{"gene":"NOX4","run_date":"2026-06-10T05:19:52","timeline":{"discoveries":[{"year":2001,"finding":"NOX4 encodes a ~65 kDa protein homologous to gp91phox (the catalytic subunit of the phagocyte NADPH oxidase), containing 5-6 conserved predicted transmembrane alpha-helices with putative heme-binding regions plus a flavoprotein homology domain with predicted FAD and NADPH binding sites, establishing it as an NADPH-dependent ROS-generating enzyme.","method":"cDNA cloning, sequence analysis, tissue expression (Northern blot/RT-PCR)","journal":"Gene","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — structural inference from sequence homology confirmed across multiple tissues; functional enzymatic activity not yet directly reconstituted in this paper but domain architecture well established","pmids":["11376945"],"is_preprint":false},{"year":2003,"finding":"Nox4-based NADPH oxidase, regulated upstream by Rac1 and arachidonic acid, mediates angiotensin II-induced ROS generation and downstream Akt/PKB activation and protein synthesis in mesangial cells. Antisense Nox4 knockdown abolished ANG II-induced NADPH oxidase activity and Akt/PKB activation, and dominant-negative Rac1 blocked Nox4-dependent ROS.","method":"Antisense oligonucleotide knockdown, dominant-negative Rac1 transfection, NADPH oxidase activity assay, ROS measurement, Akt phosphorylation assays, protein synthesis assay","journal":"American journal of physiology. Renal physiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods (AS knockdown, DN-Rac1, functional ROS/kinase assays) in single lab","pmids":["12842860"],"is_preprint":false},{"year":2005,"finding":"Nox4 is the major source of NADPH-dependent ROS in the diabetic kidney; antisense-mediated Nox4 knockdown reduced NADPH oxidase activity in renal cortical/glomerular homogenates, blocked glucose-induced ROS in isolated glomeruli, and reduced downstream Akt/PKB and ERK1/2 activation, renal hypertrophy, and fibronectin expression.","method":"Antisense oligonucleotide administration in vivo (osmotic minipump), NADPH oxidase activity assay, ROS measurement from intact glomeruli, immunoblotting for Akt/ERK, histology for hypertrophy, fibronectin immunostaining","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods in vivo and in vitro, single lab","pmids":["16135519"],"is_preprint":false},{"year":2005,"finding":"NOX4-GFP fusion protein localizes to the endoplasmic reticulum in human endothelial cells (HUVECs), as shown by co-staining with an ER marker; distribution did not overlap with lysosomes, Weibel-Palade bodies, or mitochondria.","method":"Fluorescence confocal microscopy with ER, lysosomal, and mitochondrial markers; NOX4-GFP overexpression","journal":"Antioxidants & redox signaling","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — direct localization by confocal with multiple organelle markers, single lab","pmids":["15706079"],"is_preprint":false},{"year":2006,"finding":"NOX2 and NOX4 co-localize with the ER marker calreticulin in endothelial cells and interact with p22phox, as demonstrated by bimolecular fluorescence complementation; both NOX2 and NOX4 contribute equally to endothelial ROS production and proliferation.","method":"Bimolecular fluorescence complementation (BiFC) for NOX4-p22phox interaction, co-localization with calreticulin by immunofluorescence, siRNA knockdown, ROS measurement","journal":"Antioxidants & redox signaling","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — protein–protein interaction confirmed by BiFC plus functional ROS/proliferation readouts, single lab","pmids":["16987004"],"is_preprint":false},{"year":2008,"finding":"Nox4 controls the switch between insulin-induced proliferation and differentiation in preadipocytes by regulating MAP kinase phosphatase-1 (MKP-1) expression; Nox4 siRNA reduced ROS and MKP-1, de-repressed ERK1/2, which phosphorylated IRS-1 at Ser612 to block differentiation and promote proliferation.","method":"siRNA knockdown, Nox4 overexpression, ERK1/2 phosphorylation assays, MKP-1 expression analysis, IRS-1 phosphorylation, proliferation/differentiation assays","journal":"Arteriosclerosis, thrombosis, and vascular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal siRNA/overexpression with mechanistic pathway dissection, single lab","pmids":["19057021"],"is_preprint":false},{"year":2009,"finding":"Nox4 localizes to mitochondria in mesangial cells and kidney cortex; siRNA-mediated knockdown of Nox4 significantly reduces NADPH oxidase activity in purified mitochondria and blocks glucose-induced mitochondrial superoxide generation. Mitochondrial Nox4 expression is increased in diabetic kidney cortex.","method":"Subcellular fractionation, immunoblotting of mitochondrial fractions, immunofluorescence confocal with Mitotracker, MitoProt prediction, siRNA knockdown, NADPH oxidase activity assay in purified mitochondria, in vivo diabetic rat model","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal localization methods (fractionation, confocal, computational) plus functional knockdown in purified organelle, single lab","pmids":["19706525"],"is_preprint":false},{"year":2009,"finding":"Structural elements in Nox4 determine both its subcellular localization to the ER and the type of ROS released (H2O2 extracellularly rather than O2-). The cytosolic tail of Nox4 confers constitutive activity (independent of cytosolic subunits), the N-terminal region determines ER localization, and the N-terminal part of Nox1 (but not Nox4) is cleaved. Replacing the Nox1 N-terminus with the Nox4 signal peptide redirected Nox1 from plasma membrane to vesicular structures and switched ROS from O2- to H2O2.","method":"Chimeric Nox1/Nox4 constructs expressed in HEK293 cells, TIRF microscopy, ROS type measurement, Myc-tagged Nox constructs, co-expression studies","journal":"Antioxidants & redox signaling","confidence":"High","confidence_rationale":"Tier 1 / Strong — reconstitution with chimeric constructs and domain-swap mutagenesis directly mapping structural determinants of localization and ROS type, multiple orthogonal readouts","pmids":["19061439"],"is_preprint":false},{"year":2010,"finding":"NOX4 is required for PDGF-induced cell cycle entry in normal human fibroblasts; NOX4 knockdown did not block cyclin D1 upregulation but reduced ERK1 phosphorylation hours after stimulation and increased p53 and p21 levels. Co-knockdown of NOX4 with p53 or p21 rescued Rb phosphorylation, indicating NOX4 promotes cell cycle entry by suppressing a p53/p21 checkpoint.","method":"siRNA screen, Rb phosphorylation assay, cyclin D1/p53/p21 immunoblotting, co-knockdown epistasis","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — epistasis co-knockdown experiments in human fibroblasts, single lab, multiple pathway markers","pmids":["20531308"],"is_preprint":false},{"year":2011,"finding":"Nox4 overexpression in cardiomyocytes in vivo activates the Nrf2 transcriptional pathway, leading to increased expression of antioxidant/detoxifying genes and elevated GSH and reduced:oxidized GSH ratio; these effects are abolished in Nrf2-null mice, demonstrating that Nox4-derived ROS activates Nrf2-dependent antioxidant defense.","method":"Transgenic mouse overexpression, microarray transcriptomics, Q-PCR, glutathione measurement, Nrf2 knockout genetic background","journal":"Free radical biology & medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo transgenic plus genetic epistasis with Nrf2-null, single lab","pmids":["21554947"],"is_preprint":false},{"year":2014,"finding":"NOX4 silencing in VHL-deficient renal carcinoma cells abrogates nuclear accumulation of HIF2α and blocks cell branching, invasion, colony formation, and xenograft growth, demonstrating that NOX4-derived ROS is required for HIF2α nuclear localization and renal tumorigenesis.","method":"siRNA knockdown, ROS scavengers (TEMPOL, MnSOD/catalase overexpression), nuclear fractionation for HIF2α, in vitro invasion/colony assays, murine xenograft model","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal ROS manipulation methods (siRNA, scavengers, antioxidant overexpression) with in vivo xenograft validation, single lab","pmids":["24755467"],"is_preprint":false},{"year":2014,"finding":"NOX4 activity is increased in the ER (but not mitochondria) of cardiomyocytes during energy deprivation; NOX4-derived ROS activates the PERK-eIF2α-ATF4 pathway to induce autophagy, which preserves cellular energy and limits cell death.","method":"Subcellular fractionation, NOX4 knockdown/knockout, ER-specific ROS measurement, PERK/eIF2α/ATF4 pathway activation assays, autophagy flux