{"gene":"REEP5","run_date":"2026-06-10T06:43:36","timeline":{"discoveries":[{"year":2020,"finding":"REEP5 is a cardiac-enriched SR/ER membrane protein required for SR/ER structural organization; in vitro depletion in mouse cardiac myocytes causes SR/ER membrane destabilization and luminal vacuolization, decreased myocyte contractility, and disrupted Ca2+ cycling. In vivo CRISPR/Cas9-mediated loss-of-function in zebrafish causes sensitized cardiac dysfunction, and AAV9-induced depletion in mice causes cardiac dysfunction with dilated chambers, increased fibrosis, and reduced ejection fraction.","method":"shRNA/siRNA depletion in mouse cardiomyocytes, CRISPR/Cas9 zebrafish mutants, AAV9-mediated knockdown in mice, Ca2+ imaging, contractility measurements","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal in vitro and in vivo loss-of-function models (cell culture, zebrafish, mouse) with defined cellular and physiological phenotypes, replicated across systems","pmids":["32075961"],"is_preprint":false},{"year":2018,"finding":"REEP5 acts as a sarcoplasmic reticulum membrane sculptor in cardiomyocytes. Targeted inactivation of REEP5 in rats specifically deformed cardiac SR membrane architecture (visualized by FIB-SEM 3D reconstruction) without affecting transverse tubules, resulting in normal L-type Ca2+ channel currents but depressed SR Ca2+ release, reduced excitation-contraction coupling gain, and compromised cardiac contractility. REEP5 deficiency did not alter expression of major Ca2+-handling proteins.","method":"REEP5 knockout rats, FIB-SEM 3D reconstruction of SR, simultaneous patch-clamp recording of Ca2+ currents and Ca2+ transients","journal":"Journal of the American Heart Association","confidence":"High","confidence_rationale":"Tier 1-2 / Moderate — in vivo KO with ultrastructural 3D imaging and functional electrophysiology, single lab with multiple orthogonal methods","pmids":["29431104"],"is_preprint":false},{"year":2024,"finding":"REEP5 interacts with Mitofusins 1 and 2 (MFN1/2) to mediate mitochondrial 'hitchhiking' on tubular ER along microtubules, enabling cytosolic distribution of mitochondria. REEP5 depletion reduced ER-mitochondria tethering and caused perinuclear clustering of mitochondria. Rapamycin-induced irreversible REEP5-MFN1/2 interaction caused mitochondrial hyperfusion. Disruption of MFN2-REEP5 interaction dynamics or REEP5 silencing modulated mitochondrial reactive oxygen species (ROS) production.","method":"Co-immunoprecipitation, rapamycin-induced forced dimerization, live-cell imaging, ROS measurements, siRNA knockdown, REEP5 overexpression","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal interaction validation, multiple orthogonal functional assays (forced dimerization, KD, OE), defined cellular phenotypes in single lab","pmids":["39133213"],"is_preprint":false},{"year":2016,"finding":"REEP5 physically interacts with CXCR1 (but not CXCR2) and functions as an accessory protein that promotes ligand-stimulated endocytosis of CXCR1. In the absence of REEP5, CXCR1 is present at the plasma membrane but receptor internalization and intracellular clustering of β-arrestin2 following IL-8 treatment are impaired. REEP5 depletion reduces IL-8-stimulated ERK phosphorylation, actin polymerization, and cell invasion.","method":"Co-immunoprecipitation, siRNA depletion, receptor internalization assay, β-arrestin2 clustering imaging, ERK phosphorylation assay, invasion assay, xenograft mouse model","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — Co-IP plus functional depletion assays with multiple readouts in single lab; interaction specificity shown by CXCR1 vs CXCR2 comparison","pmids":["27966653"],"is_preprint":false},{"year":2023,"finding":"REEP5 forms a complex with TRAM1 that interacts with SARS-CoV-2 NSP3 at replication organelles (ROs) and promotes viral replication.","method":"Host-viral protein-protein interactome mapping (co-expression of NSP3/NSP4/NSP6 individually and in combination), co-immunoprecipitation/MS","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2-3 / Weak — interaction identified