{"gene":"APLNR","run_date":"2026-06-09T22:02:43","timeline":{"discoveries":[{"year":2012,"finding":"APJ acts as a dual-function receptor: apelin stimulation couples APJ to Gαi signaling and elicits a cardioprotective response, whereas mechanical stretch signals through APJ in a G-protein-independent, β-arrestin-dependent manner to induce cardiomyocyte hypertrophy. Knockdown of β-arrestins blocked stretch-mediated hypertrophy, and pharmacological doses of apelin acting through Gαi prevented it.","method":"Genetic loss-of-function (APJ-null and apelin-null mice), freshly isolated cardiomyocyte stretch assay, β-arrestin knockdown, Gαi pharmacology, cardiomyocyte size measurement","journal":"Nature","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal genetic and pharmacological approaches in one study, replicated across in vivo and in vitro models, published in Nature","pmids":["22810587"],"is_preprint":false},{"year":2014,"finding":"ERG is a transcriptional activator of the APLNR gene and binds to the Aplnr promoter. Knockout of either Erg or Aplnr causes pulmonary venule-specific endothelial proliferation in vitro and pulmonary veno-occlusive disease in vivo, placing APLNR downstream of ERG in a pathway essential for venular endothelial homeostasis.","method":"Erg/Aplnr knockout mice, endothelium-directed conditional Aplnr deletion, in vitro proliferation assays, ChIP/transcriptional activation studies, patient lung tissue analysis","journal":"Circulation","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic epistasis with both global and endothelium-specific knockouts, in vitro mechanistic follow-up, and human patient validation","pmids":["25062690"],"is_preprint":false},{"year":2016,"finding":"APLNR-mediated APLN/APLNR signaling in endothelial cells induces transcription of miR-139-5p (promoted by laminar flow), which in turn suppresses endothelial CXCR4 expression. Loss of Apln, Aplnr, or endothelial-specific Aplnr caused dysregulated CXCR4 upregulation and retinal vascular defects; pharmacological inhibition of CXCR4 or augmentation of the miR-139-5p axis rescued these defects.","method":"Apln/Aplnr global and endothelial-specific knockout mice, retinal vascular phenotyping, miR-139-5p in vivo inhibition, pharmacological CXCR4 blockade","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple genetic knockouts (global and tissue-specific), pharmacological rescue experiments, and mechanistic miRNA pathway delineation in one study","pmids":["27068353"],"is_preprint":false},{"year":2017,"finding":"Endothelial APLNR signaling inactivates FOXO1 and suppresses endothelial expression of fatty acid binding protein 4 (FABP4). Conditional endothelial-specific Aplnr deletion impaired glucose utilization and abrogated apelin-induced glucose lowering; excess tissue fatty acid accumulation in skeletal muscle was the mechanistic link. Concurrent endothelial Foxo1 deletion or pharmacologic FABP4 inhibition rescued the metabolic phenotype.","method":"Endothelial-specific conditional Aplnr knockout mice, FOXO1 knockout epistasis, pharmacological FABP4 inhibition, metabolic and glucose utilization assays","journal":"Science translational medicine","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional knockout with defined cellular phenotype, epistasis with Foxo1, pharmacological rescue, multiple orthogonal metabolic readouts in one study","pmids":["28904225"],"is_preprint":false},{"year":2003,"finding":"Apelin peptides are endogenous ligands for the human APJ (APLNR) receptor. Structure-activity studies identified that leucine at position 5 and arginine at positions 2 and 4 of apelin-13 are key residues required for functional potency and binding affinity at the recombinant human APJ receptor.","method":"Radioligand binding assays, functional assays with apelin analogues at human recombinant APJ receptor, quantitative RT-PCR, immunohistochemistry","journal":"Journal of neurochemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro receptor binding and functional assays with systematic mutagenesis of the ligand, replicated across multiple analogue series","pmids":["12603839"],"is_preprint":false},{"year":2003,"finding":"APJ receptor undergoes rapid, dose-dependent ligand-induced internalization upon stimulation with Apelin-36 and Apelin-13 via clathrin-coated pits (co-localization with transferrin receptor). The intact cytoplasmic C-terminal domain of APJ is required for ligand-induced internalization. Internalized APJ recycled to the cell surface within 60 min after Apelin-13 removal, but most internalized receptor remained in the cytoplasm 2 h after Apelin-36 washout.","method":"APJ-GFP stable expression, fluorescent apelin peptide (5-CF-Apelin-13), co-localization with transferrin receptor, C-terminal truncation mutants, live cell imaging","journal":"Virology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — reconstituted receptor internalization in multiple cell types with mutagenesis (C-terminal domain) and co-localization with established clathrin pathway marker","pmids":["12667811"],"is_preprint":false},{"year":2005,"finding":"Apelin/APJ signaling via a pertussis toxin-sensitive Gi protein activates PI3K→Akt/PKB and focal adhesion kinase (FAK) phosphorylation, increases focal adhesion formation with actin reorganization, and stimulates cell motility in APJ-expressing cells.","method":"Stable APJ-expressing HEK293T cells, radioligand binding, pertussis toxin treatment, PI3K inhibitor (LY294002), phospho-Akt/FAK immunoblotting, scratch migration assay, F-actin staining","journal":"International journal of molecular medicine","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with defined receptor, multiple pharmacological inhibitors blocking defined signaling nodes, and orthogonal functional readouts (phosphorylation, morphology, migration)","pmids":["16211245"],"is_preprint":false},{"year":2000,"finding":"APJ functions as a coreceptor for primate immunodeficiency viruses (HIV-1 and SIV). APJ coreceptor activity is demonstrable even at low expression levels, and the endogenous peptide ligand apelin-13 efficiently blocked APJ coreceptor activity. APJ is N-glycosylated as determined by Western blot with a specific monoclonal antibody.","method":"Monoclonal antibody (MAb 856) production, FACS, Western blot, immunofluorescence, cell-cell fusion assays, apelin-13 blockade of coreceptor activity","journal":"Virology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — functional coreceptor assays with apelin-13 blockade, glycosylation determination by Western blot, multiple detection methods in one study","pmids":["11040134"],"is_preprint":false},{"year":2011,"finding":"Non-activated (ligand-free) APJ suppresses angiotensin II (AngII)/AT1-mediated ERK1/2 phosphorylation in a receptor-proximity-dependent manner independent of heterodimerization, whereas apelin-activated APJ reverses this suppression through Gαi (pertussis toxin-sensitive). Thus APJ has a constitutive, ligand-independent inhibitory interaction with AT1 signaling that is abolished upon apelin binding.","method":"HEK293 cell co-expression of APJ and AT1, pertussis toxin treatment, AT2 and β2-adrenergic receptor controls, ERK1/2 phosphorylation immunoblotting, AT1 blocker dose-response","journal":"Hypertension research","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — defined signaling mechanism with pharmacological controls, but single lab and single method (ERK1/2 phosphorylation) as primary readout","pmids":["21412239"],"is_preprint":false},{"year":2016,"finding":"Apelin/APLNR signaling promotes endothelial cell (EC) proliferation via PI3K/Akt-mediated activation, and apelin-12 activates NO production via the PI3K/Akt pathway in human ECs. Apelin-13 additionally activates Erk1/2 phosphorylation and enhances EC proliferation. APJ knockdown inhibited PI3K phosphorylation and impaired flow-induced eNOS and PECAM-1 expression; APJ expression is induced by shear stress independently of its ligand.","method":"siRNA knockdown of APJ or apelin in human ECs, shear stress flow chamber, NO measurement, PI3K/Akt and ERK1/2 phosphorylation immunoblotting, gene and protein expression under flow","journal":"Cellular signalling","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — defined signaling nodes with siRNA knockdown and pharmacological context, single lab, several orthogonal readouts","pmids":["25817266"],"is_preprint":false},{"year":2014,"finding":"APJ receptor acts as a static pressure sensor in cardiomyocytes. Static pressure upregulates APJ expression and activates PI3K/Akt/autophagy (LC3-II/I, Beclin-1) signaling to promote cardiomyocyte hypertrophy. APJ shRNA, PI3K inhibitor (LY294002), Akt inhibitor, and autophagy inhibitor (3-methyladenine) each reversed pressure-induced increases in cell diameter, volume, and protein content.","method":"Rat left ventricular hypertrophy model (aortic coarctation), H9c2 cardiomyocyte static pressure culture, APJ shRNA knockdown, PI3K/Akt inhibitors, LC3/Beclin-1 immunoblotting, cell size measurements","journal":"Acta biochimica et biophysica Sinica","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — in vivo and in vitro models with shRNA and pharmacological inhibitors, single lab, defined pathway but no rescue experiments","pmids":["24966188"],"is_preprint":false},{"year":2015,"finding":"Hypoxia-induced HIF-1α upregulates apelin and APLNR expression in bone marrow-derived mesenchymal stem cells (BMSCs), and this apelin/APJ/autophagy axis mediates hypoxia-induced BMSC proliferation. siRNA-HIF-1α suppressed apelin, APJ, Beclin-1, and LC3II/LC3I; siRNA-APJ suppressed Beclin-1 and LC3II/LC3I and reversed proliferation; siRNA-Beclin-1 abolished proliferation.","method":"Mouse BMSC hypoxia culture, siRNA knockdown of HIF-1α/APJ/Beclin-1, MTT and BrdU proliferation assays, Western blot for autophagy markers","journal":"Acta biochimica et biophysica Sinica","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — genetic epistasis via sequential siRNA knockdown establishing HIF-1α→apelin/APJ→autophagy→proliferation pathway, single lab","pmids":["25736405"],"is_preprint":false},{"year":2015,"finding":"Hypoxia upregulates HIF-1α, Apelin, and APLNR in endothelial progenitor cells (EPCs), and the Apelin/APLNR axis mediates hypoxia-induced EPC proliferation via downstream MAPK signaling. siRNA knockdown of Apelin or APLNR suppressed hypoxia-induced proliferation; MAPK inhibitors (SB-239063 and PD98059) eliminated Apelin upregulation-induced EPC proliferation.","method":"Human EPC hypoxia culture, siRNA knockdown of Apelin/APLNR, MAPK inhibitors, MTT proliferation assay, RT-qPCR and Western blot","journal":"Molecular medicine reports","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — siRNA knockdown and pharmacological inhibition establishing signaling pathway, single lab","pmids":["26676468"],"is_preprint":false},{"year":2012,"finding":"APLNR (Aplnr) is specifically expressed on circulating cKit+/Flk1+ cells and functions as a receptor for apelin-mediated chemoattraction during myocardial repair. Apelin injection into ischemic myocardium increased recruitment of cKit+/Flk1+/Aplnr+ cells; Aplnr knockdown in bone marrow aggravated ischemic damage and could not be rescued by apelin. Recruited cells promoted neovascularization via paracrine rather than transdifferentiation mechanisms.","method":"Bone marrow Aplnr knockdown, apelin injection into ischemic myocardium, flow cytometry for circulating progenitor populations, scar formation and cardiac function measurement","journal":"Circulation research","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — in vivo receptor knockdown with defined progenitor cell population and functional cardiac readouts, single lab","pmids":["22753078"],"is_preprint":false},{"year":2016,"finding":"APLN (apelin) acts through APLNR to increase steroidogenesis in human luteinized granulosa cells (hGCs), enhancing both basal and IGF1-induced steroid secretion by increasing HSD3B protein concentration through activation of the MAPK3/1 (ERK1/2) and AKT pathways. The APLNR antagonist ML221 reversed these effects. IGF1 increased APLNR expression in hGCs.","method":"Cultured human luteinized granulosa cells, recombinant apelin-13 and apelin-17 treatment, ML221 APLNR antagonist, pharmacological inhibitors of AKT and MAPK3/1 pathways, Western blot, steroid assays","journal":"Biology of reproduction","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — receptor antagonist and pathway inhibitors with defined steroidogenic readout, single lab","pmids":["27683264"],"is_preprint":false},{"year":2022,"finding":"In Sertoli cells, high glucose induces local APLN production, which via APJ hyperactivation suppresses carnitine production and represses cell adhesion gene expression, leading to blood-testis barrier (BTB) structural dysfunction and impaired spermatogenesis in diabetic models. Pharmacological blockade of APJ with ML221 ameliorated BTB damage and improved spermatogenesis in diabetic db/db mice and cultured human testes.","method":"Single-cell RNA sequencing of diabetic patient testes (STRT-seq), high glucose treatment of Sertoli cells, APJ antagonist (ML221) treatment in db/db mice and human testis culture, BTB structural assessment","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — scRNA-seq plus pharmacological rescue in both mouse and human models, defined mechanistic pathway (APLN→APJ→carnitine/adhesion→BTB dysfunction), replicated across species","pmids":["36443325"],"is_preprint":false},{"year":2019,"finding":"In glioblastoma, APLNR on tumor cells mediates apelin-induced migration and invasion. Apelin reduction led to accelerated tumor cell invasion by APLNR-positive cells. Mutant APLNR ligand apelin-F13A blocked both tumor angiogenesis and GBM cell invasion, and cotargeting VEGFR2 and APLNR synergistically improved survival in proneural GBM mouse models.","method":"APLN knockdown/knockout in orthotopic GBM models, apelin-F13A peptide treatment, VEGFR2 + APLNR cotargeting, stereotactic biopsy analysis, in vitro and in vivo invasion assays","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic knockdown/knockout with in vivo rescue experiments and pharmacological ligand, two orthogonal readouts (angiogenesis and invasion), single lab","pmids":["30358318"],"is_preprint":false},{"year":2019,"finding":"APJ expression in ovarian cancer cells is necessary and sufficient for pro-metastatic phenotypes (proliferation, adhesion, anoikis resistance, migration, invasion) via downstream activation of STAT3, ERK, and AKT pathways. APJ inhibitor ML221 efficiently inhibited these phenotypes in vitro, and APJ overexpression increased metastasis in vivo.","method":"APJ overexpression and knockdown in ovarian cancer cell lines, in vitro adhesion/migration/invasion assays, anoikis assay, in vivo metastasis model, STAT3/ERK/AKT phosphorylation immunoblotting, ML221 inhibitor treatment","journal":"Molecular cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — gain- and loss-of-function in vitro plus in vivo validation with defined pathway components, single lab","pmids":["30858172"],"is_preprint":false},{"year":2020,"finding":"APJ and bradykinin B2 receptor (B2R) form a functional heterodimer at the cell membrane, demonstrated by BRET, FRET, proximity ligation assay, and co-immunoprecipitation in HUVECs and transfected cells. Stimulation with apelin-13 and bradykinin increased eNOS phosphorylation via the APJ-B2R heterodimer through a PLC/ERK1/2/eNOS signaling pathway, leading to increased cell proliferation. Silencing of either APJ or B2R inhibited eNOS phosphorylation.","method":"BRET and FRET resonance energy transfer, proximity ligation assay, co-immunoprecipitation in HUVECs and HEK293 cells, siRNA knockdown of APJ or B2R, ERK1/2 and eNOS phosphorylation assays, cell proliferation assay","journal":"Cellular signalling","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — heterodimer formation confirmed by multiple biophysical methods (BRET, FRET, PLA, Co-IP) plus functional signaling validation with siRNA knockdown, single lab","pmids":["32407761"],"is_preprint":false},{"year":2023,"finding":"ELA (Elabela) binds to APJ and activates the NRF2/ARE antioxidative signaling pathway via Gα13, thereby reducing neuronal ferroptosis after cerebral ischemia/reperfusion injury. AAV-mediated APJ knockdown or the NRF2 inhibitor ML385 abolished the protective effects of ELA-32.","method":"ELA-32 peptide treatment in cerebral I/R mouse model, AAV-APJ-RNAi knockdown, NRF2 inhibitor ML385, iron deposition, lipid peroxidation, mitochondrial morphology assays, behavioral readouts","journal":"Free radical biology & medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — defined signaling axis (APJ→Gα13→NRF2/ARE) with genetic (AAV knockdown) and pharmacological (NRF2 inhibitor) validation, single lab","pmids":["36681202"],"is_preprint":false},{"year":2023,"finding":"ELA (Elabela) binds to APJ in renal tubular cells to regulate renal microvascular blood flow through two downstream mediators: arginine metabolizing enzyme ARG2 and PGE2 metabolizing enzymes CBR1/3. APJ inhibitor ML221 blocked the beneficial effects of exogenous ELA-32 on AKI, while combination treatment with ARG2 inhibitor nor-NOHA and PGE2 activator Paricalcitol alleviated injury independently of APJ, placing ARG2 and CBR1/3 downstream of the ELA-APJ axis.","method":"Renal tubule-specific Apela (ELA) knockout mice, bilateral/unilateral I/R models, RNA sequencing, ML221 APJ inhibitor, ARG2 inhibitor (nor-NOHA) and Paricalcitol combination treatment, renal blood flow and functional measurements","journal":"Theranostics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — tissue-specific knockout plus RNA-seq pathway discovery plus pharmacological rescue, multiple orthogonal approaches, single lab","pmids":["37351176"],"is_preprint":false},{"year":1999,"finding":"The murine msr/apj receptor (ortholog of human APJ/APLNR) is expressed in endothelium of primary blood vessels and the forming heart, as well as in somites, limb bud, and branchial arches during embryonic development, indicating a role in endothelial/vascular lineage specification distinct from Flk1.","method":"Molecular cloning, in situ hybridization in developing mouse embryo, comparative expression with Flk1","journal":"Mechanisms of development","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — direct localization by in situ hybridization in developing tissues, replicated across developmental stages, but no functional consequence tested","pmids":["10473142"],"is_preprint":false},{"year":2003,"finding":"APLNR (APJ receptor) mRNA is upregulated in the hypothalamic parvocellular paraventricular nucleus (pPVN) by acute and repeated restraint stress, and adrenalectomy also increased APJR mRNA in the PVN. Adrenalectomized rats showed no further increase above baseline after stress, indicating glucocorticoids negatively regulate APJR mRNA expression and mediate stress-induced regulation.","method":"In situ hybridization for APJR mRNA in rat hypothalamus, restraint stress paradigms, adrenalectomy, dual-label in situ hybridization to co-localize APJR and vasopressin mRNA","journal":"Journal of neuroendocrinology","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — defined neuroendocrine regulation of APLNR expression by glucocorticoids established via adrenalectomy and stress models, single lab","pmids":["14622440"],"is_preprint":false},{"year":2003,"finding":"APLNR (APJR) mRNA is co-expressed with vasopressin in magnocellular neurons of the hypothalamic PVN and SON, and its expression is induced by osmotic stimuli (2% NaCl loading and water deprivation). Salt-loading increased co-localization of APJR and vasopressin mRNAs in the SON, supporting a role for APJ in the autocrine/paracrine regulation of vasopressin-containing neurons and fluid homeostasis.","method":"In situ hybridization histochemistry for APJR mRNA, dual-label in situ hybridization for APJR and vasopressin in salt-loaded and water-deprived rats","journal":"Journal of neuroendocrinology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — direct localization with functional osmotic stimulus paradigm and dual-label co-localization, replicated across osmotic challenge conditions, single lab","pmids":["12787050"],"is_preprint":false},{"year":2017,"finding":"APJ activation by apelin improves AngII-induced endothelial cell senescence via the AMPK/SIRT1 signaling pathway. APJ, AMPK, or SIRT1 knockdown each attenuated the protective effects of apelin. Apelin reduced AngII-induced ROS generation and enhanced telomerase activity in HUVECs.","method":"AngII-induced HUVEC senescence model, SA-β-Gal assay, siRNA knockdown of APJ/AMPK/SIRT1, ROS detection, telomerase activity (RQ-TRAP), CCK-8 viability assay, Western blot for P21 and PAI-1","journal":"Archives of medical science","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, endpoint-driven assays with siRNA knockdown but limited mechanistic depth beyond pathway identification","pmids":["30002688"],"is_preprint":false},{"year":2019,"finding":"APLNR overexpression in nasopharyngeal carcinoma (NPC) cells inhibited migration, invasion, and EMT. Low APLNR expression activated the PI3K-AKT-mTOR signaling pathway to promote EMT. ATRA treatment upregulated APLNR in NPC cell lines and inhibited proliferation; knockdown of APLNR diminished ATRA-induced growth inhibition.","method":"APLNR overexpression and knockdown in NPC cell lines, wound-healing and Transwell migration/invasion assays, Western blot for EMT markers and PI3K-AKT-mTOR pathway components, ATRA treatment, cell cycle analysis","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — gain- and loss-of-function with defined