated conceptually across multiple labs","pmids":["16177794"],"is_preprint":false},{"year":2005,"finding":"Nesprin-1alpha serves as a receptor for mAKAP (AKAP6) on the nuclear envelope; the amino-terminal dimerization domain of nesprin-1alpha directly binds the third spectrin repeat of mAKAP to target it to the nuclear envelope; overexpression of these spectrin repeat domains displaces mAKAP from nesprin-1alpha.","method":"Co-immunoprecipitation, direct binding assays, overexpression of spectrin repeat domains, displacement experiments in myocytes","journal":"Experimental cell research","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct binding assays plus in-cell displacement experiments, domain-level mapping, mechanistically validated","pmids":["15652351"],"is_preprint":false},{"year":2005,"finding":"The mAKAP (AKAP6) complex facilitates PKA-catalyzed phosphorylation of the ryanodine receptor Ca2+-release channel and is required for adrenergic-mediated cardiomyocyte hypertrophy; calcineurin Abeta associates with mAKAP and formation of the mAKAP complex is required for full activation of the pro-hypertrophic transcription factor NFATc.","method":"RNA interference of mAKAP, expression of PKA-binding-deficient mAKAP mutant, ryanodine receptor inhibition, co-immunoprecipitation of calcineurin, NFATc reporter assays","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 2 / Strong — RNAi loss-of-function, mutant rescue, co-IP, and transcription factor reporter assays — multiple orthogonal approaches in one study","pmids":["16306226"],"is_preprint":false},{"year":2008,"finding":"mAKAP (AKAP6) organizes ubiquitin E3 ligases that control the stability of HIF-1alpha near the nuclear envelope; depletion of mAKAP or disruption of its perinuclear targeting alters HIF-1alpha stability and transcriptional activation of hypoxia-responsive genes in cardiomyocytes.","method":"mAKAP depletion (RNAi), disruption of perinuclear targeting, HIF-1alpha stability assays, transcriptional reporter assays, co-immunoprecipitation of E3 ligases","journal":"Science signaling","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function and targeting disruption with defined transcriptional readout, single lab","pmids":["19109240"],"is_preprint":false},{"year":2010,"finding":"PP2A associated with mAKAP (AKAP6) complexes (containing B56delta subunit) dephosphorylates PDE4D3 at Ser-54, reversing PKA-mediated activation of PDE4D3; a C-terminal mAKAP domain (residues 2085–2319) binds PP2A; PKA phosphorylation of B56delta enhances PP2A activity 2-fold in the complex, creating a cAMP-induced positive feedback loop.","method":"Domain mapping, co-immunoprecipitation, phosphatase activity assays, site-directed mutagenesis of B56delta PKA phosphorylation site, deletion of mAKAP C-terminal domain","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — domain mapping with deletion mutants, phosphorylation site mutagenesis, enzymatic activity assays, multiple orthogonal methods in one study","pmids":["20106966"],"is_preprint":false},{"year":2012,"finding":"mAKAP (AKAP6) organizes a calcineurin/MEF2 signaling complex in myocytes; a calcineurin/mAKAP/MEF2 complex can be isolated from C2C12 cells and cardiac myocytes; calcineurin–MEF2 association is dependent on mAKAP expression; disruption of calcineurin–mAKAP binding blunts MEF2 transcriptional activity during myoblast differentiation and inhibits adrenergic-induced cardiac hypertrophy.","method":"Co-immunoprecipitation from C2C12 cells and cardiac myocytes, dominant-interference peptide disrupting calcineurin–mAKAP binding, MEF2 transcriptional reporter assays, siRNA knockdown","journal":"Experimental cell research","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal co-IP, dominant-interference peptide with defined binding domain, transcriptional readout, and hypertrophy phenotype — multiple orthogonal methods","pmids":["23261540"],"is_preprint":false},{"year":2012,"finding":"mAKAP (AKAP6) directly binds MEF2 through discrete interaction domains; disruption of MEF2–mAKAP binding blocks MEF2 activation during early myoblast differentiation and inhibits myotube formation and expression of differentiation markers.","method":"Direct binding assays identifying discrete binding domains, dominant-interference expression, MEF2 transcriptional reporter assays, myotube formation assays","journal":"Cellular signalling","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct binding domain mapping and loss-of-function with defined phenotype, single lab","pmids":["22484155"],"is_preprint":false},{"year":2013,"finding":"Human mAKAP coding polymorphism P1400S (in the PDE4D3 binding site) reduces mAKAP–PDE4D3 binding without affecting PKA binding or activity; S2195F (near the PP2A binding site) increases PKA binding and PKA activity; L717V (flanking the spectrin repeat domain) increases PKA binding without changing PKA activity.","method":"Site-directed mutagenesis of mAKAP, co-immunoprecipitation, surface plasmon resonance (Biacore), PKA activity assays, Ca2+ measurements","journal":"Journal of molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — mutagenesis of specific residues with orthogonal binding methods (co-IP + SPR) and functional assays in single lab","pmids":["23806656"],"is_preprint":false},{"year":2015,"finding":"AKAP6 knockdown in skeletal myoblasts halts myotube formation and decreases myogenin and myosin heavy chain expression; AKAP6 promotes myogenin expression through MEF2A; myogenin in turn binds an E-box site on the AKAP6 promoter to increase AKAP6 expression, forming a positive feedback loop.","method":"siRNA knockdown, shRNA lentiviral delivery in vivo (cardiotoxin muscle injury model), chromatin immunoprecipitation, luciferase reporter assays, motor function assessment","journal":"Scientific reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vitro and in vivo loss-of-function, ChIP identifying direct promoter binding, luciferase reporter, multiple orthogonal methods","pmids":["26563778"],"is_preprint":false},{"year":2019,"finding":"AKAP6 physically interacts with phospholamban (PLN) at the perinuclear SR region in HEK-293T cells and cardiomyocytes; AKAP6 promotes Ca2+ uptake activity of SERCA1 in cotransfected cells even in the presence of PLN.","method":"Immunofluorescence colocalization, co-immunoprecipitation from HEK-293T cells and adult rat cardiomyocytes, Ca2+ uptake assay","journal":"Physiological reports","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — co-IP from two cell systems plus functional Ca2+ uptake assay, single lab","pmids":["31325238"],"is_preprint":false},{"year":2020,"finding":"AKAP6 is a key organizer of the nuclear envelope MTOC (NE-MTOC) in cardiomyocytes and osteoclasts; AKAP6 anchors centrosomal