measurement, cardiomyocyte viability","journal":"Autophagy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — organelle-specific activity localization plus pathway knockdown epistasis, single lab","pmids":["24492492"],"is_preprint":false},{"year":2015,"finding":"FLT3ITD-driven leukemic transformation elevates NOX4 expression via STAT5-mediated activation of the NOX4 promoter; NOX4-derived ROS inactivates protein-tyrosine phosphatase DEP-1/PTPRJ, sustaining FLT3ITD signaling. Nox4 knockout hematopoietic progenitors are refractory to FLT3ITD transformation in vitro, and NOX4 downregulation attenuates myeloproliferative disease in vivo.","method":"NOX4 mRNA/protein quantification in FLT3ITD cells, STAT5 ChIP on NOX4 promoter, siRNA/knockout, DEP-1 PTP activity assay, ROS measurement, in vitro transformation assay, in vivo mouse disease models","journal":"Leukemia","confidence":"High","confidence_rationale":"Tier 2 / Strong — transcriptional mechanism (STAT5→NOX4 promoter), enzymatic PTP activity assay, in vitro and in vivo rescue experiments across multiple labs/models","pmids":["26308771"],"is_preprint":false},{"year":2016,"finding":"Nox4 regulates eIF2α-mediated stress signaling by binding to the PP1-targeting subunit GADD34 at the ER and inhibiting PP1 phosphatase activity through oxidation of its metal center (not thiol oxidation), thereby sustaining eIF2α phosphorylation and ATF4 levels. This is spatially confined to the ER and does not affect PP1 targets at other locations.","method":"Co-immunoprecipitation of Nox4 with GADD34, PP1 activity assays with metal center vs. thiol oxidation characterization, eIF2α phosphorylation/ATF4 immunoblotting, ER localization, genetic knockdown/overexpression, in vivo heart ischemia-reperfusion and acute kidney injury models","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct biochemical enzymatic inhibition assay with mechanistic distinction (metal center vs. thiol), protein–protein interaction by Co-IP, in vivo disease models, multiple orthogonal methods","pmids":["26742780"],"is_preprint":false},{"year":2016,"finding":"FYN tyrosine kinase directly interacts with the C-terminal domain of NOX4 and phosphorylates it at tyrosine 566, negatively regulating NOX4-induced O2- production and apoptosis in cardiomyocytes. FYN and NOX4 co-localize in perinuclear mitochondria, ER, and nuclear fractions. FYN-deficient mice have exacerbated cardiac hypertrophy with increased ROS, rescued by Nox4 deletion.","method":"Co-immunoprecipitation, co-localization imaging, site-directed mutagenesis (Y566), NOX4 activity assay, Nox4-/- genetic rescue of FYN-/- cardiac phenotype, transverse aortic constriction model, human failing heart analysis","journal":"The Journal of clinical investigation","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct phosphorylation site identified by mutagenesis, Co-IP, in vivo genetic epistasis rescue, multiple orthogonal methods","pmids":["27525436"],"is_preprint":false},{"year":2017,"finding":"In VEGF-stimulated endothelial cells, Nox4-derived H2O2 activates Nox2, which promotes mitochondrial ROS production via S36 phosphorylation of p66Shc; this Nox4/Nox2/pSer36-p66Shc/mtROS feed-forward axis drives sustained VEGFR2 phosphorylation, EC migration, and proliferation (angiogenesis).","method":"Cytosol/mitochondria-targeted RoGFP biosensors with real-time imaging, Nox4/Nox2 siRNA, mitochondria-targeted catalase overexpression, Nox4 overexpression, p66Shc(S36A) mutant, VEGFR2 tyrosine phosphorylation assay, EC migration/proliferation assays","journal":"American journal of physiology. Cell physiology","confidence":"High","confidence_rationale":"Tier 1 / Strong — RoGFP real-time organelle-specific ROS imaging, genetic epistasis (double knockdown, mutant p66Shc), multiple orthogonal functional assays, single lab with rigorous controls","pmids":["28424170"],"is_preprint":false},{"year":2017,"finding":"NOX4 contains an ATP-binding motif; ATP directly binds and negatively regulates NOX4 activity. NOX4 localizes to the inner mitochondrial membrane, and subcellular redistribution of ATP from mitochondria acts as an allosteric switch to activate NOX4. NOX4-derived ROS inhibits PCAF-dependent acetylation and lysosomal degradation of PKM2.","method":"ATP-binding assay (identification of ATP-binding motif), subcellular fractionation for inner mitochondrial membrane localization, NOX4 activity assays with ATP titration, PKM2 acetylation/ubiquitination assays, PCAF interaction, NOX4 silencing in xenograft models","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct biochemical binding assay identifying novel allosteric ATP regulation, enzymatic activity measurement, subcellular localization, and mechanistic PKM2 downstream pathway, multiple orthogonal methods","pmids":["29051480"],"is_preprint":false},{"year":2018,"finding":"NOX4-derived H2O2 in podocytes activates TRPC6-dependent calcium influx, contributing to podocyte damage in diabetic kidney disease; SSNox4-/- rats show lower basal intracellular Ca2+ in podocytes and less DKD-associated damage, and H2O2-stimulated TRPC-dependent calcium influx is blunted in Trpc6-knockout podocytes.","method":"Nox4 knockout rat (SSNox4-/-), TRPC6/TRPC5/6 knockout mice, H2O2 stimulation, live calcium imaging, electrophysiology patch-clamp, biosensor measurements, electron microscopy","journal":"Journal of the American Society of Nephrology : JASN","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple knockout models, direct calcium imaging, electrophysiology, and electron microscopy across species, multiple orthogonal methods","pmids":["29793963"],"is_preprint":false},{"year":2019,"finding":"CD44 directly associates with NOX4 in tachypaced atrial myocytes and atrial fibrillation patient tissues; blocking HAS/HA/CD44 signaling attenuates tachypacing-induced NOX4 expression, oxidative stress, and Ca2+-handling abnormalities (ox-CaMKII/p-RyR2).","method":"Co-immunoprecipitation of CD44 with NOX4, CD44-/- mice, anti-CD44 blocking antibody, Ca2+ spark measurement, tachypacing model in vitro and ex vivo, AF patient tissue analysis","journal":"Journal of molecular and cellular cardiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP demonstrating direct CD44-NOX4 interaction plus genetic/pharmacological epistasis and functional Ca2+ imaging, single lab","pmids":["31419440"],"is_preprint":false},{"year":2020,"finding":"Nox4 upregulated at ER-mitochondria contact sites (MAMs) during stress inhibits InsP3 receptor-mediated Ca2+ transfer from ER to mitochondria by augmenting Akt-dependent phosphorylation of InsP3R, thereby reducing mitochondrial permeability transition and necrosis; in ischemia-reperfusion, Nox4 limits myocardial infarct size through this mechanism.","method":"MAM fractionation, Nox4 overexpression/knockout, InsP3R phosphorylation assays, mitochondrial Ca2+ measurement, mPT assay, cardiac ischemia-reperfusion model, cardiomyocyte/neuron stress models","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1 / Strong — mechanistic dissection at MAM with organelle-specific fractionation, InsP3R phosphorylation, Ca2+ flux and mPT measurements, in vivo cardiac model, multiple orthogonal approaches","pmids":["33001475"],"is_preprint":false},{"year":2020,"finding":"Nox4 is required for exercise-induced expression of metabolic genes (Ucp3, Hk2, Pdk4) in skeletal muscle; global and endothelial-specific Nox4 deletion impairs glucose and fatty acid oxidation after acute exercise, revealing an endothelium-to-skeletal muscle cross-talk mediated by Nox4-derived H2O2.","method":"Global and endothelial-specific Nox4 KO mice, 14C-labeled substrate oxidation assays ex vivo, qPCR/immunoblotting for metabolic genes, chronic exercise regimen with time-to-exhaustion measurement, catalase transgenic mice","journal":"Molecular metabolism","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — cell-type-specific KO models with functional metabolic assays, single lab, multiple orthogonal methods","pmids":["33400973"],"is_preprint":false},{"year":2020,"finding":"SIRT1 loss in cachectic muscle induces NF-κB signaling that upregulates FOXO transcription factors and NOX4 expression; skeletal muscle-specific Nox4 knockout or pharmacological NOX4 blockade abrogates tumor-induced cachexia in mice, placing NOX4 downstream of the SIRT1/NF-κB/FOXO axis in muscle wasting.","method":"RNA-seq, Nox4 muscle-specific KO mice, pharmacological NOX4 inhibition, SIRT1 reconstitution, co-culture and in vivo cancer cachexia models","journal":"The Journal of experimental medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — tissue-specific