by PPI mapping with functional viral replication readout, single lab, limited mechanistic detail in abstract","pmids":["37768083"],"is_preprint":false},{"year":2024,"finding":"REEP5 overexpression inhibits ER stress in cardiomyocytes following myocardial infarction, suppressing phosphorylation of PERK and IRE1α and nuclear translocation of ATF6, and reducing CHOP and cleaved caspase-12 levels to alleviate ER stress-induced apoptosis. REEP5 was found to physically bind CLEC5A, and REEP5 overexpression abolished CLEC5A-induced ER stress and apoptosis.","method":"Co-immunoprecipitation (REEP5-CLEC5A interaction), gain-of-function overexpression in MI mouse model and hypoxia cell model, western blot for ER stress markers, apoptosis assays","journal":"BMC cardiovascular disorders","confidence":"Medium","confidence_rationale":"Tier 2-3 / Weak — Co-IP for interaction, in vivo and in vitro gain-of-function with ER stress pathway readouts, single lab","pmids":["39044150"],"is_preprint":false},{"year":2026,"finding":"AAV9-shRNA knockdown of REEP5 in mouse hearts causes fragmented mitochondrial networks and increased reactive oxygen species in isolated cardiomyocytes. Loss of REEP5 alters the SR/ER membrane-shaping proteome (upregulating RTN4, ATL3, CKAP4 as partial compensation) and broadly reorganizes mitochondrial and microsomal proteomes, with depletion of mitochondrial import machinery and antioxidant enzymes, suggesting REEP5 supports SR/ER-mitochondria tethering and functional crosstalk.","method":"AAV9-shRNA knockdown in mouse heart, subcellular fractionation, DIA mass spectrometry proteomics, live-cell imaging of mitochondrial networks, ROS measurement","journal":"Molecular & cellular proteomics : MCP","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo KD combined with subcellular proteomics and live imaging, multiple orthogonal readouts in single lab","pmids":["41672147"],"is_preprint":false}],"current_model":"REEP5 is a cardiac-enriched ER/SR membrane-shaping protein that maintains SR/ER tubular architecture to support Ca2+ cycling and excitation-contraction coupling in cardiomyocytes; it also interacts with Mitofusins 1/2 to mediate mitochondrial 'hitchhiking' on tubular ER for cytosolic mitochondrial distribution, forms a complex with TRAM1 that engages SARS-CoV-2 NSP3 at replication organelles, and acts as an accessory protein promoting ligand-stimulated endocytosis of CXCR1."},"narrative":{"mechanistic_narrative":"REEP5 is an endoplasmic/sarcoplasmic reticulum membrane-shaping protein that organizes tubular SR/ER architecture to support cardiomyocyte function, including Ca2+ cycling and excitation-contraction coupling [PMID:32075961, PMID:29431104]. Loss of REEP5 destabilizes SR/ER membranes and produces luminal vacuolization, reduced contractility, and disrupted Ca2+ handling in cardiomyocytes, and causes dilated cardiomyopathy-like dysfunction in vivo across zebrafish and mouse models [PMID:32075961]; ultrastructurally, REEP5 deficiency specifically deforms SR membrane geometry while sparing transverse tubules and Ca2+-handling protein levels, yielding depressed SR Ca2+ release and reduced EC-coupling gain [PMID:29431104]. Through this membrane-shaping role REEP5 also coordinates inter-organelle contacts: it binds Mitofusins 1 and 2 to enable mitochondrial 'hitchhiking' along tubular ER on microtubules, controlling cytosolic mitochondrial distribution and ER-mitochondria tethering, with REEP5 loss causing perinuclear mitochondrial clustering and altered mitochondrial ROS [PMID:39133213], and in cardiac tissue its loss fragments mitochondrial networks, raises ROS, and remodels both the SR/ER membrane-shaping proteome and mitochondrial/microsomal proteomes [PMID:41672147]. Beyond its structural role, REEP5 overexpression suppresses PERK/IRE1α/ATF6 ER-stress signaling and apoptosis after myocardial infarction and binds CLEC5A to counter CLEC5A-induced ER stress [PMID:39044150]. REEP5 additionally acts as an accessory protein promoting ligand-stimulated endocytosis of CXCR1 [PMID:27966653], and forms a complex with TRAM1 that engages