proteins (Pcnt, AKAP9) to the nuclear envelope via its spectrin repeats, acting as an adaptor between nesprin-1alpha and Pcnt/AKAP9; AKAP6 and AKAP9 form a protein platform tethering the Golgi to the nucleus; ectopic AKAP6 expression in epithelial cells is sufficient to recruit endogenous centrosomal proteins; AKAP6 is required for cardiomyocyte hypertrophy and osteoclast bone resorption.","method":"Loss-of-function (siRNA/shRNA), gain-of-function (ectopic expression), co-immunoprecipitation, immunofluorescence in cardiomyocytes and osteoclasts, microtubule nucleation assays","journal":"eLife","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (co-IP, gain/loss-of-function, MT nucleation), multiple cell types, functional phenotype (hypertrophy, bone resorption)","pmids":["33295871"],"is_preprint":false},{"year":2021,"finding":"Myogenin transcription factor is required and sufficient for NE-MTOC formation and induces AKAP6 expression; overexpression of AKAP6beta and nesprin-1alpha together is sufficient to recruit endogenous MTOC proteins to the nuclear envelope of myoblasts in the absence of myogenin, identifying AKAP6 as a central effector of myogenin-controlled NE-MTOC assembly.","method":"Loss-of-function (myogenin KO), gain-of-function (overexpression in fibroblasts and myoblasts), promoter reporter assays, bioinformatics, immunofluorescence","journal":"eLife","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic epistasis (myogenin KO), rescue by AKAP6 overexpression, promoter studies, multiple cell types and methods","pmids":["34605406"],"is_preprint":false},{"year":2024,"finding":"AKAP6 anchors calcineurin (CaN) and NFATc4 in neurons, and this scaffolding is required for BDNF-mediated NFATc4 transcriptional activity and neuroprotection; disruption of calcineurin anchoring to AKAP6 diminishes BDNF's pro-survival effect; NFATc4 knockout mice show reduced BDNF neuroprotection in vivo.","method":"AKAP6 disruption experiments (calcineurin anchoring domain peptide), NFAT transcriptional reporter assays, NFATc4 knockout mice, neuronal survival assays","journal":"Molecular brain","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function (dominant interference + KO mouse), transcriptional reporter, in vivo validation — single lab","pmids":["39578909"],"is_preprint":false},{"year":2025,"finding":"Wnt/β-catenin pathway transcriptionally upregulates AKAP6 expression in cardiomyocytes; increased AKAP6 enhances PKA-mediated RyR2 phosphorylation, causing sarcoplasmic reticulum Ca2+ leakage and cardiomyocyte dysfunction.","method":"Wnt pathway activation, transcriptome analysis, AKAP6 overexpression/knockdown, RyR2 phosphorylation assays, Ca2+ measurements","journal":"Journal of molecular cell biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — transcriptomics identification combined with functional validation of AKAP6-RyR2 axis and Ca2+ readout, single lab","pmids":["40097291"],"is_preprint":false}],"current_model":"AKAP6 (mAKAP) is a scaffold protein anchored to the nuclear envelope of striated myocytes via its spectrin repeats binding nesprin-1alpha, where it organizes a large multi-protein signalosome integrating cAMP (PKA, PDE4D3, Epac1), Ca2+ (ryanodine receptor, calcineurin), MAP kinase (ERK5), PP2A, HIF-1alpha ubiquitin E3 ligases, and transcription factors (NFATc, MEF2) to coordinate cardiomyocyte hypertrophy, myoblast differentiation, and hypoxic gene responses; additionally, AKAP6 serves as the key organizer of the nuclear envelope microtubule-organizing center by bridging nesprin-1alpha to centrosomal proteins (Pcnt, AKAP9) and tethering the Golgi to the nucleus."},"narrative":{"mechanistic_narrative":"AKAP6 (mAKAP) is a large scaffold protein that organizes a multi-protein signalosome at the nuclear envelope of striated myocytes to integrate cAMP, Ca2+, and MAP-kinase signals controlling cardiomyocyte hypertrophy and myocyte differentiation [PMID:11296225, PMID:16177794, PMID:16306226]. It was first identified as a PKA type II regulatory subunit-binding protein targeting PKA to the sarcoplasmic reticulum of cardiac and skeletal muscle [PMID:7721854], and is anchored to the nuclear membrane through spectrin-repeat regions that bind the amino-terminal dimerization domain of nesprin-1alpha [PMID:10413680, PMID:15652351]. At this site AKAP6 nucleates a cAMP module containing PKA and PDE4D3, where PKA stimulates PDE4D3 and PKA-phosphorylation of PDE4D3 (Ser-13) raises its affinity for the scaffold, establishing a negative feedback loop for cAMP termination [PMID:11296225, PMID:15182229]; an associated PP2A/B56delta phosphatase reverses PDE4D3 activation as a counterbalancing positive-feedback arm [PMID:20106966], and an ERK5/Epac1 module couples this network to hypertrophic growth [PMID:16177794]. The complex localizes ryanodine receptor and calcineurin to enable PKA-mediated RyR phosphorylation, Ca2+ release, and calcineurin-dependent activation of the pro-hypertrophic transcription factor NFATc [PMID:12709444, PMID:16306226]. AKAP6 directly binds MEF2 and organizes calcineurin/MEF2 signaling to drive myoblast differentiation, with myogenin feeding back to upregulate AKAP6 transcription [PMID:23261540, PMID:22484155, PMID:26563778]. It also organizes ubiquitin E3 ligases that govern HIF-1alpha stability and hypoxic gene responses [PMID:19109240]. Independently of its kinase-anchoring role, AKAP6 is the central organizer of the nuclear-envelope microtubule-organizing center, bridging nesprin-1alpha to the centrosomal proteins pericentrin and AKAP9 and tethering the Golgi to the nucleus, a function induced downstream of myogenin [PMID:33295871, PMID:34605406]. Beyond striated muscle, AKAP6 anchors calcineurin and NFATc4 in neurons to mediate BDNF-dependent transcription and neuroprotection [PMID:39578909].","teleology":[{"year":1995,"claim":"Established AKAP6 as a PKA-anchoring protein, defining its founding molecular activity of targeting PKA type II to muscle sarcoplasmic reticulum.","evidence":"Interaction cloning with RII probe, overlay assay, and cAMP-agarose co-purification from muscle extracts","pmids":["7721854"],"confidence":"High","gaps":["Did not define the nuclear-envelope targeting mechanism","No downstream substrates identified at this stage"]},{"year":1999,"claim":"Mapped the spectrin-repeat regions responsible for nuclear-membrane targeting, locating the scaffold to a defined subcellular compartment in differentiated myocytes.","evidence":"GFP fusion constructs and competitive displacement of endogenous mAKAP in myocytes","pmids":["10413680"],"confidence":"High","gaps":["The nuclear-envelope receptor binding these repeats was not yet identified"]},{"year":2005,"claim":"Identified nesprin-1alpha as the nuclear-envelope