KO with in vivo rescue plus transcriptomic pathway mapping, single lab","pmids":["32441762"],"is_preprint":false},{"year":2020,"finding":"Nox4 overexpression induces oxidation of HDAC4 in HEK293 cells and endothelial cells; Nox4-derived H2O2 increases HDAC4 phosphorylation at Ser632 and disrupts the HDAC4/Mef2A complex, de-repressing Mef2A and enabling proper endothelial tube formation. A redox-insensitive HDAC4 mutant blocks tube formation, while a redox-dead Nox4 mutant fails to rescue it.","method":"Tetracycline-inducible Nox4 overexpression in HEK293, HDAC4 oxidation assay, HDAC4/Mef2A co-immunoprecipitation, Ser632 phosphorylation analysis, redox-insensitive HDAC4 mutant, redox-dead Nox4 mutant, endothelial tube formation assay","journal":"Redox biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct substrate oxidation assay, protein complex disruption by Co-IP, structure-function mutagenesis of both Nox4 and HDAC4, functional tube formation readout","pmids":["32818796"],"is_preprint":false},{"year":2021,"finding":"Nox4 promotes RANKL-induced autophagy and osteoclastogenesis by stimulating non-mitochondrial ROS production that activates the PERK/eIF-2α/ATF4 unfolded protein response pathway; inhibition of Nox4 or PERK/eIF-2α/ATF4 or ROS scavenging similarly blocks autophagy and osteoclastogenesis.","method":"Nox4 inhibitor (5-O-methyl quercetin), Nox4 shRNA knockdown, ROS scavenger (NAC), PERK inhibitor (GSK2606414), autophagy markers, osteoclastogenesis assays, pathway epistasis","journal":"Frontiers in pharmacology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic and pharmacological epistasis with multiple nodes of PERK/eIF-2α/ATF4 pathway, single lab","pmids":["34650437"],"is_preprint":false},{"year":2021,"finding":"Neuronal NOX4 promotes pathological tau accumulation by impairing autophagy-lysosomal pathway flux; global Nox4 knockout and neuronal Nox4 knockdown in mice reduced accumulation of hyperphosphorylated tau, improved macroautophagy flux, reduced neurotoxicity, and prevented cognitive decline in a tauopathy model.","method":"Global Nox4 KO mice, neuronal-targeted AAV-mediated Nox4 knockdown, humanized tauopathy mouse model (AAV-TauP301L), tau immunohistochemistry, autophagy flux assays, behavioral testing","journal":"Redox biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — neuronal-specific KD with mechanistic autophagy pathway readout and in vivo behavioral rescue, single lab","pmids":["34922273"],"is_preprint":false},{"year":2021,"finding":"NOX4 deletion in mice promotes cancerogen-induced tumor formation by reducing nuclear PP2A abundance; NOX4-derived H2O2 continuously oxidizes AKT, trapping PP2A in the cytosol, which maintains γH2AX (phospho-H2AX) levels for DNA damage recognition. Without Nox4, PP2A translocates to the nucleus, dephosphorylates γH2AX, impairing DNA damage recognition and simultaneously increasing AKT-driven proliferation.","method":"Nox4 KO mice, carcinogen-induced tumor models, AKT oxidation assay, PP2A subcellular fractionation, γH2AX immunostaining/immunoblotting, PP2A activity assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct AKT oxidation assay, PP2A subcellular localization by fractionation, γH2AX functional DNA damage readout, in vivo genetic KO with two carcinogen models","pmids":["33836590"],"is_preprint":false},{"year":2021,"finding":"CYB5R3 localizes to the mitochondrial outer membrane and directly interacts with NOX4; CYB5R3 activity and membrane translocation are required for optimal NOX4-dependent H2O2 generation via coenzyme Q (CoQ). Cyb5r3 knockdown reduces total H2O2 but increases mitochondrial O2•-, and cells lacking the CoQ-synthesizing enzyme COQ6 show decreased NOX4-derived H2O2.","method":"APEX2-based electron microscopy, proximity biotinylation, proximity ligation assay, Co-IP, CYB5R3 activity mutants, COQ6 knockdown, mitochondrial ROS species measurement, endothelium-specific Cyb5r3 KO mice","journal":"Redox biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple independent protein interaction methods (APEX2-EM, proximity biotinylation, PLA, Co-IP), organelle localization, structure-function mutants, and CoQ dependency established","pmids":["34656824"],"is_preprint":false},{"year":2023,"finding":"NOX4 locus contains iron-response element-like (IRE-like) sequences bound and repressed by iron regulatory protein 1 (IRP1); excess iron dissociates IRP1 from these sequences, activating NOX4 transcription, which increases lipid peroxides and causes ferroptosis-associated mitochondrial dysfunction in osteoblasts.","method":"IRP1 binding assays on NOX4 promoter (IRE-like sequences), NOX4 expression analysis with iron loading, ferroptosis markers (lipid peroxides, MDA), mitochondrial morphology/function assays, ferroptosis inhibitor ferrostatin-1 and iron chelator DFO in Hepc1-/- mice","journal":"Free radical biology & medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — IRP1-IRE-like sequence binding characterization on NOX4 promoter plus in vivo genetic/pharmacological validation, single lab","pmids":["36738798"],"is_preprint":false},{"year":2024,"finding":"DRD4 reduces NOX4 expression via suppression of ISG15, which ISGylates NOX4 and stabilizes it; when DRD4 is active, ISG15 levels fall, NOX4 ISGylation decreases, ubiquitination of NOX4 increases, and NOX4 is degraded via the ubiquitin-proteasome pathway, reducing oxidative stress in acute kidney injury.","method":"Transcriptome sequencing, ISG15 knockdown/overexpression, Co-IP for NOX4 ISGylation and ubiquitination, NOX4 protein stability assays, DRD4 KO/overexpression in AKI models (IRI and cisplatin), oxidative stress measurement","journal":"Redox biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biochemical ISGylation/ubiquitination assays for NOX4 stability plus in vivo genetic validation, single lab","pmids":["38354631"],"is_preprint":false},{"year":2024,"finding":"HOXD10 directly binds to the NOX4 promoter (confirmed by ChIP and dual-luciferase assay) and represses its transcription; HOXD10 overexpression attenuates TGF-β1-induced ferroptosis and renal fibrosis by reducing NOX4 expression and downstream ROS/lipid peroxide accumulation. HOXD10 is epigenetically silenced by hypermethylation in TGF-β1-treated cells.","method":"ChIP analysis, dual-luciferase reporter assay, HOXD10 overexpression (AAV in vivo), NOX4 expression assay, ferroptosis markers, bisulfite sequencing PCR for methylation, UUO fibrosis model","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — direct promoter binding confirmed by ChIP and luciferase, in vivo rescue, but single lab","pmids":["38844470"],"is_preprint":false},{"year":2024,"finding":"NOX4 interacts with activated PKCα (protein kinase C alpha) to promote ferroptosis of dopaminergic neurons; NOX4 inhibition reduced lipid peroxidation, iron accumulation, and astrocytic lipocalin-2 expression (reducing neuroinflammation) in a Parkinson's disease MPTP model. ATF3 transcriptionally increases NOX4 expression in dopaminergic neurons and astrocytes.","method":"Co-immunoprecipitation of NOX4 with PKCα, NOX4 inhibitor in MPTP mouse model, lipid peroxidation and iron assays, behavioral tests, lipocalin-2 immunostaining, ATF3 transcriptional analysis","journal":"Neural regeneration research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP for NOX4-PKCα interaction plus in vivo NOX4 inhibitor with multiple mechanistic readouts, single lab","pmids":["38993139"],"is_preprint":false},{"year":2020,"finding":"NOX4 (but not NOX2) activity and protein levels increase specifically in the ER of cardiomyocytes during energy deprivation; this ER-localized NOX4-derived ROS activates the PERK-eIF2α-ATF4 autophagy pathway, which is a critical adaptive response to energy stress.","method":"Organelle-specific NOX4 activity measurements (ER vs. mitochondria fractions), NOX4 genetic deletion, PERK/eIF2α/ATF4 pathway immunoblotting, autophagy markers, cardiomyocyte energy deprivation model","journal":"Autophagy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — organelle-resolved biochemistry with genetic KO and pathway epistasis, single lab","pmids":["24492492"],"is_preprint":false},{"year":2004,"finding":"Co-transfection of Nox4 with p22phox in osteoclasts enhances superoxide production and increases expression of cathepsin K and TRAP, with JNK activation and NF-κB inhibition, indicating p22phox is a necessary cofactor for Nox4 activity in osteoclasts.","method":"Nox4/p22phox co-transfection, superoxide assay, cathepsin K/TRAP expression, JNK/NF-κB signaling assays","journal":"Journal of cellular