SARS-CoV-2 NSP3 at replication organelles to promote viral replication [PMID:37768083].","teleology":[{"year":2016,"claim":"Established a non-structural signaling role for REEP5 by asking whether it modulates GPCR trafficking, showing it is required for ligand-stimulated CXCR1 internalization and downstream signaling.","evidence":"Co-IP, siRNA depletion, receptor internalization and β-arrestin2 clustering imaging, ERK/invasion assays, xenograft model","pmids":["27966653"],"confidence":"Medium","gaps":["Mechanism by which REEP5 promotes CXCR1 endocytosis (vesicle scission vs membrane shaping) not defined","Receptor selectivity beyond CXCR1 vs CXCR2 untested","Relationship to its ER membrane-shaping role unclear"]},{"year":2018,"claim":"Resolved whether REEP5 shapes cardiac SR membranes and whether this geometry is functionally coupled to Ca2+ release, distinguishing structural deformation from altered Ca2+-handling protein expression.","evidence":"REEP5 knockout rats with FIB-SEM 3D SR reconstruction and simultaneous patch-clamp of Ca2+ currents and transients","pmids":["29431104"],"confidence":"High","gaps":["How REEP5 physically curves/stabilizes SR membrane at molecular level not shown","Whether the same mechanism operates in non-cardiac ER untested here"]},{"year":2020,"claim":"Confirmed REEP5 as essential for SR/ER structural integrity and cardiac function across orthogonal in vitro and in vivo systems, linking membrane destabilization to contractile and Ca2+-cycling failure.","evidence":"shRNA/siRNA in mouse cardiomyocytes, CRISPR/Cas9 zebrafish, AAV9 knockdown in mice, Ca2+ imaging and contractility measurements","pmids":["32075961"],"confidence":"High","gaps":["Whether observed dilated cardiomyopathy phenotype reflects a primary membrane defect or secondary remodeling not fully separated","Human disease relevance not established"]},{"year":2023,"claim":"Extended REEP5 function to host-pathogen biology by asking whether it participates in viral replication organelle formation, identifying a REEP5-TRAM1 complex that engages SARS-CoV-2 NSP3.","evidence":"Host-viral interactome mapping with co-expression of NSP3/NSP4/NSP6 and co-IP/MS","pmids":["37768083"],"confidence":"Medium","gaps":["Direct vs indirect REEP5-NSP3 interaction not resolved","Whether membrane-shaping activity is required for RO formation untested","Limited mechanistic detail on replication promotion"]},{"year":2024,"claim":"Defined a mechanism for inter-organelle organization by asking how tubular ER positions mitochondria, showing REEP5 binds MFN1/2 to mediate mitochondrial hitchhiking and tethering.","evidence":"Co-IP, rapamycin-induced forced dimerization, live-cell imaging, ROS measurement, siRNA and overexpression","pmids":["39133213"],"confidence":"High","gaps":["Stoichiometry and structural basis of the REEP5-MFN interface unknown","How tethering dynamics regulate ROS mechanistically not defined"]},{"year":2024,"claim":"Addressed whether REEP5 protects cardiomyocytes from ER-stress injury, showing its overexpression suppresses PERK/IRE1α/ATF6 signaling and CLEC5A-driven apoptosis after myocardial infarction.","evidence":"Co-IP (REEP5-CLEC5A), gain-of-function overexpression in MI mouse and hypoxia cell models, western blot for ER-stress markers, apoptosis assays","pmids":["39044150"],"confidence":"Medium","gaps":["Whether ER-stress suppression is a direct consequence of CLEC5A binding or of membrane stabilization unclear","Loss-of-function effect on ER stress not tested","Single-lab gain-of-function design"]},{"year":2026,"claim":"Connected REEP5 SR/ER shaping to mitochondrial health at the proteome level, showing cardiac REEP5 loss fragments mitochondria, raises ROS, and remodels membrane-shaping and mitochondrial proteomes.","evidence":"AAV9-shRNA knockdown in mouse heart, subcellular fractionation with DIA mass spectrometry, live imaging of mitochondrial networks, ROS measurement","pmids":["41672147"],"confidence":"Medium","gaps":["Causal chain from SR/ER membrane defect to mitochondrial import/antioxidant proteome changes not established","Whether RTN4/ATL3/CKAP4 