EMT pathway, ATRA epistasis, single lab","pmids":["31408612"],"is_preprint":false},{"year":2021,"finding":"Adipocyte-derived apelin activates APJ on ovarian cancer cells in a paracrine manner, promoting lipid uptake via CD36 upregulation through APJ-STAT3 activation, and the accumulated lipids are utilized for fatty acid oxidation via AMPK-CPT1a axis. APJ antagonist F13A or APJ knockdown reversed lipid accumulation, migration, invasion, and omental homing in vivo.","method":"Co-culture with 3T3-L1 adipocyte conditioned media, APJ antagonist F13A, APJ knockdown, in vitro migration/invasion assays, ex vivo omentum adhesion, in vivo homing assay, CD36 and STAT3 pathway analysis, lipid droplet staining","journal":"Molecular cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — paracrine signaling mechanism defined with receptor knockdown, antagonist, and in vivo validation; multiple orthogonal readouts, single lab","pmids":["34172534"],"is_preprint":false},{"year":2021,"finding":"APLNR abrogates the stimulatory effects of 17β-estradiol on OVCAR-3 epithelial ovarian cancer cell proliferation and of IGF-1 on COV434 granulosa cancer cell proliferation via crosstalk between APLNR and estrogen receptor alpha (ERα) or IGF-1 receptor (IGF1R), respectively, independently of ERK1/2 and PI3K pathways.","method":"OVCAR-3 and COV434 cell proliferation assays, apelin treatment, ERK1/2 and PI3K pathway inhibitors, estrogen and IGF-1 stimulation, APLNR-ERα/IGF1R crosstalk analysis","journal":"Molecular biology reports","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, limited mechanistic detail on the receptor crosstalk mechanism beyond pharmacological inhibitor results","pmids":["31538301"],"is_preprint":false},{"year":2023,"finding":"ELA (Elabela) binds APJ on brain endothelial cells and activates the YAP/TAZ signaling pathway, promoting post-ischemic cerebral angiogenesis. Silencing of APJ or pharmacological blockade of YAP abolished the pro-angiogenic effects of ELA-32 in oxygen-glucose deprivation/reoxygenation conditions.","method":"ELA-32 treatment of bEnd.3 cells under OGD/R, APJ siRNA knockdown, YAP pharmacological blockade, RNA sequencing, in vivo cerebral I/R model with CBF measurement","journal":"Translational research","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — mechanistic signaling defined (APJ→YAP/TAZ) with genetic and pharmacological inhibition, RNA-seq pathway analysis, single lab","pmids":["36813109"],"is_preprint":false}],"current_model":"APLNR (APJ) is a class A GPCR that functions as a bifunctional receptor: its canonical endogenous ligands apelin (and Elabela) couple it primarily to Gαi to drive cardioprotective, vasodilatory, pro-angiogenic, and metabolic effects via downstream PI3K/AKT, ERK1/2, AMPK, and NRF2/ARE signaling, while mechanical stretch signals through APJ in a G-protein-independent, β-arrestin-dependent manner to induce cardiomyocyte hypertrophy; ligand binding induces rapid clathrin-mediated receptor internalization requiring the intact C-terminal domain; in endothelial cells, APJ signaling induces miR-139-5p to suppress CXCR4, inactivates FOXO1 to suppress FABP4 expression and regulate fatty acid uptake, and is transcriptionally activated by the ERG transcription factor to maintain venular homeostasis; in Sertoli cells, hyperactivated APJ suppresses carnitine production and cell adhesion to impair the blood-testis barrier; non-activated APJ constitutively suppresses AT1 receptor signaling while apelin-activated APJ relieves this suppression; and APJ can heterodimerize with B2R to activate PLC/ERK1/2/eNOS-dependent cell proliferation."},"narrative":{"mechanistic_narrative":"APLNR (APJ) is a class A G-protein-coupled receptor that integrates peptide-ligand and mechanical inputs to control vascular, cardiac, metabolic, and reproductive physiology [PMID:22810587, PMID:12603839]. Its endogenous apelin peptides bind the human receptor with potency dictated by defined residues of apelin-13 [PMID:12603839], coupling APJ predominantly to pertussis-toxin-sensitive Gαi to activate PI3K→Akt and FAK signaling that drives focal adhesion formation, actin reorganization, and cell motility [PMID:16211245]; the second peptide ligand Elabela engages APJ to activate Gα13-dependent NRF2/ARE antioxidative signaling and YAP/TAZ-dependent angiogenesis [PMID:36681202, PMID:36813109]. APJ is bifunctional: apelin/Gαi signaling is cardioprotective, whereas mechanical stretch or static pressure signals through APJ in a G-protein-independent, β-arrestin-dependent manner to drive cardiomyocyte hypertrophy [PMID:22810587, PMID:24966188]. Ligand binding triggers rapid clathrin-mediated internalization that requires the intact cytoplasmic C-terminal domain, with apelin-13-bound receptor recycling to the surface [PMID:12667811]. In the endothelium APJ acts downstream of the ERG transcription factor to maintain venular homeostasis [PMID:25062690] and orchestrates a miR-139-5p→CXCR4 axis controlling developmental vascular patterning [PMID:27068353] and a FOXO1→FABP4 axis governing endothelial fatty-acid handling and systemic glucose utilization [PMID:28904225]. APJ further modulates receptor crosstalk, constitutively suppressing AngII/AT1-mediated ERK1/2 signaling in the ligand-free state and relieving this suppression upon apelin binding [PMID:21412239], and forming a functional heterodimer with the bradykinin B2 receptor that drives PLC/ERK1/2/eNOS-dependent endothelial proliferation [PMID:32407761]. In disease, APJ hyperactivation by locally produced apelin suppresses carnitine production and adhesion gene expression in Sertoli cells to disrupt the blood-testis barrier in diabetes [PMID:36443325], and APJ drives pro-metastatic and pro-invasive programs in ovarian cancer and glioblastoma via STAT3, ERK, and AKT signaling [PMID:30358318, PMID:30858172].","teleology":[{"year":2000,"claim":"Established that APJ is a functional cell-surface receptor with biological activity beyond an orphan GPCR by showing it serves as a primate immunodeficiency virus coreceptor blockable by its peptide ligand.","evidence":"Cell-cell fusion and coreceptor assays with apelin-13 blockade and Western blot glycosylation analysis","pmids":["11040134"],"confidence":"High","gaps":["Does not define physiological signaling output","Coreceptor role distinct from native ligand signaling"]},{"year":2003,"claim":"Defined the molecular determinants of apelin binding to human APJ, establishing the ligand pharmacophore.","evidence":"Radioligand binding and functional assays with systematic apelin-13 analogue mutagenesis at recombinant human APJ","pmids":["12603839"],"confidence":"High","gaps":["No receptor structural model","Downstream signaling not resolved in this study"]},{"year":2003,"claim":"Answered how APJ signaling is terminated and reset by demonstrating C-terminal-dependent clathrin-mediated internalization and ligand-specific recycling.","evidence":"APJ-GFP live-cell imaging, fluorescent apelin, transferrin receptor co-localization, and C-terminal truncation mutants","pmids":["12667811"],"confidence":"High","gaps":["Specific C-terminal motifs/phosphosites not mapped","Arrestin involvement in internalization not directly tested"]},{"year":2005,"claim":"Identified the proximal G-protein-coupled signaling cascade, linking APJ to Gi-PI3K/Akt-FAK and cell motility.","evidence":"Stable APJ-HEK293T cells with pertussis toxin, PI3K inhibitor, phospho-immunoblotting, and migration assays","pmids":["16211245"],"confidence":"High","gaps":["Cell-type generality untested","Does not address mechanical/biased signaling"]},{"year":2011,"claim":"Revealed that ligand-free APJ has constitutive activity by suppressing AT1/AngII ERK1/2 signaling that apelin binding relieves.","evidence":"HEK293 APJ/AT1 co-expression, pertussis toxin, receptor controls, ERK1/2 immunoblotting","pmids":["21412239"],"confidence":"Medium","gaps":["Single ERK1/2 readout from one lab","Physiological/in vivo relevance untested"]},{"year":2012,"claim":"Defined APJ as a bifunctional receptor distinguishing protective apelin/Gαi signaling from pathological stretch-induced β-arrestin-dependent hypertrophy.","evidence":"APJ-null and apelin-null mice, isolated cardiomyocyte stretch assay, β-arrestin knockdown, Gαi pharmacology","pmids":["22810587"],"confidence":"High","gaps":["Structural basis of biased mechanosensing unknown","Arrestin effector identity in hypertrophy unresolved"]},{"year":2012,"claim":"Showed APJ functions as an apelin chemoattractant receptor on circulating progenitor cells supporting myocardial repair.","evidence":"Bone marrow Aplnr knockdown, apelin injection into ischemic myocardium, flow cytometry, cardiac function readouts","pmids":["22753078"],"confidence":"Medium","gaps":["Single lab","Paracrine effectors not identified"]},{"year":2014,"claim":"Placed APLNR in a transcriptional hierarchy by identifying ERG as a direct activator required for venular endothelial homeostasis.","evidence":"Erg/Aplnr knockout mice, endothelium-specific deletion, ChIP/promoter binding, human lung tissue","pmids":["25062690"],"confidence":"High","gaps":["Downstream effectors of APLNR in venular ECs not fully defined here","Ligand dependence of phenotype unclear"]},{"year":2014,"claim":"Extended the mechanosensor role to static pressure, linking APJ to PI3K/Akt/autophagy-driven cardiomyocyte hypertrophy.","evidence":"Rat hypertrophy model, H9c2 static pressure culture, APJ shRNA, PI3K/Akt/autophagy inhibitors","pmids":["24966188"],"confidence":"Medium","gaps":["No rescue experiments","Relationship to β-arrestin stretch pathway not reconciled"]},{"year":2015,"claim":"Identified a HIF-1α→apelin/APJ→autophagy axis driving hypoxia-induced proliferation in stem and progenitor cells.","evidence":"BMSC and EPC hypoxia cultures, sequential siRNA knockdown of HIF-1α/APJ/Beclin-1, MAPK inhibitors, proliferation assays","pmids":["25736405","26676468"],"confidence":"Medium","gaps":["Single lab per cell type","Direct receptor-effector coupling not biochemically resolved"]},{"year":2016,"claim":"Defined an endothelial APJ→miR-139-5p→CXCR4 axis controlling vascular patterning, induced by laminar flow.","evidence":"Apln/Aplnr global and endothelial-specific knockouts, retinal phenotyping, miR-139-5p inhibition, CXCR4 blockade","pmids":["27068353"],"confidence":"High","gaps":["Mechanism of miR-139-5p induction by APJ unresolved","Flow-sensing versus ligand contribution not separated"]},{"year":2016,"claim":"Connected APJ to ligand-independent shear-stress responses and NO/proliferation signaling in endothelium.","evidence":"siRNA knockdown in human ECs, shear-stress flow chamber, NO measurement, PI3K/Akt/ERK1/2 immunoblotting","pmids":["25817266"],"confidence":"Medium","gaps":["Single lab","Mechanism of shear-induced APJ upregulation unknown"]},{"year":2017,"claim":"Established APJ as a regulator of