biochemistry","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single co-transfection experiment, single lab, no direct Nox4-p22phox interaction assay","pmids":["15108351"],"is_preprint":false},{"year":2012,"finding":"NOX4 produces H2O2 constitutively rather than superoxide (O2-), making it incapable of scavenging NO; Nox4 knockout mice on an ApoE-/- background develop increased atherosclerosis, and endothelial-specific (but not macrophage-specific) Nox4 deletion increases macrophage adhesion to endothelium, demonstrating an anti-atherosclerotic endothelial function of Nox4.","method":"Tamoxifen-inducible Nox4 KO crossed with ApoE-/- mice, partial carotid artery ligation model, atherosclerosis quantification, cell-type-specific KO, macrophage adhesion assay","journal":"European heart journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — cell-type-specific genetic KO with mechanistic gene expression analysis, in vivo atherosclerosis models, single lab","pmids":["26385958"],"is_preprint":false}],"current_model":"NOX4 is a constitutively active, NADPH-dependent oxidase that produces H2O2 (rather than superoxide) primarily at the endoplasmic reticulum and ER-mitochondria contact sites (MAMs), where it mediates spatially confined redox signaling: it requires p22phox and, at the mitochondrial outer membrane, CYB5R3/coenzyme Q for full H2O2 output; its activity is allosterically inhibited by direct ATP binding and negatively regulated by FYN kinase-mediated phosphorylation of Y566; structurally, the N-terminal region determines ER localization while the cytosolic tail confers subunit-independent constitutive activity; mechanistically, NOX4-derived H2O2 inactivates PP1 metal centers to sustain eIF2α phosphorylation, oxidizes HDAC4 to de-repress Mef2A, oxidizes AKT to retain PP2A in the cytosol for DNA damage surveillance, activates Nrf2-dependent antioxidant programs, and at the MAM augments Akt-dependent InsP3R phosphorylation to restrict ER-to-mitochondria Ca2+ flux and prevent mitochondrial permeability transition, while upstream its transcription is controlled by STAT5, HOXD10, ATF3, TGF-β/Brd4-p300 epigenetic mechanisms, and IRP1 binding to IRE-like sequences in its promoter."},"narrative":{"mechanistic_narrative":"NOX4 is a constitutively active, NADPH-dependent oxidase homologous to the phagocyte catalytic subunit gp91phox that generates reactive oxygen species to drive spatially confined redox signaling [PMID:11376945, PMID:19061439]. Distinct structural elements set its output and address: the cytosolic tail confers subunit-independent constitutive activity while the N-terminal region directs ER localization and dictates that NOX4 releases H2O2 rather than superoxide [PMID:19061439], a determinant confirmed by genetic models showing NOX4 produces H2O2 constitutively [PMID:26385958]. It localizes to the endoplasmic reticulum, mitochondria, and ER-mitochondria contact sites (MAMs) and requires p22phox as a cofactor, with the partner interaction demonstrated directly in endothelial cells [PMID:15706079, PMID:19706525, PMID:16987004]; at mitochondrial membranes, CYB5R3 directly interacts with NOX4 and, through coenzyme Q, is required for optimal H2O2 output [PMID:34656824]. NOX4 activity is allosterically inhibited by direct ATP binding [PMID:29051480] and negatively regulated by FYN-mediated phosphorylation at Y566 [PMID:27525436]. The H2O2 it produces acts on defined targets to control discrete cellular programs: it binds the PP1-targeting subunit GADD34 at the ER and oxidizes the PP1 metal center to sustain eIF2α phosphorylation and ATF4-driven autophagy [PMID:26742780, PMID:24492492], oxidizes HDAC4 to disrupt the HDAC4/Mef2A complex and enable endothelial tube formation [PMID:32818796], oxidizes AKT to retain PP2A in the cytosol and preserve γH2AX-dependent DNA damage surveillance [PMID:33836590], activates Nrf2-dependent antioxidant programs [PMID:21554947], and at the MAM augments Akt-dependent InsP3R phosphorylation to restrict ER-to-mitochondria Ca2+ flux and limit mitochondrial permeability transition [PMID:33001475]. Through these outputs NOX4 governs proliferation, angiogenesis, metabolic gene expression, and cell-fate decisions including ferroptosis, and its transcription is controlled by inputs including STAT5, HOXD10, ATF3, and IRP1 binding to IRE-like promoter sequences [PMID:26308771, PMID:38844470, PMID:38993139, PMID:36738798]. NOX4-derived signaling is implicated across diabetic kidney disease, cardiac remodeling, leukemic transformation, tauopathy, and cancer [PMID:16135519, PMID:27525436, PMID:26308771, PMID:34922273].","teleology":[{"year":2001,"claim":"Establishing NOX4 as an NADPH oxidase family member answered whether it could be an enzymatic ROS source by showing it shares the catalytic architecture of gp91phox.","evidence":"cDNA cloning and sequence/domain analysis with tissue expression profiling","pmids":["11376945"],"confidence":"Medium","gaps":["Enzymatic activity not directly reconstituted in this work","ROS species (H2O2 vs O2-) not determined","Subcellular site not defined"]},{"year":2003,"claim":"Linking NOX4 to angiotensin II-induced ROS and Akt activation placed it in a defined receptor-driven signaling pathway in mesangial cells.","evidence":"Antisense knockdown, dominant-negative Rac1, ROS and Akt phosphorylation assays","pmids":["12842860"],"confidence":"Medium","gaps":["Antisense specificity limits, no genetic KO","Direct ROS species not resolved","Mechanism of Rac1/arachidonate input unclear"]},{"year":2005,"claim":"ER localization of NOX4 answered where the enzyme acts, distinguishing it from plasma-membrane oxidases.","evidence":"Confocal microscopy of NOX4-GFP with ER, lysosome, and mitochondrial markers in HUVECs","pmids":["15706079"],"confidence":"Medium","gaps":["Based on overexpressed GFP fusion","Does not address mitochondrial pools reported later","No structural basis for targeting"]},{"year":2006,"claim":"Demonstrating a direct NOX4-p22phox interaction at the ER identified the obligatory cofactor needed for endothelial ROS production.","evidence":"Bimolecular fluorescence complementation, calreticulin co-localization, siRNA, ROS assays","pmids":["16987004"],"confidence":"Medium","gaps":["BiFC can trap transient interactions","Stoichiometry not defined","Single lab"]},{"year":2009,"claim":"Domain-swap mapping established that the N-terminus dictates ER localization and the cytosolic tail confers constitutive, subunit-independent activity and H2O2 output, defining the structural logic of NOX4 specificity.","evidence":"Chimeric Nox1/Nox4 constructs, TIRF microscopy, ROS-type measurement in HEK293","pmids":["19061439"],"confidence":"High","gaps":["No atomic structure","Mechanism of H2O2 vs O2- determination at the catalytic core unresolved"]},{"year":2009,"claim":"Detection of NOX4 in mitochondria broadened its localization beyond the ER and tied it to glucose-induced mitochondrial superoxide in diabetic kidney.","evidence":"Subcellular fractionation, Mitotracker confocal, MitoProt prediction, siRNA, organelle NADPH oxidase assays","pmids":["19706525"],"confidence":"Medium","gaps":["Reconciliation with H2O2-only model not addressed here","Import/targeting mechanism unknown"]},{"year":2011,"claim":"Showing NOX4-derived ROS activates Nrf2 reframed NOX4 as a driver of antioxidant defense rather than purely oxidative damage.","evidence":"Transgenic cardiomyocyte overexpression, transcriptomics, glutathione assays, Nrf2-null epistasis","pmids":["21554947"],"confidence":"Medium","gaps":["Overexpression context","Direct sensor of NOX4 H2O2 in Keap1/Nrf2 axis not identified"]},{"year":2015,"claim":"Identifying STAT5-driven NOX4 transcription and NOX4-mediated DEP-1/PTPRJ inactivation explained how NOX4 sustains oncogenic FLT3ITD signaling in leukemia.","evidence":"STAT5 ChIP on NOX4 promoter, PTP activity assay, knockout transformation assays, in vivo disease models","pmids":["26308771"],"confidence":"High","gaps":["Site of redox-DEP1 encounter not localized","Generalizability beyond FLT3ITD leukemia"]},{"year":2016,"claim":"The GADD34/PP1 work provided a direct molecular target, showing NOX4 H2O2 oxidizes the PP1 metal center to sustain eIF2α phosphorylation in a spatially confined manner at the ER.","evidence":"Co-IP with GADD34, PP1 activity assays distinguishing metal-center vs thiol oxidation, in vivo ischemia and AKI models","pmids":["26742780"],"confidence":"High","gaps":["How H2O2 is delivered to confined PP1 pool not fully resolved","Other PP1 holoenzymes spared but mechanism of selectivity incomplete"]},{"year":2016,"claim":"Identification of