upregulation is true functional compensation untested"]},{"year":null,"claim":"How REEP5's intrinsic membrane-curving biochemistry generates SR/ER tubule geometry, and how this single activity is shared across its cardiac, inter-organelle, GPCR-trafficking, and viral roles, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structural model of REEP5 membrane-shaping mechanism in the corpus","No unifying biochemical assay linking the disparate functional contexts","No human Mendelian disease link established in the corpus"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[0,1]}],"localization":[{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[0,2,5]}],"pathway":[{"term_id":"R-HSA-397014","term_label":"Muscle contraction","supporting_discovery_ids":[0,1]},{"term_id":"R-HSA-1852241","term_label":"Organelle biogenesis and maintenance","supporting_discovery_ids":[2,6]}],"complexes":[],"partners":["MFN1","MFN2","CXCR1","TRAM1","CLEC5A"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q00765","full_name":"Receptor expression-enhancing protein 5","aliases":["Polyposis locus protein 1","Protein TB2"],"length_aa":189,"mass_kda":21.5,"function":"Plays an essential role in heart function and development by regulating the organization and function of the sarcoplasmic reticulum in cardiomyocytes","subcellular_location":"Endoplasmic reticulum membrane; Sarcoplasmic reticulum membrane","url":"https://www.uniprot.org/uniprotkb/Q00765/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/REEP5","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000129625","cell_line_id":"CID001524","localizations":[{"compartment":"er","grade":3}],"interactors":[{"gene":"ARL6IP1","stoichiometry":10.0},{"gene":"RTN4","stoichiometry":10.0},{"gene":"ATL3","stoichiometry":4.0},{"gene":"COPA","stoichiometry":4.0},{"gene":"ESYT1","stoichiometry":4.0},{"gene":"RTN3","stoichiometry":4.0},{"gene":"CALM2","stoichiometry":0.2},{"gene":"COPB2","stoichiometry":0.2},{"gene":"COPE","stoichiometry":0.2},{"gene":"RABAC1","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID001524","total_profiled":1310},"omim":[{"mim_id":"611731","title":"APC REGULATOR OF WNT SIGNALING PATHWAY; APC","url":"https://www.omim.org/entry/611731"},{"mim_id":"610243","title":"ZINC FINGER FYVE DOMAIN-CONTAINING PROTEIN 27; ZFYVE27","url":"https://www.omim.org/entry/610243"},{"mim_id":"604475","title":"RETICULON 4; RTN4","url":"https://www.omim.org/entry/604475"},{"mim_id":"125265","title":"RECEPTOR EXPRESSION-ENHANCING PROTEIN 5; REEP5","url":"https://www.omim.org/entry/125265"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Endoplasmic reticulum","reliability":"Supported"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/REEP5"},"hgnc":{"alias_symbol":["DP1","TB2","D5S346","Yip2e","POB16"],"prev_symbol":["C5orf18"]},"alphafold":{"accession":"Q00765","domains":[{"cath_id":"-","chopping":"1-51","consensus_level":"medium","plddt":72.8714,"start":1,"end":51},{"cath_id":"-","chopping":"52-187","consensus_level":"medium","plddt":82.3402,"start":52,"end":187}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q00765","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q00765-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q00765-F1-predicted_aligned_error_v6.png","plddt_mean":78.88},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=REEP5","jax_strain_url":"https://www.jax.org/strain/search?query=REEP5"},"sequence":{"accession":"Q00765","fasta_url":"https://rest.uniprot.org/uniprotkb/Q00765.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q00765/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q00765"}},"corpus_meta":[{"pmid":"27966653","id":"PMC_27966653","title":"The accessory proteins REEP5 and REEP6 refine CXCR1-mediated cellular responses and lung cancer progression.","date":"2016","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/27966653","citation_count":27,"is_preprint":false},{"pmid":"32075961","id":"PMC_32075961","title":"REEP5 depletion causes sarco-endoplasmic reticulum vacuolization and cardiac functional