receptor, explaining how AKAP6 is physically anchored to the perinuclear membrane.","evidence":"Direct binding assays and in-cell displacement mapping nesprin-1alpha N-terminus to the third spectrin repeat of mAKAP","pmids":["15652351"],"confidence":"High","gaps":["Structural basis of the spectrin-repeat/nesprin interface not resolved"]},{"year":2006,"claim":"Defined the core cAMP-handling logic of the signalosome: a PKA/PDE4D3 module with reciprocal feedback regulating local cAMP.","evidence":"Co-IP from heart tissue, kinase/PDE activity assays, PKA-binding disruption, and PDE4D3 Ser-13 mutagenesis","pmids":["11296225","15182229"],"confidence":"High","gaps":["How the cAMP module integrates with Ca2+ and MAPK arms not yet defined"]},{"year":2005,"claim":"Connected the scaffold to Ca2+ handling and hypertrophy, showing PKA anchoring enhances RyR phosphorylation and Ca2+ flux and that mAKAP is required for adrenergic hypertrophy via calcineurin/NFATc.","evidence":"PKA-binding-deficient mutant, Ca2+ transient and phosphorylation assays, RNAi, calcineurin co-IP, and NFATc reporters in cardiomyocytes","pmids":["12709444","16306226","11590243"],"confidence":"High","gaps":["Stoichiometry of RyR recruitment to nuclear-envelope mAKAP unclear","RyR co-IP/fractionation results from a single lab"]},{"year":2005,"claim":"Revealed integration of MAPK signaling, with an ERK5 module and Epac1 coupling cAMP to hypertrophic growth control through the scaffold.","evidence":"Co-IP, RNAi, pharmacology, dominant-negatives, and cardiomyocyte hypertrophy assays","pmids":["16177794"],"confidence":"High","gaps":["Quantitative contribution of ERK5 versus calcineurin arms to hypertrophy not separated"]},{"year":2010,"claim":"Added a PP2A/B56delta counter-regulatory arm and extended the scaffold to hypoxic signaling via HIF-1alpha-controlling E3 ligases.","evidence":"Domain mapping, phosphatase activity assays, B56delta mutagenesis, and HIF-1alpha stability/reporter assays with mAKAP depletion","pmids":["20106966","19109240"],"confidence":"Medium","gaps":["Identity of the HIF-1alpha E3 ligases not fully defined","HIF-1alpha findings from a single lab"]},{"year":2015,"claim":"Defined AKAP6's role in myocyte differentiation through direct MEF2/calcineurin binding and a myogenin feedback loop driving its own expression.","evidence":"MEF2 binding-domain mapping, dominant-interference peptides, ChIP of myogenin at the AKAP6 promoter, reporter and myotube assays, in vivo shRNA","pmids":["23261540","22484155","26563778"],"confidence":"High","gaps":["Discrete MEF2-binding domain structure not resolved"]},{"year":2021,"claim":"Established a kinase-anchoring-independent function: AKAP6 as the central organizer of the nuclear-envelope MTOC, bridging nesprin-1alpha to centrosomal proteins and Golgi, downstream of myogenin.","evidence":"Loss/gain-of-function, co-IP, microtubule nucleation, and myogenin KO/rescue in cardiomyocytes, osteoclasts, and myoblasts","pmids":["33295871","34605406"],"confidence":"High","gaps":["How NE-MTOC scaffolding and the signalosome roles are partitioned on the same protein is unresolved"]},{"year":2024,"claim":"Extended AKAP6 scaffolding beyond muscle, showing it anchors calcineurin/NFATc4 in neurons for BDNF-dependent transcription and neuroprotection.","evidence":"Calcineurin-anchoring disruption peptide, NFAT reporters, NFATc4 KO mice, and neuronal survival assays","pmids":["39578909"],"confidence":"Medium","gaps":["Neuronal localization of AKAP6 not mapped","Findings from a single lab"]},{"year":2025,"claim":"Placed AKAP6 downstream of Wnt/beta-catenin, linking its transcriptional upregulation to pathological RyR2 hyperphosphorylation and Ca2+ leak.","evidence":"Wnt activation, transcriptomics, AKAP6 over/knockdown, and RyR2 phosphorylation/Ca2+ assays in cardiomyocytes","pmids":["40097291"],"confidence":"Medium","gaps":["Direct Wnt-responsive element on AKAP6 promoter not confirmed","Single lab"]},{"year":null,"claim":"How the same scaffold molecule simultaneously coordinates the cAMP/Ca2+ signalosome and the NE-MTOC, and whether disease-associated coding variants alter these functions in vivo, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structural model of the assembled signalosome","In vivo phenotype of AKAP6 polymorphisms not established","Mechanism dividing scaffold pools between functions unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,3,8,13,17]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[3,11,9]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[2,8,17]}],"localization":[{"term_id":"GO:0005635","term_label":"nuclear envelope","supporting_discovery_ids":[2,8,10]},{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[0,5,16]},{"term_id":"GO:0005815","term_label":"microtubule organizing center","supporting_discovery_ids":[17]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[1,2]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[3,7,9]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[13,15,18]},{"term_id":"R-HSA-1852241","term_label":"Organelle biogenesis and maintenance","supporting_discovery_ids":[17,18]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[9,10,12]}],"complexes":["mAKAP signalosome (nuclear envelope)","nuclear-envelope MTOC (NE-MTOC)"],"partners":["PRKAR2/PKA","PDE4D3","RYR2","PPP3CB (CALCINEURIN)","PPP2 (PP2A B56DELTA)","SYNE1 (NESPRIN-1ALPHA)","PCNT","AKAP9"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q13023","full_name":"A-kinase anchor protein 6","aliases":["A-kinase anchor protein 100 kDa","AKAP 100","Protein kinase A-anchoring protein 6","PRKA6","mAKAP"],"length_aa":2319,"mass_kda":256.7,"function":"Binds to type II regulatory subunits of protein kinase A and anchors/targets them to the nuclear membrane or sarcoplasmic reticulum. May act as an adapter for assembling multiprotein complexes","subcellular_location":"Sarcoplasmic reticulum; Nucleus membrane","url":"https://www.uniprot.org/uniprotkb/Q13023/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/AKAP6","classification":"Not Classified","n_dependent_lines":2,"n_total_lines":1208,"dependency_fraction":0.0016556291390728477},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/AKAP6","total_profiled":1310},"omim":[{"mim_id":"606057","title":"RAP GUANINE NUCLEOTIDE EXCHANGE FACTOR 3; RAPGEF3","url":"https://www.omim.org/entry/606057"},{"mim_id":"604691","title":"A-KINASE ANCHOR PROTEIN 6; AKAP6","url":"https://www.omim.org/entry/604691"},{"mim_id":"600620","title":"FK506-BINDING PROTEIN 1B; FKBP1B","url":"https://www.omim.org/entry/600620"},{"mim_id":"600129","title":"PHOSPHODIESTERASE 4D; PDE4D","url":"https://www.omim.org/entry/600129"},{"mim_id":"188830","title":"PROTEIN KINASE, cAMP-DEPENDENT, REGULATORY, TYPE