systemic metabolism through an endothelial FOXO1→FABP4 fatty-acid handling axis.","evidence":"Endothelial-specific Aplnr knockout, Foxo1 epistasis, FABP4 inhibition, glucose utilization assays","pmids":["28904225"],"confidence":"High","gaps":["Direct biochemical link from APJ to FOXO1 inactivation not mapped","G-protein dependence not dissected"]},{"year":2019,"claim":"Demonstrated APJ drives pro-metastatic and invasive tumor programs via STAT3/ERK/AKT in ovarian cancer and glioblastoma.","evidence":"Gain/loss-of-function in tumor cells, in vivo metastasis/invasion models, ML221 and apelin-F13A antagonists, VEGFR2 cotargeting","pmids":["30858172","30358318"],"confidence":"Medium","gaps":["Single lab per tumor type","Context-dependent tumor-suppressive role elsewhere unexplained"]},{"year":2020,"claim":"Showed APJ engages in receptor crosstalk by heterodimerizing with the bradykinin B2 receptor to drive eNOS-dependent endothelial proliferation.","evidence":"BRET, FRET, proximity ligation, co-IP in HUVECs/HEK293, siRNA knockdown, ERK1/2/eNOS phosphorylation","pmids":["32407761"],"confidence":"Medium","gaps":["Stoichiometry and structural interface unknown","In vivo relevance of heterodimer untested"]},{"year":2022,"claim":"Defined a pathological APJ hyperactivation mechanism disrupting the blood-testis barrier in diabetes via carnitine and adhesion suppression.","evidence":"scRNA-seq of diabetic testes, high-glucose Sertoli cell treatment, ML221 rescue in db/db mice and human testis culture","pmids":["36443325"],"confidence":"High","gaps":["Signaling node linking APJ to carnitine repression unmapped","Receptor coupling (G-protein vs arrestin) not defined"]},{"year":2023,"claim":"Established Elabela as a second APJ ligand engaging distinct Gα13/NRF2 and YAP/TAZ pathways in protective cytoprotection and angiogenesis.","evidence":"ELA-32 treatment in cerebral and renal I/R models, AAV/siRNA APJ knockdown, NRF2 and YAP inhibitors, RNA-seq","pmids":["36681202","36813109","37351176"],"confidence":"Medium","gaps":["Ligand bias between apelin and Elabela not structurally defined","Single lab per organ system"]},{"year":null,"claim":"How APJ structurally encodes biased signaling among Gαi, Gα13, β-arrestin, and mechanical inputs, and how distinct ligands and heterodimers select these outputs, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No receptor structure or biased-agonism model in the corpus","Mechanism of mechanosensing unknown","Determinants of apelin vs Elabela pathway selection undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060089","term_label":"molecular transducer activity","supporting_discovery_ids":[0,4,6]},{"term_id":"GO:0048018","term_label":"receptor ligand activity","supporting_discovery_ids":[4]},{"term_id":"GO:0001618","term_label":"virus receptor activity","supporting_discovery_ids":[7]},{"term_id":"GO:0120274","term_label":"virus coreceptor activity","supporting_discovery_ids":[7]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[5,7,18]},{"term_id":"GO:0031410","term_label":"cytoplasmic vesicle","supporting_discovery_ids":[5]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[0,6,8]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[1,2,21]},{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[3,26]},{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[10,11]}],"complexes":["APJ-B2R heterodimer"],"partners":["APLN","APELA","BDKRB2","AGTR1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P35414","full_name":"Apelin receptor","aliases":["Angiotensin receptor-like 1","G-protein coupled receptor APJ","G-protein coupled receptor HG11"],"length_aa":380,"mass_kda":42.7,"function":"G protein-coupled receptor for peptide hormones apelin (APLN) and apelin receptor early endogenous ligand (APELA/ELA), that plays a role in the regulation of normal cardiovascular function and fluid homeostasis (PubMed:11090199, PubMed:22810587, PubMed:25639753, PubMed:28137936, PubMed:35817871, PubMed:38428423). When acting as apelin receptor, activates both G(i) protein pathway that inhibits adenylate cyclase activity, and the beta-arrestin pathway that promotes internalization of the receptor (PubMed:11090199, PubMed:25639753, PubMed:28137936, PubMed:35817871, PubMed:38428423). APLNR/APJ also functions as mechanoreceptor that is activated by pathological stimuli in a G-protein-independent fashion to induce beta-arrestin signaling, hence eliciting cardiac hypertrophy (PubMed:22810587, PubMed:38428423). However, the presence of apelin ligand blunts cardiac hypertrophic induction from APLNR/APJ on response to pathological stimuli (PubMed:22810587, PubMed:38428423). Plays a key role in early development such as gastrulation, blood vessels formation and heart morphogenesis by acting as a APELA receptor (By similarity). May promote angioblast migration toward the embryonic midline, i.e. the position of the future vessel formation, during vasculogenesis (By similarity). Promotes sinus venosus (SV)-derived endothelial cells migration into the developing heart to promote coronary blood vessel development (By similarity). Also plays a role in various processes in adults such as regulation of blood vessel formation, blood pressure, heart contractility and heart failure (PubMed:25639753, PubMed:28137936) (Microbial infection) Alternative coreceptor with CD4 for HIV-1 infection; may be involved in the development of AIDS dementia (PubMed:11090199)","subcellular_location":"Cell membrane","url":"https://www.uniprot.org/uniprotkb/P35414/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/APLNR","classification":"Not Classified","n_dependent_lines":3,"n_total_lines":1208,"dependency_fraction":0.0024834437086092716},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/APLNR","total_profiled":1310},"omim":[{"mim_id":"615594","title":"APELIN RECEPTOR EARLY ENDOGENOUS LIGAND; APELA","url":"https://www.omim.org/entry/615594"},{"mim_id":"600052","title":"APELIN RECEPTOR; APLNR","url":"https://www.omim.org/entry/600052"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"brain","ntpm":77.5},{"tissue":"lymphoid tissue","ntpm":101.5},{"tissue":"placenta","ntpm":87.2}],"url":"https://www.proteinatlas.org/search/APLNR"},"hgnc":{"alias_symbol":["FLJ90771","APJ","APJR"],"prev_symbol":["AGTRL1"]},"alphafold":{"accession":"P35414","domains":[{"cath_id":"1.20.1070.10","chopping":"28-318","consensus_level":"high","plddt":91.8456,"start":28,"end":318}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P35414","model_url":"https://alphafold.ebi.ac.uk/files/AF-P35414-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P35414-F1-predicted_aligned_error_v6.png","plddt_mean":81.69},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=APLNR","jax_strain_url":"https://www.jax.org/strain/search?query=APLNR"},"sequence":{"accession":"P35414","fasta_url":"https://rest.uniprot.org/uniprotkb/P35414.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P35414/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P35414"}},"corpus_meta":[{"pmid":"12603839","id":"PMC_12603839","title":"Pharmacological 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Overview.","date":"2021","source":"Cells","url":"https://pubmed.ncbi.nlm.nih.gov/35011661","citation_count":32,"is_preprint":false},{"pmid":"21412239","id":"PMC_21412239","title":"Non-activated APJ suppresses the angiotensin II type 1 receptor, whereas apelin-activated APJ acts conversely.","date":"2011","source":"Hypertension research : official journal of the Japanese Society of Hypertension","url":"https://pubmed.ncbi.nlm.nih.gov/21412239","citation_count":32,"is_preprint":false},{"pmid":"16211245","id":"PMC_16211245","title":"G protein-coupled APJ receptor signaling induces focal adhesion formation and cell motility.","date":"2005","source":"International journal of molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/16211245","citation_count":32,"is_preprint":false},{"pmid":"30380415","id":"PMC_30380415","title":"Apj+ Vessels Drive Tumor Growth and Represent a Tractable Therapeutic Target.","date":"2018","source":"Cell 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gastroenterology","url":"https://pubmed.ncbi.nlm.nih.gov/20725750","citation_count":30,"is_preprint":false},{"pmid":"33640616","id":"PMC_33640616","title":"Neuroprotective gain of Apelin/APJ system.","date":"2021","source":"Neuropeptides","url":"https://pubmed.ncbi.nlm.nih.gov/33640616","citation_count":29,"is_preprint":false},{"pmid":"30070701","id":"PMC_30070701","title":"Targeting drugs to APJ receptor: From signaling to pathophysiological effects.","date":"2018","source":"Journal of cellular physiology","url":"https://pubmed.ncbi.nlm.nih.gov/30070701","citation_count":29,"is_preprint":false},{"pmid":"33436040","id":"PMC_33436040","title":"Hsa_circ_0123190 acts as a competitive endogenous RNA to regulate APLNR expression by sponging hsa-miR-483-3p in lupus nephritis.","date":"2021","source":"Arthritis research & therapy","url":"https://pubmed.ncbi.nlm.nih.gov/33436040","citation_count":28,"is_preprint":false},{"pmid":"33504680","id":"PMC_33504680","title":"Apelin/APJ relieve diabetic cardiomyopathy by reducing microvascular dysfunction.","date":"2021","source":"The Journal of endocrinology","url":"https://pubmed.ncbi.nlm.nih.gov/33504680","citation_count":28,"is_preprint":false},{"pmid":"24966188","id":"PMC_24966188","title":"A static pressure sensitive receptor APJ promote H9c2 cardiomyocyte hypertrophy via PI3K-autophagy pathway.","date":"2014","source":"Acta biochimica et biophysica Sinica","url":"https://pubmed.ncbi.nlm.nih.gov/24966188","citation_count":28,"is_preprint":false},{"pmid":"36378421","id":"PMC_36378421","title":"Apelin/APJ system: an emerging therapeutic target for neurological diseases.","date":"2022","source":"Molecular biology reports","url":"https://pubmed.ncbi.nlm.nih.gov/36378421","citation_count":26,"is_preprint":false},{"pmid":"31408612","id":"PMC_31408612","title":"APLNR is involved in ATRA-induced growth inhibition of nasopharyngeal carcinoma and may suppress EMT through PI3K-Akt-mTOR signaling.","date":"2019","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/31408612","citation_count":26,"is_preprint":false},{"pmid":"20099713","id":"PMC_20099713","title":"Apelin and its receptor APJ in human aortic valve stenosis.","date":"2009","source":"The Journal of heart valve disease","url":"https://pubmed.ncbi.nlm.nih.gov/20099713","citation_count":26,"is_preprint":false},{"pmid":"37540330","id":"PMC_37540330","title":"ELABELA/APJ Axis Prevents Diabetic Glomerular Endothelial Injury by Regulating AMPK/NLRP3 Pathway.","date":"2023","source":"Inflammation","url":"https://pubmed.ncbi.nlm.nih.gov/37540330","citation_count":24,"is_preprint":false},{"pmid":"32194492","id":"PMC_32194492","title":"The Protective Effects and Mechanisms of Apelin/APJ System on Ischemic Stroke: A Promising Therapeutic