defects.","date":"2020","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/32075961","citation_count":24,"is_preprint":false},{"pmid":"23411922","id":"PMC_23411922","title":"Microsatellite instability--MSI markers (BAT26, BAT25, D2S123, D5S346, D17S250) in rectal cancer.","date":"2012","source":"Arquivos brasileiros de cirurgia digestiva : ABCD = Brazilian archives of digestive surgery","url":"https://pubmed.ncbi.nlm.nih.gov/23411922","citation_count":24,"is_preprint":false},{"pmid":"29431104","id":"PMC_29431104","title":"REEP5 (Receptor Accessory Protein 5) Acts as a Sarcoplasmic Reticulum Membrane Sculptor to Modulate Cardiac Function.","date":"2018","source":"Journal of the American Heart Association","url":"https://pubmed.ncbi.nlm.nih.gov/29431104","citation_count":19,"is_preprint":false},{"pmid":"22795047","id":"PMC_22795047","title":"Association of APC and REEP5 gene polymorphisms with major depression disorder and treatment response to antidepressants in a Han Chinese population.","date":"2012","source":"General hospital psychiatry","url":"https://pubmed.ncbi.nlm.nih.gov/22795047","citation_count":17,"is_preprint":false},{"pmid":"37221870","id":"PMC_37221870","title":"Comparison of interferon-gamma production between TB1 and TB2 tubes of QuantiFERON-TB Gold Plus: a meta-analysis.","date":"2023","source":"Clinical chemistry and laboratory medicine","url":"https://pubmed.ncbi.nlm.nih.gov/37221870","citation_count":11,"is_preprint":false},{"pmid":"39133213","id":"PMC_39133213","title":"Dynamic interaction of REEP5-MFN1/2 enables mitochondrial hitchhiking on tubular ER.","date":"2024","source":"The Journal of cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/39133213","citation_count":11,"is_preprint":false},{"pmid":"37768083","id":"PMC_37768083","title":"The REEP5/TRAM1 complex binds SARS-CoV-2 NSP3 and promotes virus replication.","date":"2023","source":"Journal of virology","url":"https://pubmed.ncbi.nlm.nih.gov/37768083","citation_count":11,"is_preprint":false},{"pmid":"30514508","id":"PMC_30514508","title":"Characterization of specific CD4 and CD8 T-cell responses in QuantiFERON TB Gold-Plus TB1 and TB2 tubes.","date":"2018","source":"Tuberculosis (Edinburgh, Scotland)","url":"https://pubmed.ncbi.nlm.nih.gov/30514508","citation_count":10,"is_preprint":false},{"pmid":"7654261","id":"PMC_7654261","title":"Correspondence of RLGS-M spot behavior with tissue expression on mouse homologue of DP1/TB2 gene.","date":"1995","source":"Biochemical and biophysical research 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In vivo CRISPR/Cas9-mediated loss-of-function in zebrafish causes sensitized cardiac dysfunction, and AAV9-induced depletion in mice causes cardiac dysfunction with dilated chambers, increased fibrosis, and reduced ejection fraction.\",\n      \"method\": \"shRNA/siRNA depletion in mouse cardiomyocytes, CRISPR/Cas9 zebrafish mutants, AAV9-mediated knockdown in mice, Ca2+ imaging, contractility measurements\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal in vitro and in vivo loss-of-function models (cell culture, zebrafish, mouse) with defined cellular and physiological phenotypes, replicated across systems\",\n      \"pmids\": [\"32075961\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"REEP5 acts as a sarcoplasmic reticulum membrane sculptor in cardiomyocytes. Targeted inactivation of REEP5 in rats specifically deformed cardiac SR membrane architecture (visualized by FIB-SEM 3D reconstruction) without affecting transverse tubules, resulting in normal L-type Ca2+ channel currents but depressed SR Ca2+ release, reduced excitation-contraction coupling gain, and compromised cardiac contractility. REEP5 deficiency did not alter expression of major Ca2+-handling proteins.