I, ALPHA; PRKAR1A","url":"https://www.omim.org/entry/188830"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"heart muscle","ntpm":51.1},{"tissue":"skeletal muscle","ntpm":44.4},{"tissue":"tongue","ntpm":47.4}],"url":"https://www.proteinatlas.org/search/AKAP6"},"hgnc":{"alias_symbol":["KIAA0311","mAKAP","AKAP100","PRKA6","ADAP6"],"prev_symbol":[]},"alphafold":{"accession":"Q13023","domains":[{"cath_id":"1.20.58.60","chopping":"766-938","consensus_level":"medium","plddt":79.971,"start":766,"end":938},{"cath_id":"1.20.58.60","chopping":"963-1068","consensus_level":"medium","plddt":79.1276,"start":963,"end":1068},{"cath_id":"1.20.58.60","chopping":"1075-1226","consensus_level":"medium","plddt":69.8801,"start":1075,"end":1226},{"cath_id":"1.20.58","chopping":"61-170","consensus_level":"medium","plddt":70.6499,"start":61,"end":170}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q13023","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q13023-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q13023-F1-predicted_aligned_error_v6.png","plddt_mean":42.16},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=AKAP6","jax_strain_url":"https://www.jax.org/strain/search?query=AKAP6"},"sequence":{"accession":"Q13023","fasta_url":"https://rest.uniprot.org/uniprotkb/Q13023.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q13023/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q13023"}},"corpus_meta":[{"pmid":"16177794","id":"PMC_16177794","title":"The protein kinase A anchoring protein mAKAP coordinates two integrated cAMP effector pathways.","date":"2005","source":"Nature","url":"https://pubmed.ncbi.nlm.nih.gov/16177794","citation_count":450,"is_preprint":false},{"pmid":"11296225","id":"PMC_11296225","title":"mAKAP assembles a protein kinase A/PDE4 phosphodiesterase cAMP signaling module.","date":"2001","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/11296225","citation_count":389,"is_preprint":false},{"pmid":"10413680","id":"PMC_10413680","title":"mAKAP: an A-kinase anchoring protein targeted to the nuclear membrane of differentiated myocytes.","date":"1999","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/10413680","citation_count":154,"is_preprint":false},{"pmid":"11590243","id":"PMC_11590243","title":"mAKAP and the ryanodine receptor are part of a multi-component signaling complex on the cardiomyocyte nuclear envelope.","date":"2001","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/11590243","citation_count":128,"is_preprint":false},{"pmid":"16306226","id":"PMC_16306226","title":"The mAKAP complex participates in the induction of cardiac myocyte hypertrophy by adrenergic receptor signaling.","date":"2005","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/16306226","citation_count":109,"is_preprint":false},{"pmid":"15652351","id":"PMC_15652351","title":"Nesprin-1alpha contributes to the targeting of mAKAP to the cardiac myocyte nuclear envelope.","date":"2005","source":"Experimental cell research","url":"https://pubmed.ncbi.nlm.nih.gov/15652351","citation_count":100,"is_preprint":false},{"pmid":"7721854","id":"PMC_7721854","title":"Cloning and characterization of A-kinase anchor protein 100 (AKAP100). A protein that targets A-kinase to the sarcoplasmic reticulum.","date":"1995","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/7721854","citation_count":94,"is_preprint":false},{"pmid":"9679148","id":"PMC_9679148","title":"A-kinase anchoring protein 100 (AKAP100) is localized in multiple subcellular compartments in the adult rat heart.","date":"1998","source":"The Journal of cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/9679148","citation_count":90,"is_preprint":false},{"pmid":"15182229","id":"PMC_15182229","title":"PKA-phosphorylation of PDE4D3 facilitates recruitment of the mAKAP signalling complex.","date":"2004","source":"The Biochemical journal","url":"https://pubmed.ncbi.nlm.nih.gov/15182229","citation_count":90,"is_preprint":false},{"pmid":"20106966","id":"PMC_20106966","title":"cAMP-stimulated protein phosphatase 2A activity associated with muscle A kinase-anchoring protein (mAKAP) signaling complexes inhibits the phosphorylation and activity of the cAMP-specific phosphodiesterase PDE4D3.","date":"2010","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/20106966","citation_count":81,"is_preprint":false},{"pmid":"12709444","id":"PMC_12709444","title":"Targeting of protein kinase A by muscle A kinase-anchoring protein (mAKAP) regulates phosphorylation and function of the skeletal muscle ryanodine receptor.","date":"2003","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/12709444","citation_count":57,"is_preprint":false},{"pmid":"16460834","id":"PMC_16460834","title":"The mAKAP signaling complex: integration of cAMP, calcium, and MAP kinase signaling pathways.","date":"2006","source":"European journal of cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/16460834","citation_count":56,"is_preprint":false},{"pmid":"19109240","id":"PMC_19109240","title":"mAKAP compartmentalizes oxygen-dependent control of HIF-1alpha.","date":"2008","source":"Science signaling","url":"https://pubmed.ncbi.nlm.nih.gov/19109240","citation_count":54,"is_preprint":false},{"pmid":"25551320","id":"PMC_25551320","title":"mAKAP-a master scaffold for cardiac remodeling.","date":"2015","source":"Journal of cardiovascular pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/25551320","citation_count":44,"is_preprint":false},{"pmid":"33295871","id":"PMC_33295871","title":"AKAP6 orchestrates the nuclear envelope microtubule-organizing center by linking golgi and nucleus via AKAP9.","date":"2020","source":"eLife","url":"https://pubmed.ncbi.nlm.nih.gov/33295871","citation_count":43,"is_preprint":false},{"pmid":"23261540","id":"PMC_23261540","title":"Regulation of MEF2 transcriptional activity by calcineurin/mAKAP complexes.","date":"2012","source":"Experimental cell research","url":"https://pubmed.ncbi.nlm.nih.gov/23261540","citation_count":35,"is_preprint":false},{"pmid":"17487687","id":"PMC_17487687","title":"The mAKAP signalosome and cardiac myocyte hypertrophy.","date":"2007","source":"IUBMB life","url":"https://pubmed.ncbi.nlm.nih.gov/17487687","citation_count":30,"is_preprint":false},{"pmid":"22484155","id":"PMC_22484155","title":"Myocyte enhancer factor 2 (MEF2) tethering to muscle selective A-kinase anchoring protein (mAKAP) is necessary for myogenic differentiation.","date":"2012","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/22484155","citation_count":23,"is_preprint":false},{"pmid":"26563778","id":"PMC_26563778","title":"AKAP6 inhibition impairs myoblast differentiation and muscle regeneration: Positive loop between AKAP6 and myogenin.","date":"2015","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/26563778","citation_count":20,"is_preprint":false},{"pmid":"34605406","id":"PMC_34605406","title":"Myogenin