Target.","date":"2020","source":"Frontiers in neurology","url":"https://pubmed.ncbi.nlm.nih.gov/32194492","citation_count":24,"is_preprint":false},{"pmid":"32066307","id":"PMC_32066307","title":"A patent review of apelin receptor (APJR) modulators (2014-2019).","date":"2020","source":"Expert opinion on therapeutic patents","url":"https://pubmed.ncbi.nlm.nih.gov/32066307","citation_count":23,"is_preprint":false},{"pmid":"33912466","id":"PMC_33912466","title":"Study Progression of Apelin/APJ Signaling and Apela in Different Types of Cancer.","date":"2021","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/33912466","citation_count":23,"is_preprint":false},{"pmid":"34174264","id":"PMC_34174264","title":"Apelin/APJ system: A novel therapeutic target for locomotor system diseases.","date":"2021","source":"European journal of pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/34174264","citation_count":23,"is_preprint":false},{"pmid":"35598689","id":"PMC_35598689","title":"The Apelin/APLNR system modulates tumor immune response by reshaping the tumor microenvironment.","date":"2022","source":"Gene","url":"https://pubmed.ncbi.nlm.nih.gov/35598689","citation_count":22,"is_preprint":false},{"pmid":"30626933","id":"PMC_30626933","title":"The biological function of ELABELA and APJ signaling in the cardiovascular system and pre-eclampsia.","date":"2019","source":"Hypertension research : official journal of the Japanese Society of Hypertension","url":"https://pubmed.ncbi.nlm.nih.gov/30626933","citation_count":20,"is_preprint":false},{"pmid":"34115895","id":"PMC_34115895","title":"Impact of the apelin/APJ axis in the pathogenesis of Parkinson's disease with therapeutic potential.","date":"2021","source":"Journal of neuroscience research","url":"https://pubmed.ncbi.nlm.nih.gov/34115895","citation_count":20,"is_preprint":false},{"pmid":"30631305","id":"PMC_30631305","title":"Characterization of the Apelin/Elabela Receptors (APLNR) in Chickens, Turtles, and Zebrafish: Identification of a Novel Apelin-Specific Receptor in Teleosts.","date":"2018","source":"Frontiers in endocrinology","url":"https://pubmed.ncbi.nlm.nih.gov/30631305","citation_count":19,"is_preprint":false},{"pmid":"37242650","id":"PMC_37242650","title":"APJ as Promising Therapeutic Target of Peptide Analogues in Myocardial Infarction- and Hypertension-Induced Heart Failure.","date":"2023","source":"Pharmaceutics","url":"https://pubmed.ncbi.nlm.nih.gov/37242650","citation_count":17,"is_preprint":false},{"pmid":"36813109","id":"PMC_36813109","title":"Elabela-APJ axis mediates angiogenesis via YAP/TAZ pathway in cerebral ischemia/reperfusion injury.","date":"2023","source":"Translational research : the journal of laboratory and clinical medicine","url":"https://pubmed.ncbi.nlm.nih.gov/36813109","citation_count":16,"is_preprint":false},{"pmid":"24411164","id":"PMC_24411164","title":"The effects of water immersion and restraint stress on the expressions of apelin, apelin receptor (APJR) and apoptosis rate in the rat heart.","date":"2014","source":"Acta histochemica","url":"https://pubmed.ncbi.nlm.nih.gov/24411164","citation_count":15,"is_preprint":false},{"pmid":"29395316","id":"PMC_29395316","title":"Apelin/APJ expression in the heart and kidneys of hypertensive rats.","date":"2018","source":"Acta histochemica","url":"https://pubmed.ncbi.nlm.nih.gov/29395316","citation_count":15,"is_preprint":false},{"pmid":"31645268","id":"PMC_31645268","title":"The influence of post-infarct heart failure and high fat diet on the expression of apelin APJ and vasopressin V1a and V1b receptors.","date":"2019","source":"Neuropeptides","url":"https://pubmed.ncbi.nlm.nih.gov/31645268","citation_count":15,"is_preprint":false},{"pmid":"31525345","id":"PMC_31525345","title":"Role of apelin/APJ system in hypothalamic-pituitary axis.","date":"2019","source":"Clinica chimica acta; international journal of clinical chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/31525345","citation_count":15,"is_preprint":false},{"pmid":"31538301","id":"PMC_31538301","title":"Apelin abrogates the stimulatory effects of 17β-estradiol and insulin-like growth factor-1 on proliferation of epithelial and granulosa ovarian cancer cell lines via crosstalk between APLNR and ERα/IGF1R.","date":"2019","source":"Molecular biology reports","url":"https://pubmed.ncbi.nlm.nih.gov/31538301","citation_count":15,"is_preprint":false},{"pmid":"32407761","id":"PMC_32407761","title":"Roles for heterodimerization of APJ and B2R in promoting cell proliferation via ERK1/2-eNOS signaling pathway.","date":"2020","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/32407761","citation_count":15,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":45492,"output_tokens":8405,"usd":0.131275,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":18217,"output_tokens":5046,"usd":0.108618,"stage2_stop_reason":"end_turn"},"total_usd":0.239893,"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\": 2012,\n      \"finding\": \"APJ acts as a dual-function receptor: apelin stimulation couples APJ to Gαi signaling and elicits a cardioprotective response, whereas mechanical stretch signals through APJ in a G-protein-independent, β-arrestin-dependent manner to induce cardiomyocyte hypertrophy. Knockdown of β-arrestins blocked stretch-mediated hypertrophy, and pharmacological doses of apelin acting through Gαi prevented it.\",\n      \"method\": \"Genetic loss-of-function (APJ-null and apelin-null mice), freshly isolated cardiomyocyte stretch assay, β-arrestin knockdown, Gαi pharmacology, cardiomyocyte size measurement\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal genetic and pharmacological approaches in one study, replicated across in vivo and in vitro models, published in Nature\",\n      \"pmids\": [\"22810587\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"ERG is a transcriptional activator of the APLNR gene and binds to the Aplnr promoter. Knockout of either Erg or Aplnr causes pulmonary venule-specific endothelial proliferation in vitro and pulmonary veno-occlusive disease in vivo, placing APLNR downstream of ERG in a pathway essential for venular endothelial homeostasis.\",\n      \"method\": \"Erg/Aplnr knockout mice, endothelium-directed conditional Aplnr deletion, in vitro proliferation assays, ChIP/transcriptional activation studies, patient lung tissue analysis\",\n      \"journal\": \"Circulation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic epistasis with both global and endothelium-specific knockouts, in vitro mechanistic follow-up, and human patient validation\",\n      \"pmids\": [\"25062690\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"APLNR-mediated APLN/APLNR signaling in endothelial cells induces transcription of miR-139-5p (promoted by laminar flow), which in turn suppresses endothelial CXCR4 expression. Loss of Apln, Aplnr, or endothelial-specific Aplnr caused dysregulated CXCR4 upregulation and retinal vascular defects; pharmacological inhibition of CXCR4 or augmentation of the miR-139-5p axis rescued these defects.\",\n      \"method\": \"Apln/Aplnr global and endothelial-specific knockout mice, retinal vascular phenotyping, miR-139-5p in vivo inhibition, pharmacological CXCR4 blockade\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple genetic knockouts (global and tissue-specific), pharmacological rescue experiments, and mechanistic miRNA pathway delineation in one study\",\n      \"pmids\": [\"27068353\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Endothelial APLNR signaling inactivates FOXO1 and suppresses endothelial expression of fatty acid binding protein 4 (FABP4). Conditional endothelial-specific Aplnr deletion impaired glucose utilization and abrogated apelin-induced glucose lowering; excess tissue fatty acid accumulation in skeletal muscle was the mechanistic link. Concurrent endothelial Foxo1 deletion or pharmacologic FABP4 inhibition rescued the metabolic phenotype.\",\n      \"method\": \"Endothelial-specific conditional Aplnr knockout mice, FOXO1 knockout epistasis, pharmacological FABP4 inhibition, metabolic and glucose utilization assays\",\n      \"journal\": \"Science translational medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional knockout with defined cellular phenotype, epistasis with Foxo1, pharmacological rescue, multiple orthogonal metabolic readouts in one study\",\n      \"pmids\": [\"28904225\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Apelin peptides are endogenous ligands for the human APJ (APLNR) receptor. Structure-activity studies identified that leucine at position 5 and arginine at positions 2 and 4 of apelin-13 are key residues required for functional potency and binding affinity at the recombinant human APJ receptor.\",\n      \"method\": \"Radioligand binding assays, functional assays with apelin analogues at human recombinant APJ receptor, quantitative RT-PCR, immunohistochemistry\",\n      \"journal\": \"Journal of neurochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro receptor binding and functional assays with systematic mutagenesis of the ligand, replicated across multiple analogue series\",\n      \"pmids\": [\"12603839\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"APJ receptor undergoes rapid, dose-dependent ligand-induced internalization upon stimulation with Apelin-36 and Apelin-13 via clathrin-coated pits (co-localization with transferrin receptor). The intact cytoplasmic C-terminal domain of APJ is required for ligand-induced internalization. Internalized APJ recycled to the cell surface within 60 min after Apelin-13 removal, but most internalized receptor remained in the cytoplasm 2 h after Apelin-36 washout.\",\n      \"method\": \"APJ-GFP stable expression, fluorescent apelin peptide (5-CF-Apelin-13), co-localization with transferrin receptor, C-terminal truncation mutants, live cell imaging\",\n      \"journal\": \"Virology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — reconstituted receptor internalization in multiple cell types with mutagenesis (C-terminal domain) and co-localization with established clathrin pathway marker\",\n      \"pmids\": [\"12667811\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Apelin/APJ signaling via a pertussis toxin-sensitive Gi protein activates PI3K→Akt/PKB and focal adhesion kinase (FAK) phosphorylation, increases focal adhesion formation with actin reorganization, and stimulates cell motility in APJ-expressing cells.