\",\n      \"method\": \"REEP5 knockout rats, FIB-SEM 3D reconstruction of SR, simultaneous patch-clamp recording of Ca2+ currents and Ca2+ transients\",\n      \"journal\": \"Journal of the American Heart Association\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Moderate — in vivo KO with ultrastructural 3D imaging and functional electrophysiology, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"29431104\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"REEP5 interacts with Mitofusins 1 and 2 (MFN1/2) to mediate mitochondrial 'hitchhiking' on tubular ER along microtubules, enabling cytosolic distribution of mitochondria. REEP5 depletion reduced ER-mitochondria tethering and caused perinuclear clustering of mitochondria. Rapamycin-induced irreversible REEP5-MFN1/2 interaction caused mitochondrial hyperfusion. Disruption of MFN2-REEP5 interaction dynamics or REEP5 silencing modulated mitochondrial reactive oxygen species (ROS) production.\",\n      \"method\": \"Co-immunoprecipitation, rapamycin-induced forced dimerization, live-cell imaging, ROS measurements, siRNA knockdown, REEP5 overexpression\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal interaction validation, multiple orthogonal functional assays (forced dimerization, KD, OE), defined cellular phenotypes in single lab\",\n      \"pmids\": [\"39133213\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"REEP5 physically interacts with CXCR1 (but not CXCR2) and functions as an accessory protein that promotes ligand-stimulated endocytosis of CXCR1. In the absence of REEP5, CXCR1 is present at the plasma membrane but receptor internalization and intracellular clustering of β-arrestin2 following IL-8 treatment are impaired. REEP5 depletion reduces IL-8-stimulated ERK phosphorylation, actin polymerization, and cell invasion.\",\n      \"method\": \"Co-immunoprecipitation, siRNA depletion, receptor internalization assay, β-arrestin2 clustering imaging, ERK phosphorylation assay, invasion assay, xenograft mouse model\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — Co-IP plus functional depletion assays with multiple readouts in single lab; interaction specificity shown by CXCR1 vs CXCR2 comparison\",\n      \"pmids\": [\"27966653\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"REEP5 forms a complex with TRAM1 that interacts with SARS-CoV-2 NSP3 at replication organelles (ROs) and promotes viral replication.\",\n      \"method\": \"Host-viral protein-protein interactome mapping (co-expression of NSP3/NSP4/NSP6 individually and in combination), co-immunoprecipitation/MS\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Weak — interaction identified by PPI mapping with functional viral replication readout, single lab, limited mechanistic detail in abstract\",\n      \"pmids\": [\"37768083\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"REEP5 overexpression inhibits ER stress in cardiomyocytes following myocardial infarction, suppressing phosphorylation of PERK and IRE1α and nuclear translocation of ATF6, and reducing CHOP and cleaved caspase-12 levels to alleviate ER stress-induced apoptosis. REEP5 was found to physically bind CLEC5A, and REEP5 overexpression abolished CLEC5A-induced ER stress and apoptosis.\",\n      \"method\": \"Co-immunoprecipitation (REEP5-CLEC5A interaction), gain-of-function overexpression in MI mouse model and hypoxia cell model, western blot for ER stress markers, apoptosis assays\",\n      \"journal\": \"BMC cardiovascular disorders\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Weak — Co-IP for interaction, in vivo and in vitro gain-of-function with ER stress pathway readouts, single lab\",\n      \"pmids\": [\"39044150\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"AAV9-shRNA knockdown of REEP5 in mouse hearts causes fragmented mitochondrial networks and increased reactive oxygen species in isolated cardiomyocytes. Loss of REEP5 alters the SR/ER membrane-shaping proteome (upregulating RTN4, ATL3, CKAP4 as partial compensation) and broadly reorganizes mitochondrial and microsomal proteomes, with depletion of mitochondrial import machinery and antioxidant enzymes, suggesting REEP5 supports SR/ER-mitochondria tethering and functional crosstalk.