controls via AKAP6 non-centrosomal microtubule-organizing center formation at the nuclear envelope.","date":"2021","source":"eLife","url":"https://pubmed.ncbi.nlm.nih.gov/34605406","citation_count":17,"is_preprint":false},{"pmid":"23806656","id":"PMC_23806656","title":"Protein kinase A and phosphodiesterase-4D3 binding to coding polymorphisms of cardiac muscle anchoring protein (mAKAP).","date":"2013","source":"Journal of molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/23806656","citation_count":15,"is_preprint":false},{"pmid":"31325238","id":"PMC_31325238","title":"AKAP6 and phospholamban colocalize and interact in HEK-293T cells and primary murine cardiomyocytes.","date":"2019","source":"Physiological reports","url":"https://pubmed.ncbi.nlm.nih.gov/31325238","citation_count":8,"is_preprint":false},{"pmid":"40097291","id":"PMC_40097291","title":"Wnt/β-catenin pathway induces cardiac dysfunction via AKAP6-mediated RyR2 phosphorylation and sarcoplasmic reticulum calcium leakage.","date":"2025","source":"Journal of molecular cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/40097291","citation_count":3,"is_preprint":false},{"pmid":"39578909","id":"PMC_39578909","title":"AKAP6 controls NFATc4 activity for BDNF-mediated neuroprotection.","date":"2024","source":"Molecular brain","url":"https://pubmed.ncbi.nlm.nih.gov/39578909","citation_count":1,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":12474,"output_tokens":5455,"usd":0.059624,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":13890,"output_tokens":4049,"usd":0.085337,"stage2_stop_reason":"end_turn"},"total_usd":0.144961,"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\": 1995,\n      \"finding\": \"AKAP100 (AKAP6) was identified as a PKA type II regulatory subunit (RII)-binding protein that targets PKA type II to the sarcoplasmic reticulum in cardiac and skeletal muscle cells, demonstrated by RII overlay assay and co-purification with RII from cell extracts.\",\n      \"method\": \"Interaction cloning with RII probe, solid-phase overlay assay, immunohistochemistry, cAMP-agarose affinity co-purification\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (interaction cloning, overlay assay, affinity purification, immunohistochemistry) in foundational study, independently replicated in subsequent work\",\n      \"pmids\": [\"7721854\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"AKAP100 (AKAP6) localizes to multiple subcellular compartments in adult rat cardiomyocytes including the nucleus, sarcolemma, intercalated disc, Z-line, and transverse tubule/junctional SR; RII (but not RI) co-localizes with AKAP100 at these sites.\",\n      \"method\": \"Immunofluorescence, confocal microscopy, double immunostaining with alpha-actinin and ryanodine receptor antibodies\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — localization established by immunofluorescence/confocal across multiple structures, single lab but multiple markers\",\n      \"pmids\": [\"9679148\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"mAKAP (AKAP6) is targeted to the nuclear membrane of differentiated myocytes; nuclear membrane targeting is conferred by two regions (residues 772–915 and 915–1065) containing spectrin-like repeat sequences, as shown by GFP fusion constructs and displacement experiments.\",\n      \"method\": \"GFP fusion construct expression in myocytes, heterologous overexpression of targeting domains to displace endogenous mAKAP, immunolocalization\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — domain mapping with GFP fusions and competitive displacement, replicated in subsequent studies confirming nuclear envelope localization\",\n      \"pmids\": [\"10413680\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"mAKAP (AKAP6) assembles a cAMP signaling module containing PKA and PDE4D3 at the nuclear envelope of cardiac myocytes; tonic PDE4D3 activity reduces anchored PKA activity, while PKA activation stimulates mAKAP-associated PDE4D3, forming a negative feedback loop; disruption of PKA–mAKAP interaction prevents PKA-mediated enhancement of PDE4D3 activity.\",\n      \"method\": \"Co-immunoprecipitation from heart tissue, functional kinase and phosphodiesterase assays, PKA-mAKAP interaction disruption experiments\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — biochemical reconstitution of complex in native tissue, functional assays with disruption mutants, multiple orthogonal approaches\",\n      \"pmids\": [\"11296225\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"The mAKAP (AKAP6) complex at the cardiac nuclear envelope also includes ryanodine receptors and protein phosphatase 2A; a subset of cardiac ryanodine receptor binds to mAKAP at the nuclear envelope, potentially enabling PKA-mediated phosphorylation of ryanodine receptor.\",\n      \"method\": \"Co-immunoprecipitation, immunohistochemistry, tissue fractionation\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — co-IP and fractionation from cardiac tissue, single lab, two orthogonal methods\",\n      \"pmids\": [\"11590243\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Anchoring of PKA by mAKAP (AKAP6) at the sarcoplasmic reticulum co-localizes with RyR1 and increases PKA-dependent phosphorylation of RyR1 and Ca2+ efflux through RyR1; a PKA-binding-deficient mAKAP mutant (mAKAP-P) fails to enhance RyR1 phosphorylation or Ca2+ transient amplitude.\",\n      \"method\": \"Immunoelectron microscopy, Ca2+ transient measurements, phosphorylation assays in CHO cells stably expressing RyR1, expression of wild-type vs. PKA-binding-deficient mAKAP mutant\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — structure-function analysis with loss-of-function mutant, multiple readouts (phosphorylation + Ca2+ flux), replicated conceptually in cardiac studies\",\n      \"pmids\": [\"12709444\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"PKA phosphorylation of PDE4D3 on Ser-13 increases the affinity of PDE4D3 for mAKAP (AKAP6), facilitating recruitment of PDE4D3 to the mAKAP signaling complex for faster cAMP signal termination.\",\n      \"method\": \"In vitro phosphorylation assays, co-immunoprecipitation, site-directed mutagenesis of PDE4D3 Ser-13, cellular experiments\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — site-directed mutagenesis identifying specific phosphorylation site combined with in vitro and cellular binding assays\",\n      \"pmids\": [\"15182229\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"mAKAP (AKAP6) coordinates two integrated cAMP effector pathways: anchored PKA stimulates PDE4D3 to reduce local cAMP, while an mAKAP-associated ERK5 module suppresses PDE4D3; PDE4D3 also recruits Epac1 to enable cAMP-dependent attenuation of ERK5; anchored ERK5 can induce cardiomyocyte hypertrophy.