FYN-mediated Y566 phosphorylation defined a negative post-translational regulatory input restraining NOX4 in the heart.","evidence":"Co-IP, Y566 site-directed mutagenesis, activity assays, Nox4-/- rescue of FYN-/- cardiac phenotype","pmids":["27525436"],"confidence":"High","gaps":["Structural effect of Y566 phosphorylation on catalysis unknown","Upstream control of FYN-NOX4 axis"]},{"year":2017,"claim":"Discovery of an ATP-binding motif established direct allosteric inhibition of NOX4 by ATP, providing a metabolic switch coupling energy state to oxidase activity.","evidence":"ATP-binding and titration activity assays, inner mitochondrial membrane fractionation, PKM2 downstream pathway, xenografts","pmids":["29051480"],"confidence":"High","gaps":["ATP-binding site not structurally resolved","Inner-membrane localization claim contrasts with ER/MAM models"]},{"year":2017,"claim":"The Nox4/Nox2/p66Shc feed-forward axis showed how NOX4 H2O2 amplifies mitochondrial ROS and VEGFR2 signaling to drive angiogenesis.","evidence":"Organelle-targeted RoGFP imaging, double knockdown, p66Shc(S36A) mutant, EC migration/proliferation assays","pmids":["28424170"],"confidence":"High","gaps":["Direct molecular link from NOX4 H2O2 to Nox2 activation not defined","Spatial coupling of the two oxidases unresolved"]},{"year":2018,"claim":"NOX4 H2O2 activation of TRPC6-dependent calcium influx connected the oxidase to podocyte calcium dysregulation in diabetic kidney disease.","evidence":"Nox4 KO rat, TRPC6 KO, H2O2 stimulation, calcium imaging, patch-clamp, EM","pmids":["29793963"],"confidence":"High","gaps":["Direct oxidative modification of TRPC6 not demonstrated","Cell-type generality unclear"]},{"year":2019,"claim":"Direct CD44-NOX4 association linked HA/CD44 signaling to NOX4-driven Ca2+-handling abnormalities in atrial fibrillation.","evidence":"Co-IP, CD44-/- mice, blocking antibody, Ca2+ spark measurement, AF patient tissue","pmids":["31419440"],"confidence":"Medium","gaps":["Single Co-IP without reciprocal structural mapping","Whether CD44 regulates NOX4 activity or only expression unclear"]},{"year":2020,"claim":"Localizing NOX4 to MAMs and showing it augments Akt-dependent InsP3R phosphorylation defined a cytoprotective role restricting ER-to-mitochondria Ca2+ flux and limiting permeability transition.","evidence":"MAM fractionation, overexpression/KO, InsP3R phosphorylation, mitochondrial Ca2+ and mPT assays, cardiac ischemia-reperfusion","pmids":["33001475"],"confidence":"High","gaps":["How NOX4 H2O2 promotes Akt activity at MAM not fully resolved","Direct InsP3R oxidation not shown"]},{"year":2020,"claim":"ER-confined NOX4 activation of the PERK-eIF2α-ATF4 pathway during energy stress established NOX4 as an inducer of adaptive autophagy.","evidence":"Organelle-resolved NOX4 activity, genetic deletion, UPR pathway immunoblotting, autophagy flux in cardiomyocytes","pmids":["24492492"],"confidence":"Medium","gaps":["Direct PERK oxidation/sensor not identified","Relationship to the GADD34/PP1 mechanism not integrated"]},{"year":2020,"claim":"Cell-type-specific KO showed endothelial NOX4-derived H2O2 mediates endothelium-to-muscle cross-talk required for exercise-induced metabolic gene expression.","evidence":"Global and endothelial-specific Nox4 KO, 14C substrate oxidation, qPCR, catalase transgenics","pmids":["33400973"],"confidence":"Medium","gaps":["Identity of the cross-talk signal beyond H2O2 unknown","Direct muscle target genes' redox sensors undefined"]},{"year":2020,"claim":"Placing NOX4 downstream of SIRT1/NF-κB/FOXO defined it as an effector in tumor-induced muscle wasting.","evidence":"RNA-seq, muscle-specific Nox4 KO, pharmacological inhibition, SIRT1 reconstitution, cachexia models","pmids":["32441762"],"confidence":"Medium","gaps":["Downstream NOX4 targets in wasting muscle not identified","Direct FOXO-NOX4 promoter binding not shown here"]},{"year":2020,"claim":"Showing NOX4 oxidizes HDAC4 to disrupt the HDAC4/Mef2A complex provided a direct chromatin-linked substrate enabling endothelial tube formation.","evidence":"Inducible overexpression, HDAC4 oxidation assay, HDAC4/Mef2A Co-IP, redox-dead NOX4 and redox-insensitive HDAC4 mutants, tube formation","pmids":["32818796"],"confidence":"High","gaps":["Spatial route of H2O2 to nuclear/cytosolic HDAC4 unclear","Overexpression-based system"]},{"year":2010,"claim":"NOX4 was shown to promote cell cycle entry by suppressing a p53/p21 checkpoint downstream of PDGF.","evidence":"siRNA, Rb phosphorylation, cyclin D1/p53/p21 immunoblotting, co-knockdown epistasis in fibroblasts","pmids":["20531308"],"confidence":"Medium","gaps":["Direct redox target upstream of p53/p21 not identified","Single cell type"]},{"year":2014,"claim":"NOX4-derived ROS was found necessary for HIF2α nuclear accumulation and tumorigenesis in VHL-deficient renal carcinoma.","evidence":"siRNA, ROS scavengers, MnSOD/catalase, nuclear fractionation, invasion/colony assays, xenografts","pmids":["24755467"],"confidence":"Medium","gaps":["Molecular target linking NOX4 ROS to HIF2α transport not defined","Single tumor model"]},{"year":2021,"claim":"NOX4 oxidation of AKT to retain PP2A in the cytosol revealed a tumor-suppressive role maintaining γH2AX-dependent DNA damage surveillance.","evidence":"Nox4 KO mice, carcinogen models, AKT oxidation assay, PP2A fractionation, γH2AX readouts","pmids":["33836590"],"confidence":"High","gaps":["Site of AKT oxidation and its kinetics not fully mapped","How redox state directs PP2A shuttling mechanistically"]},{"year":2021,"claim":"Identifying CYB5R3 as a direct NOX4 partner at the mitochondrial outer membrane defined a CoQ-dependent requirement for full H2O2 output.","evidence":"APEX2-EM, proximity biotinylation, PLA, Co-IP, CYB5R3 activity mutants, COQ6 knockdown, endothelial Cyb5r3 KO mice","pmids":["34656824"],"confidence":"High","gaps":["Electron-transfer mechanism between CYB5R3/CoQ and NOX4 not biochemically reconstituted","Reconciliation with ER pool of NOX4"]},{"year":2021,"claim":"NOX4-driven non-mitochondrial ROS was shown to activate PERK/eIF2α/ATF4 to promote autophagy and osteoclastogenesis, extending the UPR-autophagy axis to bone.","evidence":"Nox4 inhibitor and shRNA, NAC, PERK inhibitor, autophagy and osteoclastogenesis assays","pmids":["34650437"],"confidence":"Medium","gaps":["Pharmacological inhibitor specificity","Direct UPR sensor target not identified"]},{"year":2021,"claim":"Neuronal NOX4 was found to impair autophagy-lysosomal flux and drive tau accumulation, linking the oxidase to tauopathy.","evidence":"Global KO and neuronal AAV knockdown, tauopathy mouse model, autophagy flux, behavior","pmids":["34922273"],"confidence":"Medium","gaps":["Molecular redox target controlling autophagy flux not identified","Single model"]},{"year":2023,"claim":"IRP1 binding to IRE-like sequences in the NOX4 locus established an iron-sensing transcriptional input coupling iron overload to NOX4-driven ferroptosis.","evidence":"IRP1 binding assays on NOX4 promoter, iron loading, ferroptosis markers, Hepc1-/- mice with ferrostatin/DFO","pmids":["36738798"],"confidence":"Medium","gaps":["IRE-like element function not validated by mutagenesis here","Direct vs indirect IRP1 effect"]},{"year":2024,"claim":"ISG15-mediated ISGylation stabilizing NOX4 against ubiquitin-proteasome degradation, controlled by DRD4, defined a post-translational stability switch in acute kidney injury.","evidence":"Transcriptomics, ISG15 manipulation, Co-IP for ISGylation/ubiquitination, stability assays, DRD4 KO/overexpression AKI models","pmids":["38354631"],"confidence":"Medium","gaps":["ISGylation sites on NOX4 not mapped","E3 ligase mediating NOX4 ubiquitination unidentified"]},{"year":2024,"claim":"HOXD10 was shown to directly bind and repress the NOX4 promoter, and its hypermethylation de-represses NOX4 to drive TGF-β1-induced ferroptosis and renal fibrosis.","evidence":"ChIP, dual-luciferase, HOXD10 AAV overexpression, ferroptosis markers, bisulfite sequencing, UUO model","pmids":["38844470"],"confidence":"Medium","gaps":["Single lab","Interaction with other NOX4 transcriptional regulators not integrated"]},{"year":2024,"claim":"NOX4 interaction with activated PKCα and ATF3-driven transcription was linked to ferroptosis of dopaminergic neurons in a Parkinson's model.","evidence":"Co-IP with PKCα, NOX4 inhibitor in MPTP mice, lipid peroxidation/iron assays, ATF3 analysis","pmids":["38993139"],"confidence":"Medium","gaps":["Direct functional consequence of PKCα binding on NOX4 activity unclear","Single model and lab"]},{"year":null,"claim":"How NOX4's multiple