\",\n      \"method\": \"AAV9-shRNA knockdown in mouse heart, subcellular fractionation, DIA mass spectrometry proteomics, live-cell imaging of mitochondrial networks, ROS measurement\",\n      \"journal\": \"Molecular & cellular proteomics : MCP\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo KD combined with subcellular proteomics and live imaging, multiple orthogonal readouts in single lab\",\n      \"pmids\": [\"41672147\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"REEP5 is a cardiac-enriched ER/SR membrane-shaping protein that maintains SR/ER tubular architecture to support Ca2+ cycling and excitation-contraction coupling in cardiomyocytes; it also interacts with Mitofusins 1/2 to mediate mitochondrial 'hitchhiking' on tubular ER for cytosolic mitochondrial distribution, forms a complex with TRAM1 that engages SARS-CoV-2 NSP3 at replication organelles, and acts as an accessory protein promoting ligand-stimulated endocytosis of CXCR1.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"REEP5 is an endoplasmic/sarcoplasmic reticulum membrane-shaping protein that organizes tubular SR/ER architecture to support cardiomyocyte function, including Ca2+ cycling and excitation-contraction coupling [#0, #1]. Loss of REEP5 destabilizes SR/ER membranes and produces luminal vacuolization, reduced contractility, and disrupted Ca2+ handling in cardiomyocytes, and causes dilated cardiomyopathy-like dysfunction in vivo across zebrafish and mouse models [#0]; ultrastructurally, REEP5 deficiency specifically deforms SR membrane geometry while sparing transverse tubules and Ca2+-handling protein levels, yielding depressed SR Ca2+ release and reduced EC-coupling gain [#1]. Through this membrane-shaping role REEP5 also coordinates inter-organelle contacts: it binds Mitofusins 1 and 2 to enable mitochondrial 'hitchhiking' along tubular ER on microtubules, controlling cytosolic mitochondrial distribution and ER-mitochondria tethering, with REEP5 loss causing perinuclear mitochondrial clustering and altered mitochondrial ROS [#2], and in cardiac tissue its loss fragments mitochondrial networks, raises ROS, and remodels both the SR/ER membrane-shaping proteome and mitochondrial/microsomal proteomes [#6]. Beyond its structural role, REEP5 overexpression suppresses PERK/IRE1\\u03b1/ATF6 ER-stress signaling and apoptosis after myocardial infarction and binds CLEC5A to counter CLEC5A-induced ER stress [#5]. REEP5 additionally acts as an accessory protein promoting ligand-stimulated endocytosis of CXCR1 [#3], and forms a complex with TRAM1 that engages SARS-CoV-2 NSP3 at replication organelles to promote viral replication [#4].\",\n  \"teleology\": [\n    {\n      \"year\": 2016,\n      \"claim\": \"Established a non-structural signaling role for REEP5 by asking whether it modulates GPCR trafficking, showing it is required for ligand-stimulated CXCR1 internalization and downstream signaling.\",\n      \"evidence\": \"Co-IP, siRNA depletion, receptor internalization and \\u03b2-arrestin2 clustering imaging, ERK/invasion assays, xenograft model\",\n      \"pmids\": [\"27966653\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\n        \"Mechanism by which REEP5 promotes CXCR1 endocytosis (vesicle scission vs membrane shaping) not defined\",\n        \"Receptor selectivity beyond CXCR1 vs CXCR2 untested\",\n        \"Relationship to its ER membrane-shaping role unclear\"\n      ]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Resolved whether REEP5 shapes cardiac SR membranes and whether this geometry is functionally coupled to Ca2+ release, distinguishing structural deformation from altered Ca2+-handling protein expression.\",\n      \"evidence\": \"REEP5 knockout rats with FIB-SEM 3D SR reconstruction and simultaneous patch-clamp of Ca2+ currents and transients\",\n      \"pmids\": [\"29431104\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\n        \"How REEP5 physically curves/stabilizes SR membrane at molecular level not shown\",\n        \"Whether the same mechanism operates in non-cardiac ER untested here\"\n      ]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Confirmed REEP5 as essential for SR/ER structural integrity and cardiac function across orthogonal in vitro and in vivo systems, linking membrane destabilization to contractile and Ca2+-cycling failure.