\",\n      \"method\": \"Co-immunoprecipitation, pharmacological inhibitors, RNA interference, dominant-negative constructs, cardiomyocyte hypertrophy assays\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (co-IP, RNAi, pharmacology, dominant-negative), replicated conceptually across multiple labs\",\n      \"pmids\": [\"16177794\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Nesprin-1alpha serves as a receptor for mAKAP (AKAP6) on the nuclear envelope; the amino-terminal dimerization domain of nesprin-1alpha directly binds the third spectrin repeat of mAKAP to target it to the nuclear envelope; overexpression of these spectrin repeat domains displaces mAKAP from nesprin-1alpha.\",\n      \"method\": \"Co-immunoprecipitation, direct binding assays, overexpression of spectrin repeat domains, displacement experiments in myocytes\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct binding assays plus in-cell displacement experiments, domain-level mapping, mechanistically validated\",\n      \"pmids\": [\"15652351\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"The mAKAP (AKAP6) complex facilitates PKA-catalyzed phosphorylation of the ryanodine receptor Ca2+-release channel and is required for adrenergic-mediated cardiomyocyte hypertrophy; calcineurin Abeta associates with mAKAP and formation of the mAKAP complex is required for full activation of the pro-hypertrophic transcription factor NFATc.\",\n      \"method\": \"RNA interference of mAKAP, expression of PKA-binding-deficient mAKAP mutant, ryanodine receptor inhibition, co-immunoprecipitation of calcineurin, NFATc reporter assays\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — RNAi loss-of-function, mutant rescue, co-IP, and transcription factor reporter assays — multiple orthogonal approaches in one study\",\n      \"pmids\": [\"16306226\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"mAKAP (AKAP6) organizes ubiquitin E3 ligases that control the stability of HIF-1alpha near the nuclear envelope; depletion of mAKAP or disruption of its perinuclear targeting alters HIF-1alpha stability and transcriptional activation of hypoxia-responsive genes in cardiomyocytes.\",\n      \"method\": \"mAKAP depletion (RNAi), disruption of perinuclear targeting, HIF-1alpha stability assays, transcriptional reporter assays, co-immunoprecipitation of E3 ligases\",\n      \"journal\": \"Science signaling\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function and targeting disruption with defined transcriptional readout, single lab\",\n      \"pmids\": [\"19109240\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"PP2A associated with mAKAP (AKAP6) complexes (containing B56delta subunit) dephosphorylates PDE4D3 at Ser-54, reversing PKA-mediated activation of PDE4D3; a C-terminal mAKAP domain (residues 2085–2319) binds PP2A; PKA phosphorylation of B56delta enhances PP2A activity 2-fold in the complex, creating a cAMP-induced positive feedback loop.\",\n      \"method\": \"Domain mapping, co-immunoprecipitation, phosphatase activity assays, site-directed mutagenesis of B56delta PKA phosphorylation site, deletion of mAKAP C-terminal domain\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — domain mapping with deletion mutants, phosphorylation site mutagenesis, enzymatic activity assays, multiple orthogonal methods in one study\",\n      \"pmids\": [\"20106966\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"mAKAP (AKAP6) organizes a calcineurin/MEF2 signaling complex in myocytes; a calcineurin/mAKAP/MEF2 complex can be isolated from C2C12 cells and cardiac myocytes; calcineurin–MEF2 association is dependent on mAKAP expression; disruption of calcineurin–mAKAP binding blunts MEF2 transcriptional activity during myoblast differentiation and inhibits adrenergic-induced cardiac hypertrophy.\",\n      \"method\": \"Co-immunoprecipitation from C2C12 cells and cardiac myocytes, dominant-interference peptide disrupting calcineurin–mAKAP binding, MEF2 transcriptional reporter assays, siRNA knockdown\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal co-IP, dominant-interference peptide with defined binding domain, transcriptional readout, and hypertrophy phenotype — multiple orthogonal methods\",\n      \"pmids\": [\"23261540\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"mAKAP (AKAP6) directly binds MEF2 through discrete interaction domains; disruption of MEF2–mAKAP binding blocks MEF2 activation during early myoblast differentiation and inhibits myotube formation and expression of differentiation markers.\",\n      \"method\": \"Direct binding assays identifying discrete binding domains, dominant-interference expression, MEF2 transcriptional reporter assays, myotube formation assays\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding domain mapping and loss-of-function with defined phenotype, single lab\",\n      \"pmids\": [\"22484155\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Human mAKAP coding polymorphism P1400S (in the PDE4D3 binding site) reduces mAKAP–PDE4D3 binding without affecting PKA binding or activity; S2195F (near the PP2A binding site) increases PKA binding and PKA activity; L717V (flanking the spectrin repeat domain) increases PKA binding without changing PKA activity.\",\n      \"method\": \"Site-directed mutagenesis of mAKAP, co-immunoprecipitation, surface plasmon resonance (Biacore), PKA activity assays, Ca2+ measurements\",\n      \"journal\": \"Journal of molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — mutagenesis of specific residues with orthogonal binding methods (co-IP + SPR) and functional assays in single lab\",\n      \"pmids\": [\"23806656\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"AKAP6 knockdown in skeletal myoblasts halts myotube formation and decreases myogenin and myosin heavy chain expression; AKAP6 promotes myogenin expression through MEF2A; myogenin in turn binds an E-box site on the AKAP6 promoter to increase AKAP6 expression, forming a positive feedback loop.\",\n      \"method\": \"siRNA knockdown, shRNA lentiviral delivery in vivo (cardiotoxin muscle injury model), chromatin immunoprecipitation, luciferase reporter assays, motor function assessment\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vitro and in vivo loss-of-function, ChIP identifying direct promoter binding, luciferase reporter, multiple orthogonal methods\",\n      \"pmids\": [\"26563778\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"AKAP6 physically interacts with phospholamban (PLN) at the perinuclear SR region in HEK-293T cells and cardiomyocytes; AKAP6 promotes Ca2+ uptake activity of SERCA1 in cotransfected cells even in the presence of PLN.