subcellular pools (ER, MAM, mitochondrial membranes) are coordinated, and how a single diffusible H2O2 output achieves the documented target selectivity (PP1, AKT, HDAC4, InsP3R), remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No atomic structure of human NOX4 to explain H2O2 vs O2- output","Mechanism conferring spatial confinement and target specificity of NOX4 H2O2 not defined","Reconciliation of ER vs inner/outer mitochondrial membrane localization claims"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016491","term_label":"oxidoreductase activity","supporting_discovery_ids":[0,7,33]},{"term_id":"GO:0140657","term_label":"ATP-dependent activity","supporting_discovery_ids":[16]}],"localization":[{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[3,4,11,13,31]},{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[6,16,26]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[7]}],"pathway":[{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[13,9,11,31]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[2,12,15,17,25]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[27,29,30,19]},{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[11,23,24,31]}],"complexes":[],"partners":["CYBA","FYN","CYB5R3","PPP1R15A","HDAC4","CD44","PRKCA"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9NPH5","full_name":"NADPH oxidase 4","aliases":["Kidney oxidase-1","KOX-1","Kidney superoxide-producing NADPH oxidase","Renal NAD(P)H-oxidase"],"length_aa":578,"mass_kda":66.9,"function":"NADPH oxidase that catalyzes predominantly the reduction of oxygen to H2O2 (PubMed:14966267, PubMed:15356101, PubMed:15927447, PubMed:21343298, PubMed:25062272). Can also catalyze to a smaller extent, the reduction of oxygen to superoxide (PubMed:10869423, PubMed:11032835, PubMed:15155719, PubMed:15572675, PubMed:15927447, PubMed:16019190, PubMed:16179589, PubMed:16230378, PubMed:16324151, PubMed:25062272). May function as an oxygen sensor regulating the KCNK3/TASK-1 potassium channel and HIF1A activity (PubMed:16019190). May regulate insulin signaling cascade (PubMed:14966267). May play a role in apoptosis, bone resorption and lipolysaccharide-mediated activation of NFKB (PubMed:15356101, PubMed:15572675). May produce superoxide in the nucleus and play a role in regulating gene expression upon cell stimulation (PubMed:16324151). Promotes ferroptosis, reactive oxygen species production and reduced glutathione (GSH) levels by activating NLRP3 inflammasome activation and cytokine release (PubMed:39909992) NADPH oxidase that catalyzes the generation of superoxide from molecular oxygen utilizing NADPH as an electron donor (PubMed:15721269, PubMed:23393389). Involved in redox signaling in vascular cells (PubMed:23393389). Modulates the nuclear activation of ERK1/2 and the ELK1 transcription factor, and is capable of inducing nuclear DNA damage (PubMed:23393389) Lacks superoxide-generating NADPH oxidase activity","subcellular_location":"Cytoplasm; Cytoplasm, perinuclear region","url":"https://www.uniprot.org/uniprotkb/Q9NPH5/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/NOX4","classification":"Not Classified","n_dependent_lines":1,"n_total_lines":1208,"dependency_fraction":0.0008278145695364238},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/NOX4","total_profiled":1310},"omim":[{"mim_id":"617509","title":"VON WILLEBRAND FACTOR A DOMAIN-CONTAINING PROTEIN 8; VWA8","url":"https://www.omim.org/entry/617509"},{"mim_id":"606255","title":"STATURE AS A QUANTITATIVE TRAIT","url":"https://www.omim.org/entry/606255"},{"mim_id":"605441","title":"ADIPOCYTE-, C1q-, AND COLLAGEN DOMAIN-CONTAINING; ADIPOQ","url":"https://www.omim.org/entry/605441"},{"mim_id":"605261","title":"NADPH OXIDASE 4; NOX4","url":"https://www.omim.org/entry/605261"},{"mim_id":"600524","title":"RYK RECEPTOR-LIKE TYROSINE KINASE; RYK","url":"https://www.omim.org/entry/600524"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Group enriched","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"blood vessel","ntpm":35.3},{"tissue":"kidney","ntpm":128.9}],"url":"https://www.proteinatlas.org/search/NOX4"},"hgnc":{"alias_symbol":["KOX-1","KOX"],"prev_symbol":[]},"alphafold":{"accession":"Q9NPH5","domains":[{"cath_id":"-","chopping":"6-159_211-218_277-297","consensus_level":"high","plddt":91.6423,"start":6,"end":297},{"cath_id":"2.40.30.10","chopping":"309-387_404-414","consensus_level":"high","plddt":91.8128,"start":309,"end":414},{"cath_id":"3.40.50.80","chopping":"417-576","consensus_level":"high","plddt":92.9165,"start":417,"end":576}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NPH5","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NPH5-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NPH5-F1-predicted_aligned_error_v6.png","plddt_mean":87.0},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=NOX4","jax_strain_url":"https://www.jax.org/strain/search?query=NOX4"},"sequence":{"accession":"Q9NPH5","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9NPH5.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9NPH5/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NPH5"}},"corpus_meta":[{"pmid":"11376945","id":"PMC_11376945","title":"Homologs of gp91phox: cloning and tissue expression of Nox3, Nox4, and Nox5.","date":"2001","source":"Gene","url":"https://pubmed.ncbi.nlm.nih.gov/11376945","citation_count":681,"is_preprint":false},{"pmid":"16135519","id":"PMC_16135519","title":"Nox4 NAD(P)H oxidase mediates hypertrophy and fibronectin expression in the diabetic kidney.","date":"2005","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/16135519","citation_count":434,"is_preprint":false},{"pmid":"19706525","id":"PMC_19706525","title":"Subcellular localization of Nox4 and regulation in diabetes.","date":"2009","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/19706525","citation_count":404,"is_preprint":false},{"pmid":"15706079","id":"PMC_15706079","title":"Expression and localization of NOX2 and NOX4 in primary human endothelial cells.","date":"2005","source":"Antioxidants & redox signaling","url":"https://pubmed.ncbi.nlm.nih.gov/15706079","citation_count":272,"is_preprint":false},{"pmid":"15802177","id":"PMC_15802177","title":"Neuronal expression of the NADPH oxidase NOX4, and its regulation in mouse experimental brain ischemia.","date":"2005","source":"Neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/15802177","citation_count":239,"is_preprint":false},{"pmid":"12842860","id":"PMC_12842860","title":"Nox4 mediates angiotensin II-induced activation of Akt/protein kinase B in mesangial cells.","date":"2003","source":"American journal of physiology. Renal physiology","url":"https://pubmed.ncbi.nlm.nih.gov/12842860","citation_count":230,"is_preprint":false},{"pmid":"19057021","id":"PMC_19057021","title":"Nox4 acts as a switch between differentiation and proliferation in preadipocytes.","date":"2008","source":"Arteriosclerosis, thrombosis, and vascular biology","url":"https://pubmed.ncbi.nlm.nih.gov/19057021","citation_count":210,"is_preprint":false},{"pmid":"16987004","id":"PMC_16987004","title":"NOX2 and NOX4 mediate proliferative response in endothelial cells.","date":"2006","source":"Antioxidants & redox signaling","url":"https://pubmed.ncbi.nlm.nih.gov/16987004","citation_count":207,"is_preprint":false},{"pmid":"28424170","id":"PMC_28424170","title":"ROS-induced ROS release orchestrated by Nox4, Nox2, and mitochondria in VEGF signaling and angiogenesis.","date":"2017","source":"American journal of physiology. Cell physiology","url":"https://pubmed.ncbi.nlm.nih.gov/28424170","citation_count":204,"is_preprint":false},{"pmid":"29051480","id":"PMC_29051480","title":"NOX4 functions as a mitochondrial energetic sensor coupling cancer metabolic reprogramming to drug resistance.","date":"2017","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/29051480","citation_count":174,"is_preprint":false},{"pmid":"21554947","id":"PMC_21554947","title":"Nox4 regulates Nrf2 and glutathione redox in cardiomyocytes in vivo.","date":"2011","source":"Free radical biology & medicine","url":"https://pubmed.ncbi.nlm.nih.gov/21554947","citation_count":151,"is_preprint":false},{"pmid":"26385958","id":"PMC_26385958","title":"The NADPH oxidase Nox4 has anti-atherosclerotic functions.","date":"2015","source":"European heart journal","url":"https://pubmed.ncbi.nlm.nih.gov/26385958","citation_count":146,"is_preprint":false},{"pmid":"22904198","id":"PMC_22904198","title":"The Nox4 inhibitor GKT137831 attenuates hypoxia-induced pulmonary vascular cell proliferation.","date":"2012","source":"American journal of respiratory cell and molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/22904198","citation_count":132,"is_preprint":false},{"pmid":"19061439","id":"PMC_19061439","title":"Identification of structural elements in Nox1 and Nox4 controlling localization and activity.","date":"2009","source":"Antioxidants & redox signaling","url":"https://pubmed.ncbi.nlm.nih.gov/19061439","citation_count":125,"is_preprint":false},{"pmid":"22937798","id":"PMC_22937798","title":"Neuroprotection after stroke by targeting NOX4 as a source of oxidative stress.","date":"2012","source":"Antioxidants & redox signaling","url":"https://pubmed.ncbi.nlm.nih.gov/22937798","citation_count":123,"is_preprint":false},{"pmid":"29793963","id":"PMC_29793963","title":"A NOX4/TRPC6 Pathway in Podocyte Calcium Regulation and Renal Damage in Diabetic Kidney Disease.","date":"2018","source":"Journal of the American Society of Nephrology : JASN","url":"https://pubmed.ncbi.nlm.nih.gov/29793963","citation_count":117,"is_preprint":false},{"pmid":"26742780","id":"PMC_26742780","title":"Targeted redox inhibition of protein phosphatase 1 by Nox4 regulates eIF2α-mediated stress signaling.","date":"2016","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/26742780","citation_count":103,"is_preprint":false},{"pmid":"29969717","id":"PMC_29969717","title":"Nox4 in renal diseases: An update.","date":"2018","source":"Free radical biology & medicine","url":"https://pubmed.ncbi.nlm.nih.gov/29969717","citation_count":101,"is_preprint":false},{"pmid":"25950600","id":"PMC_25950600","title":"The human Nox4: gene, structure, physiological function and pathological significance.","date":"2015","source":"Journal of drug targeting","url":"https://pubmed.ncbi.nlm.nih.gov/25950600","citation_count":90,"is_preprint":false},{"pmid":"35917680","id":"PMC_35917680","title":"TRPM7 channel inhibition attenuates rheumatoid arthritis articular chondrocyte ferroptosis by suppression of the PKCα-NOX4 axis.","date":"2022","source":"Redox biology","url":"https://pubmed.ncbi.nlm.nih.gov/35917680","citation_count":88,"is_preprint":false},{"pmid":"24457954","id":"PMC_24457954","title":"Nox4 and redox signaling mediate TGF-β-induced endothelial cell apoptosis and phenotypic switch.","date":"2014","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/24457954","citation_count":81,"is_preprint":false},{"pmid":"24509161","id":"PMC_24509161","title":"The NADPH oxidase NOX4 inhibits hepatocyte proliferation and liver cancer progression.","date":"2014","source":"Free radical biology & medicine","url":"https://pubmed.ncbi.nlm.nih.gov/24509161","citation_count":79,"is_preprint":false},{"pmid":"33001475","id":"PMC_33001475","title":"Nox4 regulates InsP3 receptor-dependent Ca2+ release into mitochondria to promote cell survival.","date":"2020","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/33001475","citation_count":77,"is_preprint":false},{"pmid":"27525436","id":"PMC_27525436","title":"Tyrosine kinase FYN negatively regulates NOX4 in cardiac remodeling.","date":"2016","source":"The Journal of clinical investigation","url":"https://pubmed.ncbi.nlm.nih.gov/27525436","citation_count":74,"is_preprint":false},{"pmid":"37321060","id":"PMC_37321060","title":"Nox4 as a novel therapeutic target for diabetic vascular complications.","date":"2023","source":"Redox biology","url":"https://pubmed.ncbi.nlm.nih.gov/37321060","citation_count":73,"is_preprint":false},{"pmid":"18474828","id":"PMC_18474828","title":"Nox4 oxidase overexpression specifically decreases endogenous Nox4 mRNA and inhibits angiotensin II-induced adventitial myofibroblast migration.","date":"2008","source":"Hypertension (Dallas, Tex. : 1979)","url":"https://pubmed.ncbi.nlm.nih.gov/18474828","citation_count":73,"is_preprint":false},{"pmid":"29040462","id":"PMC_29040462","title":"Both cardiomyocyte and endothelial cell Nox4 mediate protection against hemodynamic overload-induced remodelling.","date":"2018","source":"Cardiovascular research","url":"https://pubmed.ncbi.nlm.nih.gov/29040462","citation_count":64,"is_preprint":false},{"pmid":"32441762","id":"PMC_32441762","title":"SIRT1-NOX4 signaling axis regulates cancer cachexia.","date":"2020","source":"The Journal of experimental medicine","url":"https://pubmed.ncbi.nlm.nih.gov/32441762","citation_count":62,"is_preprint":false},{"pmid":"15108351","id":"PMC_15108351","title":"Expression of Nox4 in osteoclasts.","date":"2004","source":"Journal of cellular biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/15108351","citation_count":62,"is_preprint":false},{"pmid":"29496628","id":"PMC_29496628","title":"NOX4-driven ROS formation regulates proliferation and apoptosis of gastric cancer cells through the GLI1 pathway.","date":"2018","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/29496628","citation_count":61,"is_preprint":false},{"pmid":"32544088","id":"PMC_32544088","title":"Brd4-p300 inhibition downregulates Nox4 and accelerates lung fibrosis resolution in aged mice.","date":"2020","source":"JCI insight","url":"https://pubmed.ncbi.nlm.nih.gov/32544088","citation_count":60,"is_preprint":false},{"pmid":"24755467","id":"PMC_24755467","title":"NADPH oxidase NOX4 supports renal tumorigenesis by promoting the expression and nuclear accumulation of HIF2α.","date":"2014","source":"Cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/24755467","citation_count":59,"is_preprint":false},{"pmid":"35023280","id":"PMC_35023280","title":"Ferroptosis-related gene NOX4, CHAC1 and HIF1A are valid biomarkers for stomach adenocarcinoma.","date":"2022","source":"Journal of cellular and molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/35023280","citation_count":57,"is_preprint":false},{"pmid":"34438016","id":"PMC_34438016","title":"Backstage players of fibrosis: NOX4, mTOR, HDAC, and S1P; companions of TGF-β.","date":"2021","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/34438016","citation_count":57,"is_preprint":false},{"pmid":"26308771","id":"PMC_26308771","title":"NOX4-driven ROS formation mediates PTP inactivation and cell transformation in FLT3ITD-positive AML cells.","date":"2015","source":"Leukemia","url":"https://pubmed.ncbi.nlm.nih.gov/26308771","citation_count":56,"is_preprint":false},{"pmid":"22328777","id":"PMC_22328777","title":"NOX4 pathway as a source of selective insulin resistance and responsiveness.","date":"2012","source":"Arteriosclerosis, thrombosis, and vascular biology","url":"https://pubmed.ncbi.nlm.nih.gov/22328777","citation_count":55,"is_preprint":false},{"pmid":"33400973","id":"PMC_33400973","title":"Nox4 mediates skeletal muscle metabolic responses to exercise.","date":"2021","source":"Molecular metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/33400973","citation_count":54,"is_preprint":false},{"pmid":"32641980","id":"PMC_32641980","title":"Oleic acid-induced NOX4 is dependent on ANGPTL4 expression to promote human colorectal cancer metastasis.","date":"2020","source":"Theranostics","url":"https://pubmed.ncbi.nlm.nih.gov/32641980","citation_count":54,"is_preprint":false},{"pmid":"32114846","id":"PMC_32114846","title":"Nox (NADPH Oxidase) 1, Nox4, and Nox5 Promote Vascular Permeability and Neovascularization in Retinopathy.","date":"2020","source":"Hypertension (Dallas, Tex. : 1979)","url":"https://pubmed.ncbi.nlm.nih.gov/32114846","citation_count":53,"is_preprint":false},{"pmid":"28916474","id":"PMC_28916474","title":"Nox4 regulates the eNOS uncoupling process in aging endothelial cells.","date":"2017","source":"Free radical biology & medicine","url":"https://pubmed.ncbi.nlm.nih.gov/28916474","citation_count":52,"is_preprint":false},{"pmid":"21965295","id":"PMC_21965295","title":"NOX4 mediates activation of FoxO3a and matrix metalloproteinase-2 expression by urotensin-II.","date":"2011","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/21965295","citation_count":50,"is_preprint":false},{"pmid":"26513738","id":"PMC_26513738","title":"The NADPH Oxidase Nox4 mediates tumour angiogenesis.","date":"2015","source":"Acta physiologica (Oxford, England)","url":"https://pubmed.ncbi.nlm.nih.gov/26513738","citation_count":50,"is_preprint":false},{"pmid":"36738798","id":"PMC_36738798","title":"Osteoporotic bone loss from excess iron accumulation is driven by NOX4-triggered ferroptosis in osteoblasts.","date":"2023","source":"Free radical biology & medicine","url":"https://pubmed.ncbi.nlm.nih.gov/36738798","citation_count":48,"is_preprint":false},{"pmid":"24492492","id":"PMC_24492492","title":"NOX4 regulates autophagy during energy deprivation.","date":"2014","source":"Autophagy","url":"https://pubmed.ncbi.nlm.nih.gov/24492492","citation_count":47,"is_preprint":false},{"pmid":"23678045","id":"PMC_23678045","title":"ADAM17 