\",\n      \"evidence\": \"shRNA/siRNA in mouse cardiomyocytes, CRISPR/Cas9 zebrafish, AAV9 knockdown in mice, Ca2+ imaging and contractility measurements\",\n      \"pmids\": [\"32075961\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\n        \"Whether observed dilated cardiomyopathy phenotype reflects a primary membrane defect or secondary remodeling not fully separated\",\n        \"Human disease relevance not established\"\n      ]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Extended REEP5 function to host-pathogen biology by asking whether it participates in viral replication organelle formation, identifying a REEP5-TRAM1 complex that engages SARS-CoV-2 NSP3.\",\n      \"evidence\": \"Host-viral interactome mapping with co-expression of NSP3/NSP4/NSP6 and co-IP/MS\",\n      \"pmids\": [\"37768083\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\n        \"Direct vs indirect REEP5-NSP3 interaction not resolved\",\n        \"Whether membrane-shaping activity is required for RO formation untested\",\n        \"Limited mechanistic detail on replication promotion\"\n      ]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defined a mechanism for inter-organelle organization by asking how tubular ER positions mitochondria, showing REEP5 binds MFN1/2 to mediate mitochondrial hitchhiking and tethering.\",\n      \"evidence\": \"Co-IP, rapamycin-induced forced dimerization, live-cell imaging, ROS measurement, siRNA and overexpression\",\n      \"pmids\": [\"39133213\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\n        \"Stoichiometry and structural basis of the REEP5-MFN interface unknown\",\n        \"How tethering dynamics regulate ROS mechanistically not defined\"\n      ]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Addressed whether REEP5 protects cardiomyocytes from ER-stress injury, showing its overexpression suppresses PERK/IRE1\\u03b1/ATF6 signaling and CLEC5A-driven apoptosis after myocardial infarction.\",\n      \"evidence\": \"Co-IP (REEP5-CLEC5A), gain-of-function overexpression in MI mouse and hypoxia cell models, western blot for ER-stress markers, apoptosis assays\",\n      \"pmids\": [\"39044150\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\n        \"Whether ER-stress suppression is a direct consequence of CLEC5A binding or of membrane stabilization unclear\",\n        \"Loss-of-function effect on ER stress not tested\",\n        \"Single-lab gain-of-function design\"\n      ]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Connected REEP5 SR/ER shaping to mitochondrial health at the proteome level, showing cardiac REEP5 loss fragments mitochondria, raises ROS, and remodels membrane-shaping and mitochondrial proteomes.\",\n      \"evidence\": \"AAV9-shRNA knockdown in mouse heart, subcellular fractionation with DIA mass spectrometry, live imaging of mitochondrial networks, ROS measurement\",\n      \"pmids\": [\"41672147\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\n        \"Causal chain from SR/ER membrane defect to mitochondrial import/antioxidant proteome changes not established\",\n        \"Whether RTN4/ATL3/CKAP4 upregulation is true functional compensation untested\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How REEP5's intrinsic membrane-curving biochemistry generates SR/ER tubule geometry, and how this single activity is shared across its cardiac, inter-organelle, GPCR-trafficking, and viral roles, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\n        \"No structural model of REEP5 membrane-shaping mechanism in the corpus\",\n        \"No unifying biochemical assay linking the disparate functional contexts\",\n        \"No human Mendelian disease link established in the corpus\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [0, 2, 5]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-397014\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"R-HSA-1852241\", \"supporting_discovery_ids\": [2, 6]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"MFN1\", \"MFN2\", \"CXCR1\", \"TRAM1\", \"CLEC5A\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":4,"faith_total":5,"faith_pct":80.0}}