\",\n      \"method\": \"Immunofluorescence colocalization, co-immunoprecipitation from HEK-293T cells and adult rat cardiomyocytes, Ca2+ uptake assay\",\n      \"journal\": \"Physiological reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — co-IP from two cell systems plus functional Ca2+ uptake assay, single lab\",\n      \"pmids\": [\"31325238\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"AKAP6 is a key organizer of the nuclear envelope MTOC (NE-MTOC) in cardiomyocytes and osteoclasts; AKAP6 anchors centrosomal proteins (Pcnt, AKAP9) to the nuclear envelope via its spectrin repeats, acting as an adaptor between nesprin-1alpha and Pcnt/AKAP9; AKAP6 and AKAP9 form a protein platform tethering the Golgi to the nucleus; ectopic AKAP6 expression in epithelial cells is sufficient to recruit endogenous centrosomal proteins; AKAP6 is required for cardiomyocyte hypertrophy and osteoclast bone resorption.\",\n      \"method\": \"Loss-of-function (siRNA/shRNA), gain-of-function (ectopic expression), co-immunoprecipitation, immunofluorescence in cardiomyocytes and osteoclasts, microtubule nucleation assays\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (co-IP, gain/loss-of-function, MT nucleation), multiple cell types, functional phenotype (hypertrophy, bone resorption)\",\n      \"pmids\": [\"33295871\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Myogenin transcription factor is required and sufficient for NE-MTOC formation and induces AKAP6 expression; overexpression of AKAP6beta and nesprin-1alpha together is sufficient to recruit endogenous MTOC proteins to the nuclear envelope of myoblasts in the absence of myogenin, identifying AKAP6 as a central effector of myogenin-controlled NE-MTOC assembly.\",\n      \"method\": \"Loss-of-function (myogenin KO), gain-of-function (overexpression in fibroblasts and myoblasts), promoter reporter assays, bioinformatics, immunofluorescence\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic epistasis (myogenin KO), rescue by AKAP6 overexpression, promoter studies, multiple cell types and methods\",\n      \"pmids\": [\"34605406\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"AKAP6 anchors calcineurin (CaN) and NFATc4 in neurons, and this scaffolding is required for BDNF-mediated NFATc4 transcriptional activity and neuroprotection; disruption of calcineurin anchoring to AKAP6 diminishes BDNF's pro-survival effect; NFATc4 knockout mice show reduced BDNF neuroprotection in vivo.\",\n      \"method\": \"AKAP6 disruption experiments (calcineurin anchoring domain peptide), NFAT transcriptional reporter assays, NFATc4 knockout mice, neuronal survival assays\",\n      \"journal\": \"Molecular brain\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function (dominant interference + KO mouse), transcriptional reporter, in vivo validation — single lab\",\n      \"pmids\": [\"39578909\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Wnt/β-catenin pathway transcriptionally upregulates AKAP6 expression in cardiomyocytes; increased AKAP6 enhances PKA-mediated RyR2 phosphorylation, causing sarcoplasmic reticulum Ca2+ leakage and cardiomyocyte dysfunction.\",\n      \"method\": \"Wnt pathway activation, transcriptome analysis, AKAP6 overexpression/knockdown, RyR2 phosphorylation assays, Ca2+ measurements\",\n      \"journal\": \"Journal of molecular cell biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — transcriptomics identification combined with functional validation of AKAP6-RyR2 axis and Ca2+ readout, single lab\",\n      \"pmids\": [\"40097291\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"AKAP6 (mAKAP) is a scaffold protein anchored to the nuclear envelope of striated myocytes via its spectrin repeats binding nesprin-1alpha, where it organizes a large multi-protein signalosome integrating cAMP (PKA, PDE4D3, Epac1), Ca2+ (ryanodine receptor, calcineurin), MAP kinase (ERK5), PP2A, HIF-1alpha ubiquitin E3 ligases, and transcription factors (NFATc, MEF2) to coordinate cardiomyocyte hypertrophy, myoblast differentiation, and hypoxic gene responses; additionally, AKAP6 serves as the key organizer of the nuclear envelope microtubule-organizing center by bridging nesprin-1alpha to centrosomal proteins (Pcnt, AKAP9) and tethering the Golgi to the nucleus.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"AKAP6 (mAKAP) is a large scaffold protein that organizes a multi-protein signalosome at the nuclear envelope of striated myocytes to integrate cAMP, Ca2+, and MAP-kinase signals controlling cardiomyocyte hypertrophy and myocyte differentiation [#3, #7, #9]. It was first identified as a PKA type II regulatory subunit-binding protein targeting PKA to the sarcoplasmic reticulum of cardiac and skeletal muscle [#0], and is anchored to the nuclear membrane through spectrin-repeat regions that bind the amino-terminal dimerization domain of nesprin-1alpha [#2, #8]. At this site AKAP6 nucleates a cAMP module containing PKA and PDE4D3, where PKA stimulates PDE4D3 and PKA-phosphorylation of PDE4D3 (Ser-13) raises its affinity for the scaffold, establishing a negative feedback loop for cAMP termination [#3, #6]; an associated PP2A/B56delta phosphatase reverses PDE4D3 activation as a counterbalancing positive-feedback arm [#11], and an ERK5/Epac1 module couples this network to hypertrophic growth [#7]. The complex localizes ryanodine receptor and calcineurin to enable PKA-mediated RyR phosphorylation, Ca2+ release, and calcineurin-dependent activation of the pro-hypertrophic transcription factor NFATc [#5, #9]. AKAP6 directly binds MEF2 and organizes calcineurin/MEF2 signaling to drive myoblast differentiation, with myogenin feeding back to upregulate AKAP6 transcription [#12, #13, #15]. It also organizes ubiquitin E3 ligases that govern HIF-1alpha stability and hypoxic gene responses [#10]. Independently of its kinase-anchoring role, AKAP6 is the central organizer of the nuclear-envelope microtubule-organizing center, bridging nesprin-1alpha to the centrosomal proteins pericentrin and AKAP9 and tethering the Golgi to the nucleus, a function induced downstream of myogenin [#17, #18]. Beyond striated muscle, AKAP6 anchors calcineurin and NFATc4 in neurons to mediate BDNF-dependent transcription and neuroprotection [#19].\",\n  \"teleology\": [\n    {\n      \"year\": 1995,\n      \"claim\": \"Established AKAP6 as a PKA-anchoring protein, defining its founding molecular activity of targeting PKA type II to muscle sarcoplasmic reticulum.