\",\n      \"method\": \"Stable APJ-expressing HEK293T cells, radioligand binding, pertussis toxin treatment, PI3K inhibitor (LY294002), phospho-Akt/FAK immunoblotting, scratch migration assay, F-actin staining\",\n      \"journal\": \"International journal of molecular medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with defined receptor, multiple pharmacological inhibitors blocking defined signaling nodes, and orthogonal functional readouts (phosphorylation, morphology, migration)\",\n      \"pmids\": [\"16211245\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"APJ functions as a coreceptor for primate immunodeficiency viruses (HIV-1 and SIV). APJ coreceptor activity is demonstrable even at low expression levels, and the endogenous peptide ligand apelin-13 efficiently blocked APJ coreceptor activity. APJ is N-glycosylated as determined by Western blot with a specific monoclonal antibody.\",\n      \"method\": \"Monoclonal antibody (MAb 856) production, FACS, Western blot, immunofluorescence, cell-cell fusion assays, apelin-13 blockade of coreceptor activity\",\n      \"journal\": \"Virology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional coreceptor assays with apelin-13 blockade, glycosylation determination by Western blot, multiple detection methods in one study\",\n      \"pmids\": [\"11040134\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Non-activated (ligand-free) APJ suppresses angiotensin II (AngII)/AT1-mediated ERK1/2 phosphorylation in a receptor-proximity-dependent manner independent of heterodimerization, whereas apelin-activated APJ reverses this suppression through Gαi (pertussis toxin-sensitive). Thus APJ has a constitutive, ligand-independent inhibitory interaction with AT1 signaling that is abolished upon apelin binding.\",\n      \"method\": \"HEK293 cell co-expression of APJ and AT1, pertussis toxin treatment, AT2 and β2-adrenergic receptor controls, ERK1/2 phosphorylation immunoblotting, AT1 blocker dose-response\",\n      \"journal\": \"Hypertension research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — defined signaling mechanism with pharmacological controls, but single lab and single method (ERK1/2 phosphorylation) as primary readout\",\n      \"pmids\": [\"21412239\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Apelin/APLNR signaling promotes endothelial cell (EC) proliferation via PI3K/Akt-mediated activation, and apelin-12 activates NO production via the PI3K/Akt pathway in human ECs. Apelin-13 additionally activates Erk1/2 phosphorylation and enhances EC proliferation. APJ knockdown inhibited PI3K phosphorylation and impaired flow-induced eNOS and PECAM-1 expression; APJ expression is induced by shear stress independently of its ligand.\",\n      \"method\": \"siRNA knockdown of APJ or apelin in human ECs, shear stress flow chamber, NO measurement, PI3K/Akt and ERK1/2 phosphorylation immunoblotting, gene and protein expression under flow\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — defined signaling nodes with siRNA knockdown and pharmacological context, single lab, several orthogonal readouts\",\n      \"pmids\": [\"25817266\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"APJ receptor acts as a static pressure sensor in cardiomyocytes. Static pressure upregulates APJ expression and activates PI3K/Akt/autophagy (LC3-II/I, Beclin-1) signaling to promote cardiomyocyte hypertrophy. APJ shRNA, PI3K inhibitor (LY294002), Akt inhibitor, and autophagy inhibitor (3-methyladenine) each reversed pressure-induced increases in cell diameter, volume, and protein content.\",\n      \"method\": \"Rat left ventricular hypertrophy model (aortic coarctation), H9c2 cardiomyocyte static pressure culture, APJ shRNA knockdown, PI3K/Akt inhibitors, LC3/Beclin-1 immunoblotting, cell size measurements\",\n      \"journal\": \"Acta biochimica et biophysica Sinica\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — in vivo and in vitro models with shRNA and pharmacological inhibitors, single lab, defined pathway but no rescue experiments\",\n      \"pmids\": [\"24966188\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Hypoxia-induced HIF-1α upregulates apelin and APLNR expression in bone marrow-derived mesenchymal stem cells (BMSCs), and this apelin/APJ/autophagy axis mediates hypoxia-induced BMSC proliferation. siRNA-HIF-1α suppressed apelin, APJ, Beclin-1, and LC3II/LC3I; siRNA-APJ suppressed Beclin-1 and LC3II/LC3I and reversed proliferation; siRNA-Beclin-1 abolished proliferation.\",\n      \"method\": \"Mouse BMSC hypoxia culture, siRNA knockdown of HIF-1α/APJ/Beclin-1, MTT and BrdU proliferation assays, Western blot for autophagy markers\",\n      \"journal\": \"Acta biochimica et biophysica Sinica\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — genetic epistasis via sequential siRNA knockdown establishing HIF-1α→apelin/APJ→autophagy→proliferation pathway, single lab\",\n      \"pmids\": [\"25736405\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Hypoxia upregulates HIF-1α, Apelin, and APLNR in endothelial progenitor cells (EPCs), and the Apelin/APLNR axis mediates hypoxia-induced EPC proliferation via downstream MAPK signaling. siRNA knockdown of Apelin or APLNR suppressed hypoxia-induced proliferation; MAPK inhibitors (SB-239063 and PD98059) eliminated Apelin upregulation-induced EPC proliferation.\",\n      \"method\": \"Human EPC hypoxia culture, siRNA knockdown of Apelin/APLNR, MAPK inhibitors, MTT proliferation assay, RT-qPCR and Western blot\",\n      \"journal\": \"Molecular medicine reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — siRNA knockdown and pharmacological inhibition establishing signaling pathway, single lab\",\n      \"pmids\": [\"26676468\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"APLNR (Aplnr) is specifically expressed on circulating cKit+/Flk1+ cells and functions as a receptor for apelin-mediated chemoattraction during myocardial repair. Apelin injection into ischemic myocardium increased recruitment of cKit+/Flk1+/Aplnr+ cells; Aplnr knockdown in bone marrow aggravated ischemic damage and could not be rescued by apelin. Recruited cells promoted neovascularization via paracrine rather than transdifferentiation mechanisms.\",\n      \"method\": \"Bone marrow Aplnr knockdown, apelin injection into ischemic myocardium, flow cytometry for circulating progenitor populations, scar formation and cardiac function measurement\",\n      \"journal\": \"Circulation research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — in vivo receptor knockdown with defined progenitor cell population and functional cardiac readouts, single lab\",\n      \"pmids\": [\"22753078\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"APLN (apelin) acts through APLNR to increase steroidogenesis in human luteinized granulosa cells (hGCs), enhancing both basal and IGF1-induced steroid secretion by increasing HSD3B protein concentration through activation of the MAPK3/1 (ERK1/2) and AKT pathways. The APLNR antagonist ML221 reversed these effects. IGF1 increased APLNR expression in hGCs.\",\n      \"method\": \"Cultured human luteinized granulosa cells, recombinant apelin-13 and apelin-17 treatment, ML221 APLNR antagonist, pharmacological inhibitors of AKT and MAPK3/1 pathways, Western blot, steroid assays\",\n      \"journal\": \"Biology of reproduction\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — receptor antagonist and pathway inhibitors with defined steroidogenic readout, single lab\",\n      \"pmids\": [\"27683264\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"In Sertoli cells, high glucose induces local APLN production, which via APJ hyperactivation suppresses carnitine production and represses cell adhesion gene expression, leading to blood-testis barrier (BTB) structural dysfunction and impaired spermatogenesis in diabetic models. Pharmacological blockade of APJ with ML221 ameliorated BTB damage and improved spermatogenesis in diabetic db/db mice and cultured human testes.\",\n      \"method\": \"Single-cell RNA sequencing of diabetic patient testes (STRT-seq), high glucose treatment of Sertoli cells, APJ antagonist (ML221) treatment in db/db mice and human testis culture, BTB structural assessment\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — scRNA-seq plus pharmacological rescue in both mouse and human models, defined mechanistic pathway (APLN→APJ→carnitine/adhesion→BTB dysfunction), replicated across species\",\n      \"pmids\": [\"36443325\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"In glioblastoma, APLNR on tumor cells mediates apelin-induced migration and invasion. Apelin reduction led to accelerated tumor cell invasion by APLNR-positive cells. Mutant APLNR ligand apelin-F13A blocked both tumor angiogenesis and GBM cell invasion, and cotargeting VEGFR2 and APLNR synergistically improved survival in proneural GBM mouse models.\",\n      \"method\": \"APLN knockdown/knockout in orthotopic GBM models, apelin-F13A peptide treatment, VEGFR2 + APLNR cotargeting, stereotactic biopsy analysis, in vitro and in vivo invasion assays\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic knockdown/knockout with in vivo rescue experiments and pharmacological ligand, two orthogonal readouts (angiogenesis and invasion), single lab\",\n      \"pmids\": [\"30358318\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"APJ expression in ovarian cancer cells is necessary and sufficient for pro-metastatic phenotypes (proliferation, adhesion, anoikis resistance, migration, invasion) via downstream activation of STAT3, ERK, and AKT pathways. APJ inhibitor ML221 efficiently inhibited these phenotypes in vitro, and APJ overexpression increased metastasis in vivo.\",\n      \"method\": \"APJ overexpression and knockdown in ovarian cancer cell lines, in vitro adhesion/migration/invasion assays, anoikis assay, in vivo metastasis model, STAT3/ERK/AKT phosphorylation immunoblotting, ML221 inhibitor treatment\",\n      \"journal\": \"Molecular cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — gain- and loss-of-function in vitro plus in vivo validation with defined pathway components, single lab\",\n      \"pmids\": [\"30858172\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"APJ and bradykinin B2 receptor (B2R) form a functional heterodimer at the cell membrane, demonstrated by BRET, FRET, proximity ligation assay, and co-immunoprecipitation in HUVECs and transfected cells. Stimulation with apelin-13 and bradykinin increased eNOS phosphorylation via the APJ-B2R heterodimer through a PLC/ERK1/2/eNOS signaling pathway, leading to increased cell proliferation. Silencing of either APJ or B2R inhibited eNOS phosphorylation.\",\n      \"method\": \"BRET and FRET resonance energy transfer, proximity ligation assay, co-immunoprecipitation in HUVECs and HEK293 cells, siRNA knockdown of APJ or B2R, ERK1/2 and eNOS phosphorylation assays, cell proliferation assay\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — heterodimer formation confirmed by multiple biophysical methods (BRET, FRET, PLA, Co-IP) plus functional signaling validation with siRNA knockdown, single lab\",\n      \"pmids\": [\"32407761\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ELA (Elabela) binds to APJ and activates the NRF2/ARE antioxidative signaling pathway via Gα13, thereby reducing neuronal ferroptosis after cerebral ischemia/reperfusion injury. AAV-mediated APJ knockdown or the NRF2 inhibitor ML385 abolished the protective effects of ELA-32.\",\n      \"method\": \"ELA-32 peptide treatment in cerebral I/R mouse model, AAV-APJ-RNAi knockdown, NRF2 inhibitor ML385, iron deposition, lipid peroxidation, mitochondrial morphology assays, behavioral readouts\",\n      \"journal\": \"Free radical biology & medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — defined signaling axis (APJ→Gα13→NRF2/ARE) with genetic (AAV knockdown) and pharmacological (NRF2 inhibitor) validation, single lab\",\n      \"pmids\": [\"36681202\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ELA (Elabela) binds to APJ in renal tubular cells to regulate renal microvascular blood flow through two downstream mediators: arginine metabolizing enzyme ARG2 and PGE2 metabolizing enzymes CBR1/3. APJ inhibitor ML221 blocked the beneficial effects of exogenous ELA-32 on AKI, while combination treatment with ARG2 inhibitor nor-NOHA and PGE2 activator Paricalcitol alleviated injury independently of APJ, placing ARG2 and CBR1/3 downstream of the ELA-APJ axis.\",\n      \"method\": \"Renal tubule-specific Apela (ELA) knockout mice, bilateral/unilateral I/R models, RNA sequencing, ML221 APJ inhibitor, ARG2 inhibitor (nor-NOHA) and Paricalcitol combination treatment, renal blood flow and functional measurements\",\n      \"journal\": \"Theranostics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — tissue-specific knockout plus RNA-seq pathway discovery plus pharmacological rescue, multiple orthogonal approaches, single lab\",\n      \"pmids\": [\"37351176\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"The murine msr/apj receptor (ortholog of human APJ/APLNR) is expressed in endothelium of primary blood vessels and the forming heart, as well as in somites, limb bud, and branchial arches during embryonic development, indicating a role in endothelial/vascular lineage specification distinct from Flk1.\",\n      \"method\": \"Molecular cloning, in situ hybridization in developing mouse embryo, comparative expression with Flk1\",\n      \"journal\": \"Mechanisms of development\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — direct localization by in situ hybridization in developing tissues, replicated across developmental stages, but no functional consequence tested\",\n      \"pmids\": [\"10473142\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"APLNR (APJ receptor) mRNA is upregulated in the hypothalamic parvocellular paraventricular nucleus (pPVN) by acute and repeated restraint stress, and adrenalectomy also increased APJR mRNA in the PVN. Adrenalectomized rats showed no further increase above baseline after stress, indicating glucocorticoids negatively regulate APJR mRNA expression and mediate stress-induced regulation.\",\n      \"method\": \"In situ hybridization for APJR mRNA in rat hypothalamus, restraint stress paradigms, adrenalectomy, dual-label in situ hybridization to co-localize APJR and vasopressin mRNA\",\n      \"journal\": \"Journal of neuroendocrinology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — defined neuroendocrine regulation of APLNR expression by glucocorticoids established via adrenalectomy and stress models, single lab\",\n      \"pmids\": [\"14622440\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"APLNR (APJR) mRNA is co-expressed with vasopressin in magnocellular neurons of the hypothalamic PVN and SON, and its expression is induced by osmotic stimuli (2% NaCl loading and water deprivation). Salt-loading increased co-localization of APJR and vasopressin mRNAs in the SON, supporting a role for APJ in the autocrine/paracrine regulation of vasopressin-containing neurons and fluid homeostasis.\",\n      \"method\": \"In situ hybridization histochemistry for APJR mRNA, dual-label in situ hybridization for APJR and vasopressin in salt-loaded and water-deprived rats\",\n      \"journal\": \"Journal of neuroendocrinology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — direct localization with functional osmotic stimulus paradigm and dual-label co-localization, replicated across osmotic challenge conditions, single lab\",\n      \"pmids\": [\"12787050\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"APJ activation by apelin improves AngII-induced endothelial cell senescence via the AMPK/SIRT1 signaling pathway. APJ, AMPK, or SIRT1 knockdown each attenuated the protective effects of apelin. Apelin reduced AngII-induced ROS generation and enhanced telomerase activity in HUVECs.\",\n      \"method\": \"AngII-induced HUVEC senescence model, SA-β-Gal assay, siRNA knockdown of APJ/AMPK/SIRT1, ROS detection, telomerase activity (RQ-TRAP), CCK-8 viability assay, Western blot for P21 and PAI-1\",\n      \"journal\": \"Archives of medical science\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, endpoint-driven assays with siRNA knockdown but limited mechanistic depth beyond pathway identification\",\n      \"pmids\": [\"30002688\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"APLNR overexpression in nasopharyngeal carcinoma (NPC) cells inhibited migration, invasion, and EMT. Low APLNR expression activated the PI3K-AKT-mTOR signaling pathway to promote EMT. ATRA treatment upregulated APLNR in NPC cell lines and inhibited proliferation; knockdown of APLNR diminished ATRA-induced growth inhibition.\",\n      \"method\": \"APLNR overexpression and knockdown in NPC cell lines, wound-healing and Transwell migration/invasion assays, Western blot for EMT markers and PI3K-AKT-mTOR pathway components, ATRA treatment, cell cycle analysis\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — gain- and loss-of-function with defined EMT pathway, ATRA epistasis, single lab\",\n      \"pmids\": [\"31408612\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Adipocyte-derived apelin activates APJ on ovarian cancer cells in a paracrine manner, promoting lipid uptake via CD36 upregulation through APJ-STAT3 activation, and the accumulated lipids are utilized for fatty acid oxidation via AMPK-CPT1a axis. APJ antagonist F13A or APJ knockdown reversed lipid accumulation, migration, invasion, and omental homing in vivo.\",\n      \"method\": \"Co-culture with 3T3-L1 adipocyte conditioned media, APJ antagonist F13A, APJ knockdown, in vitro migration/invasion assays, ex vivo omentum adhesion, in vivo homing assay, CD36 and STAT3 pathway analysis, lipid droplet staining\",\n      \"journal\": \"Molecular cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — paracrine signaling mechanism defined with receptor knockdown, antagonist, and in vivo validation; multiple orthogonal readouts, single lab\",\n      \"pmids\": [\"34172534\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"APLNR abrogates the stimulatory effects of 17β-estradiol on OVCAR-3 epithelial ovarian cancer cell proliferation and of IGF-1 on COV434 granulosa cancer cell proliferation via crosstalk between APLNR and estrogen receptor alpha (ERα) or IGF-1 receptor (IGF1R), respectively, independently of ERK1/2 and PI3K pathways.\",\n      \"method\": \"OVCAR-3 and COV434 cell proliferation assays, apelin treatment, ERK1/2 and PI3K pathway inhibitors, estrogen and IGF-1 stimulation, APLNR-ERα/IGF1R crosstalk analysis\",\n      \"journal\": \"Molecular biology reports\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, limited mechanistic detail on the receptor crosstalk mechanism beyond pharmacological inhibitor results\",\n      \"pmids\": [\"31538301\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ELA (Elabela) binds APJ on brain endothelial cells and activates the YAP/TAZ signaling pathway, promoting post-ischemic cerebral angiogenesis. Silencing of APJ or pharmacological blockade of YAP abolished the pro-angiogenic effects of ELA-32 in oxygen-glucose deprivation/reoxygenation conditions.\",\n      \"method\": \"ELA-32 treatment of bEnd.3 cells under OGD/R, APJ siRNA knockdown, YAP pharmacological blockade, RNA sequencing, in vivo cerebral I/R model with CBF measurement\",\n      \"journal\": \"Translational research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — mechanistic signaling defined (APJ→YAP/TAZ) with genetic and pharmacological inhibition, RNA-seq pathway analysis, single lab\",\n      \"pmids\": [\"36813109\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"APLNR (APJ) is a class A GPCR that functions as a bifunctional receptor: its canonical endogenous ligands apelin (and Elabela) couple it primarily to Gαi to drive cardioprotective, vasodilatory, pro-angiogenic, and metabolic effects via downstream PI3K/AKT, ERK1/2, AMPK, and NRF2/ARE signaling, while mechanical stretch signals through APJ in a G-protein-independent, β-arrestin-dependent manner to induce cardiomyocyte hypertrophy; ligand binding induces rapid clathrin-mediated receptor internalization requiring the intact C-terminal domain; in endothelial cells, APJ signaling induces miR-139-5p to suppress CXCR4, inactivates FOXO1 to suppress FABP4 expression and regulate fatty acid uptake, and is transcriptionally activated by the ERG transcription factor to maintain venular homeostasis; in Sertoli cells, hyperactivated APJ suppresses carnitine production and cell adhesion to impair the blood-testis barrier; non-activated APJ constitutively suppresses AT1 receptor signaling while apelin-activated APJ relieves this suppression; and APJ can heterodimerize with B2R to activate PLC/ERK1/2/eNOS-dependent cell proliferation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"APLNR (APJ) is a class A G-protein-coupled receptor that integrates peptide-ligand and mechanical inputs to control vascular, cardiac, metabolic, and reproductive physiology [#0, #4]. Its endogenous apelin peptides bind the human receptor with potency dictated by defined residues of apelin-13 [#4], coupling APJ predominantly to pertussis-toxin-sensitive G\\u03b1i to activate PI3K\\u2192Akt and FAK signaling that drives focal adhesion formation, actin reorganization, and cell motility [#6]; the second peptide ligand Elabela engages APJ to activate G\\u03b113-dependent NRF2/ARE antioxidative signaling and YAP/TAZ-dependent angiogenesis [#19, #28]. APJ is bifunctional: apelin/G\\u03b1i signaling is cardioprotective, whereas mechanical stretch or static pressure signals through APJ in a G-protein-independent, \\u03b2-arrestin-dependent manner to drive cardiomyocyte hypertrophy [#0, #10]. Ligand binding triggers rapid clathrin-mediated internalization that requires the intact cytoplasmic C-terminal domain, with apelin-13-bound receptor recycling to the surface [#5]. In the endothelium APJ acts downstream of the ERG transcription factor to maintain venular homeostasis [#1] and orchestrates a miR-139-5p\\u2192CXCR4 axis controlling developmental vascular patterning [#2] and a FOXO1\\u2192FABP4 axis governing endothelial fatty-acid handling and systemic glucose utilization [#3]. APJ further modulates receptor crosstalk, constitutively suppressing AngII/AT1-mediated ERK1/2 signaling in the ligand-free state and relieving this suppression upon apelin binding [#8], and forming a functional heterodimer with the bradykinin B2 receptor that drives PLC/ERK1/2/eNOS-dependent endothelial proliferation [#18]. In disease, APJ hyperactivation by locally produced apelin suppresses carnitine production and adhesion gene expression in Sertoli cells to disrupt the blood-testis barrier in diabetes [#15], and APJ drives pro-metastatic and pro-invasive programs in ovarian cancer and glioblastoma via STAT3, ERK, and AKT signaling [#16, #17].