\",\n      \"evidence\": \"Interaction cloning with RII probe, overlay assay, and cAMP-agarose co-purification from muscle extracts\",\n      \"pmids\": [\"7721854\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define the nuclear-envelope targeting mechanism\", \"No downstream substrates identified at this stage\"]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"Mapped the spectrin-repeat regions responsible for nuclear-membrane targeting, locating the scaffold to a defined subcellular compartment in differentiated myocytes.\",\n      \"evidence\": \"GFP fusion constructs and competitive displacement of endogenous mAKAP in myocytes\",\n      \"pmids\": [\"10413680\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"The nuclear-envelope receptor binding these repeats was not yet identified\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Identified nesprin-1alpha as the nuclear-envelope receptor, explaining how AKAP6 is physically anchored to the perinuclear membrane.\",\n      \"evidence\": \"Direct binding assays and in-cell displacement mapping nesprin-1alpha N-terminus to the third spectrin repeat of mAKAP\",\n      \"pmids\": [\"15652351\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of the spectrin-repeat/nesprin interface not resolved\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Defined the core cAMP-handling logic of the signalosome: a PKA/PDE4D3 module with reciprocal feedback regulating local cAMP.\",\n      \"evidence\": \"Co-IP from heart tissue, kinase/PDE activity assays, PKA-binding disruption, and PDE4D3 Ser-13 mutagenesis\",\n      \"pmids\": [\"11296225\", \"15182229\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How the cAMP module integrates with Ca2+ and MAPK arms not yet defined\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Connected the scaffold to Ca2+ handling and hypertrophy, showing PKA anchoring enhances RyR phosphorylation and Ca2+ flux and that mAKAP is required for adrenergic hypertrophy via calcineurin/NFATc.\",\n      \"evidence\": \"PKA-binding-deficient mutant, Ca2+ transient and phosphorylation assays, RNAi, calcineurin co-IP, and NFATc reporters in cardiomyocytes\",\n      \"pmids\": [\"12709444\", \"16306226\", \"11590243\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Stoichiometry of RyR recruitment to nuclear-envelope mAKAP unclear\", \"RyR co-IP/fractionation results from a single lab\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Revealed integration of MAPK signaling, with an ERK5 module and Epac1 coupling cAMP to hypertrophic growth control through the scaffold.\",\n      \"evidence\": \"Co-IP, RNAi, pharmacology, dominant-negatives, and cardiomyocyte hypertrophy assays\",\n      \"pmids\": [\"16177794\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Quantitative contribution of ERK5 versus calcineurin arms to hypertrophy not separated\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Added a PP2A/B56delta counter-regulatory arm and extended the scaffold to hypoxic signaling via HIF-1alpha-controlling E3 ligases.\",\n      \"evidence\": \"Domain mapping, phosphatase activity assays, B56delta mutagenesis, and HIF-1alpha stability/reporter assays with mAKAP depletion\",\n      \"pmids\": [\"20106966\", \"19109240\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Identity of the HIF-1alpha E3 ligases not fully defined\", \"HIF-1alpha findings from a single lab\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Defined AKAP6's role in myocyte differentiation through direct MEF2/calcineurin binding and a myogenin feedback loop driving its own expression.\",\n      \"evidence\": \"MEF2 binding-domain mapping, dominant-interference peptides, ChIP of myogenin at the AKAP6 promoter, reporter and myotube assays, in vivo shRNA\",\n      \"pmids\": [\"23261540\", \"22484155\", \"26563778\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Discrete MEF2-binding domain structure not resolved\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Established a kinase-anchoring-independent function: AKAP6 as the central organizer of the nuclear-envelope MTOC, bridging nesprin-1alpha to centrosomal proteins and Golgi, downstream of myogenin.\",\n      \"evidence\": \"Loss/gain-of-function, co-IP, microtubule nucleation, and myogenin KO/rescue in cardiomyocytes, osteoclasts, and myoblasts\",\n      \"pmids\": [\"33295871\", \"34605406\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How NE-MTOC scaffolding and the signalosome roles are partitioned on the same protein is unresolved\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Extended AKAP6 scaffolding beyond muscle, showing it anchors calcineurin/NFATc4 in neurons for BDNF-dependent transcription and neuroprotection.\",\n      \"evidence\": \"Calcineurin-anchoring disruption peptide, NFAT reporters, NFATc4 KO mice, and neuronal survival assays\",\n      \"pmids\": [\"39578909\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Neuronal localization of AKAP6 not mapped\", \"Findings from a single lab\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Placed AKAP6 downstream of Wnt/beta-catenin, linking its transcriptional upregulation to pathological RyR2 hyperphosphorylation and Ca2+ leak.\",\n      \"evidence\": \"Wnt activation, transcriptomics, AKAP6 over/knockdown, and RyR2 phosphorylation/Ca2+ assays in cardiomyocytes\",\n      \"pmids\": [\"40097291\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct Wnt-responsive element on AKAP6 promoter not confirmed\", \"Single lab\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the same scaffold molecule simultaneously coordinates the cAMP/Ca2+ signalosome and the NE-MTOC, and whether disease-associated coding variants alter these functions in vivo, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structural model of the assembled signalosome\", \"In vivo phenotype of AKAP6 polymorphisms not established\", \"Mechanism dividing scaffold pools between functions unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 3, 8, 13, 17]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [3, 11, 9]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [2, 8, 17]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005635\", \"supporting_discovery_ids\": [2, 8, 10]},\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [0, 5, 16]},\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [17]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [1, 2]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [3, 7, 9]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [13, 15, 18]},\n      {\"term_id\": \"R-HSA-1852241\", \"supporting_discovery_ids\": [17, 18]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [9, 10, 12]}\n    ],\n    \"complexes\": [\"mAKAP signalosome (nuclear envelope)\", \"nuclear-envelope MTOC (NE-MTOC)\"],\n    \"partners\": [\"PRKAR2/PKA\", \"PDE4D3\", \"RYR2\", \"PPP3CB (calcineurin)\", \"PPP2 (PP2A B56delta)\", \"SYNE1 (nesprin-1alpha)\", \"PCNT\", \"AKAP9\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}