\",\n  \"teleology\": [\n    {\n      \"year\": 2000,\n      \"claim\": \"Established that APJ is a functional cell-surface receptor with biological activity beyond an orphan GPCR by showing it serves as a primate immunodeficiency virus coreceptor blockable by its peptide ligand.\",\n      \"evidence\": \"Cell-cell fusion and coreceptor assays with apelin-13 blockade and Western blot glycosylation analysis\",\n      \"pmids\": [\"11040134\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not define physiological signaling output\", \"Coreceptor role distinct from native ligand signaling\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Defined the molecular determinants of apelin binding to human APJ, establishing the ligand pharmacophore.\",\n      \"evidence\": \"Radioligand binding and functional assays with systematic apelin-13 analogue mutagenesis at recombinant human APJ\",\n      \"pmids\": [\"12603839\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No receptor structural model\", \"Downstream signaling not resolved in this study\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Answered how APJ signaling is terminated and reset by demonstrating C-terminal-dependent clathrin-mediated internalization and ligand-specific recycling.\",\n      \"evidence\": \"APJ-GFP live-cell imaging, fluorescent apelin, transferrin receptor co-localization, and C-terminal truncation mutants\",\n      \"pmids\": [\"12667811\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Specific C-terminal motifs/phosphosites not mapped\", \"Arrestin involvement in internalization not directly tested\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Identified the proximal G-protein-coupled signaling cascade, linking APJ to Gi-PI3K/Akt-FAK and cell motility.\",\n      \"evidence\": \"Stable APJ-HEK293T cells with pertussis toxin, PI3K inhibitor, phospho-immunoblotting, and migration assays\",\n      \"pmids\": [\"16211245\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Cell-type generality untested\", \"Does not address mechanical/biased signaling\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Revealed that ligand-free APJ has constitutive activity by suppressing AT1/AngII ERK1/2 signaling that apelin binding relieves.\",\n      \"evidence\": \"HEK293 APJ/AT1 co-expression, pertussis toxin, receptor controls, ERK1/2 immunoblotting\",\n      \"pmids\": [\"21412239\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single ERK1/2 readout from one lab\", \"Physiological/in vivo relevance untested\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Defined APJ as a bifunctional receptor distinguishing protective apelin/G\\u03b1i signaling from pathological stretch-induced \\u03b2-arrestin-dependent hypertrophy.\",\n      \"evidence\": \"APJ-null and apelin-null mice, isolated cardiomyocyte stretch assay, \\u03b2-arrestin knockdown, G\\u03b1i pharmacology\",\n      \"pmids\": [\"22810587\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of biased mechanosensing unknown\", \"Arrestin effector identity in hypertrophy unresolved\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Showed APJ functions as an apelin chemoattractant receptor on circulating progenitor cells supporting myocardial repair.\",\n      \"evidence\": \"Bone marrow Aplnr knockdown, apelin injection into ischemic myocardium, flow cytometry, cardiac function readouts\",\n      \"pmids\": [\"22753078\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab\", \"Paracrine effectors not identified\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Placed APLNR in a transcriptional hierarchy by identifying ERG as a direct activator required for venular endothelial homeostasis.\",\n      \"evidence\": \"Erg/Aplnr knockout mice, endothelium-specific deletion, ChIP/promoter binding, human lung tissue\",\n      \"pmids\": [\"25062690\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Downstream effectors of APLNR in venular ECs not fully defined here\", \"Ligand dependence of phenotype unclear\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Extended the mechanosensor role to static pressure, linking APJ to PI3K/Akt/autophagy-driven cardiomyocyte hypertrophy.\",\n      \"evidence\": \"Rat hypertrophy model, H9c2 static pressure culture, APJ shRNA, PI3K/Akt/autophagy inhibitors\",\n      \"pmids\": [\"24966188\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No rescue experiments\", \"Relationship to \\u03b2-arrestin stretch pathway not reconciled\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Identified a HIF-1\\u03b1\\u2192apelin/APJ\\u2192autophagy axis driving hypoxia-induced proliferation in stem and progenitor cells.\",\n      \"evidence\": \"BMSC and EPC hypoxia cultures, sequential siRNA knockdown of HIF-1\\u03b1/APJ/Beclin-1, MAPK inhibitors, proliferation assays\",\n      \"pmids\": [\"25736405\", \"26676468\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab per cell type\", \"Direct receptor-effector coupling not biochemically resolved\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Defined an endothelial APJ\\u2192miR-139-5p\\u2192CXCR4 axis controlling vascular patterning, induced by laminar flow.\",\n      \"evidence\": \"Apln/Aplnr global and endothelial-specific knockouts, retinal phenotyping, miR-139-5p inhibition, CXCR4 blockade\",\n      \"pmids\": [\"27068353\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism of miR-139-5p induction by APJ unresolved\", \"Flow-sensing versus ligand contribution not separated\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Connected APJ to ligand-independent shear-stress responses and NO/proliferation signaling in endothelium.\",\n      \"evidence\": \"siRNA knockdown in human ECs, shear-stress flow chamber, NO measurement, PI3K/Akt/ERK1/2 immunoblotting\",\n      \"pmids\": [\"25817266\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab\", \"Mechanism of shear-induced APJ upregulation unknown\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Established APJ as a regulator of systemic metabolism through an endothelial FOXO1\\u2192FABP4 fatty-acid handling axis.\",\n      \"evidence\": \"Endothelial-specific Aplnr knockout, Foxo1 epistasis, FABP4 inhibition, glucose utilization assays\",\n      \"pmids\": [\"28904225\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct biochemical link from APJ to FOXO1 inactivation not mapped\", \"G-protein dependence not dissected\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Demonstrated APJ drives pro-metastatic and invasive tumor programs via STAT3/ERK/AKT in ovarian cancer and glioblastoma.\",\n      \"evidence\": \"Gain/loss-of-function in tumor cells, in vivo metastasis/invasion models, ML221 and apelin-F13A antagonists, VEGFR2 cotargeting\",\n      \"pmids\": [\"30858172\", \"30358318\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab per tumor type\", \"Context-dependent tumor-suppressive role elsewhere unexplained\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Showed APJ engages in receptor crosstalk by heterodimerizing with the bradykinin B2 receptor to drive eNOS-dependent endothelial proliferation.\",\n      \"evidence\": \"BRET, FRET, proximity ligation, co-IP in HUVECs/HEK293, siRNA knockdown, ERK1/2/eNOS phosphorylation\",\n      \"pmids\": [\"32407761\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Stoichiometry and structural interface unknown\", \"In vivo relevance of heterodimer untested\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Defined a pathological APJ hyperactivation mechanism disrupting the blood-testis barrier in diabetes via carnitine and adhesion suppression.\",\n      \"evidence\": \"scRNA-seq of diabetic testes, high-glucose Sertoli cell treatment, ML221 rescue in db/db mice and human testis culture\",\n      \"pmids\": [\"36443325\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Signaling node linking APJ to carnitine repression unmapped\", \"Receptor coupling (G-protein vs arrestin) not defined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Established Elabela as a second APJ ligand engaging distinct G\\u03b113/NRF2 and YAP/TAZ pathways in protective cytoprotection and angiogenesis.\",\n      \"evidence\": \"ELA-32 treatment in cerebral and renal I/R models, AAV/siRNA APJ knockdown, NRF2 and YAP inhibitors, RNA-seq\",\n      \"pmids\": [\"36681202\", \"36813109\", \"37351176\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Ligand bias between apelin and Elabela not structurally defined\", \"Single lab per organ system\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How APJ structurally encodes biased signaling among G\\u03b1i, G\\u03b113, \\u03b2-arrestin, and mechanical inputs, and how distinct ligands and heterodimers select these outputs, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No receptor structure or biased-agonism model in the corpus\", \"Mechanism of mechanosensing unknown\", \"Determinants of apelin vs Elabela pathway selection undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060089\", \"supporting_discovery_ids\": [0, 4, 6]},\n      {\"term_id\": \"GO:0048018\", \"supporting_discovery_ids\": [4]},\n      {\"term_id\": \"GO:0001618\", \"supporting_discovery_ids\": [7]},\n      {\"term_id\": \"GO:0120274\", \"supporting_discovery_ids\": [7]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [5, 7, 18]},\n      {\"term_id\": \"GO:0031410\", \"supporting_discovery_ids\": [5]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0, 6, 8]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [1, 2, 21]},\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [3, 26]},\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [10, 11]}\n    ],\n    \"complexes\": [\"APJ-B2R heterodimer\"],\n    \"partners\": [\"APLN\", \"APELA\", \"BDKRB2\", \"AGTR1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":{"gene":"APLNR","tier":"GROUNDING","verdict":"Evidence-grounding concern","subtype":"fabrication","uniprot_band":"rich","rules_fired":"R7","issue":"R7: fabricated (no corpus paper): 30358318"},"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}