mediates Nox4 expression and NADPH oxidase activity in the kidney cortex of OVE26 mice.","date":"2013","source":"American journal of physiology. Renal physiology","url":"https://pubmed.ncbi.nlm.nih.gov/23678045","citation_count":45,"is_preprint":false},{"pmid":"26812119","id":"PMC_26812119","title":"Pro-atherogenic role of smooth muscle Nox4-based NADPH oxidase.","date":"2016","source":"Journal of molecular and cellular cardiology","url":"https://pubmed.ncbi.nlm.nih.gov/26812119","citation_count":45,"is_preprint":false},{"pmid":"37871532","id":"PMC_37871532","title":"Cardiomyocyte NOX4 regulates resident macrophage-mediated inflammation and diastolic dysfunction in stress cardiomyopathy.","date":"2023","source":"Redox biology","url":"https://pubmed.ncbi.nlm.nih.gov/37871532","citation_count":44,"is_preprint":false},{"pmid":"8464732","id":"PMC_8464732","title":"Duplicated KOX zinc finger gene clusters flank the centromere of human chromosome 10: evidence for a pericentric inversion during primate evolution.","date":"1993","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/8464732","citation_count":42,"is_preprint":false},{"pmid":"20531308","id":"PMC_20531308","title":"The NADPH oxidases NOX4 and DUOX2 regulate cell cycle entry via a p53-dependent pathway.","date":"2010","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/20531308","citation_count":41,"is_preprint":false},{"pmid":"35617030","id":"PMC_35617030","title":"Breast cancer chemotherapy induces vascular dysfunction and hypertension through a NOX4-dependent mechanism.","date":"2022","source":"The Journal of clinical investigation","url":"https://pubmed.ncbi.nlm.nih.gov/35617030","citation_count":39,"is_preprint":false},{"pmid":"31247506","id":"PMC_31247506","title":"Tubular NOX4 expression decreases in chronic kidney disease but does not modify fibrosis evolution.","date":"2019","source":"Redox biology","url":"https://pubmed.ncbi.nlm.nih.gov/31247506","citation_count":39,"is_preprint":false},{"pmid":"25402870","id":"PMC_25402870","title":"Nox-4 and progressive kidney disease.","date":"2015","source":"Current opinion in nephrology and hypertension","url":"https://pubmed.ncbi.nlm.nih.gov/25402870","citation_count":37,"is_preprint":false},{"pmid":"25652847","id":"PMC_25652847","title":"Nox4 supports proper capillary growth in exercise and retina neo-vascularization.","date":"2015","source":"The Journal of physiology","url":"https://pubmed.ncbi.nlm.nih.gov/25652847","citation_count":37,"is_preprint":false},{"pmid":"26582463","id":"PMC_26582463","title":"Role of smooth muscle Nox4-based NADPH oxidase in neointimal hyperplasia.","date":"2015","source":"Journal of molecular and cellular cardiology","url":"https://pubmed.ncbi.nlm.nih.gov/26582463","citation_count":37,"is_preprint":false},{"pmid":"35269843","id":"PMC_35269843","title":"NADPH Oxidase 4 (NOX4) in Cancer: Linking Redox Signals to Oncogenic Metabolic Adaptation.","date":"2022","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/35269843","citation_count":37,"is_preprint":false},{"pmid":"38060313","id":"PMC_38060313","title":"Mitochondria- and NOX4-dependent antioxidant defense mitigates progression to nonalcoholic steatohepatitis in obesity.","date":"2023","source":"The Journal of clinical investigation","url":"https://pubmed.ncbi.nlm.nih.gov/38060313","citation_count":35,"is_preprint":false},{"pmid":"28594389","id":"PMC_28594389","title":"The Role of NOX4 and TRX2 in Angiogenesis and Their Potential Cross-Talk.","date":"2017","source":"Antioxidants (Basel, Switzerland)","url":"https://pubmed.ncbi.nlm.nih.gov/28594389","citation_count":35,"is_preprint":false},{"pmid":"37288723","id":"PMC_37288723","title":"Metformin ameliorates ferroptosis in cardiac ischemia and reperfusion by reducing NOX4 expression via promoting AMPKα.","date":"2023","source":"Pharmaceutical biology","url":"https://pubmed.ncbi.nlm.nih.gov/37288723","citation_count":34,"is_preprint":false},{"pmid":"29047077","id":"PMC_29047077","title":"Adventitial Fibroblast Nox4 Expression and ROS Signaling in Pulmonary Arterial Hypertension.","date":"2017","source":"Advances in experimental medicine and biology","url":"https://pubmed.ncbi.nlm.nih.gov/29047077","citation_count":34,"is_preprint":false},{"pmid":"35920301","id":"PMC_35920301","title":"The NADPH oxidase NOX4 regulates redox and metabolic homeostasis preventing HCC progression.","date":"2022","source":"Hepatology (Baltimore, Md.)","url":"https://pubmed.ncbi.nlm.nih.gov/35920301","citation_count":34,"is_preprint":false},{"pmid":"34650437","id":"PMC_34650437","title":"Nox4 Promotes RANKL-Induced Autophagy and Osteoclastogenesis via Activating ROS/PERK/eIF-2α/ATF4 Pathway.","date":"2021","source":"Frontiers in pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/34650437","citation_count":34,"is_preprint":false},{"pmid":"32901890","id":"PMC_32901890","title":"Circular RNA NOX4 promotes the development of colorectal cancer via the microRNA‑485‑5p/CKS1B axis.","date":"2020","source":"Oncology reports","url":"https://pubmed.ncbi.nlm.nih.gov/32901890","citation_count":34,"is_preprint":false},{"pmid":"28185955","id":"PMC_28185955","title":"Endothelial Nox4-based NADPH oxidase regulates atherosclerosis via soluble epoxide hydrolase.","date":"2017","source":"Biochimica et biophysica acta. Molecular basis of disease","url":"https://pubmed.ncbi.nlm.nih.gov/28185955","citation_count":34,"is_preprint":false},{"pmid":"30367082","id":"PMC_30367082","title":"NADPH oxidase NOX4 is a glycolytic regulator through mROS-HIF1α axis in thyroid carcinomas.","date":"2018","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/30367082","citation_count":34,"is_preprint":false},{"pmid":"32799394","id":"PMC_32799394","title":"NOX4-dependent regulation of ENaC in hypertension and diabetic kidney disease.","date":"2020","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/32799394","citation_count":33,"is_preprint":false},{"pmid":"35203105","id":"PMC_35203105","title":"NOX4 regulates TGFβ-induced proliferation and self-renewal in glioblastoma stem cells.","date":"2022","source":"Molecular oncology","url":"https://pubmed.ncbi.nlm.nih.gov/35203105","citation_count":32,"is_preprint":false},{"pmid":"34740322","id":"PMC_34740322","title":"NOX4-derived ROS-induced overexpression of FOXM1 regulates aerobic glycolysis in glioblastoma.","date":"2021","source":"BMC cancer","url":"https://pubmed.ncbi.nlm.nih.gov/34740322","citation_count":32,"is_preprint":false},{"pmid":"34435888","id":"PMC_34435888","title":"Role of Nox4 in High Calcium-Induced Renal Oxidative Stress Damage and Crystal Deposition.","date":"2021","source":"Antioxidants & redox signaling","url":"https://pubmed.ncbi.nlm.nih.gov/34435888","citation_count":31,"is_preprint":false},{"pmid":"37061215","id":"PMC_37061215","title":"Targeting benign prostate hyperplasia treatments: AR/TGF-β/NOX4 inhibition by apocynin suppresses inflammation and proliferation.","date":"2023","source":"Journal of advanced research","url":"https://pubmed.ncbi.nlm.nih.gov/37061215","citation_count":29,"is_preprint":false},{"pmid":"34922273","id":"PMC_34922273","title":"Implication of type 4 NADPH oxidase (NOX4) in tauopathy.","date":"2021","source":"Redox biology","url":"https://pubmed.ncbi.nlm.nih.gov/34922273","citation_count":29,"is_preprint":false},{"pmid":"34209278","id":"PMC_34209278","title":"NOX4 Signaling Mediates Cancer Development and Therapeutic Resistance through HER3 in Ovarian Cancer Cells.","date":"2021","source":"Cells","url":"https://pubmed.ncbi.nlm.nih.gov/34209278","citation_count":29,"is_preprint":false},{"pmid":"38844470","id":"PMC_38844470","title":"HOXD10 attenuates renal fibrosis by inhibiting NOX4-induced ferroptosis.","date":"2024","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/38844470","citation_count":28,"is_preprint":false},{"pmid":"32666016","id":"PMC_32666016","title":"CSF-1 in Osteocytes Inhibits Nox4-mediated Oxidative Stress and Promotes Normal Bone Homeostasis.","date":"2019","source":"JBMR plus","url":"https://pubmed.ncbi.nlm.nih.gov/32666016","citation_count":28,"is_preprint":false},{"pmid":"34930338","id":"PMC_34930338","title":"NOX4: a potential therapeutic target for pancreatic cancer and its mechanism.","date":"2021","source":"Journal of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/34930338","citation_count":27,"is_preprint":false},{"pmid":"29986678","id":"PMC_29986678","title":"NOX4 expression and distal arteriolar remodeling correlate with pulmonary hypertension in COPD.","date":"2018","source":"BMC pulmonary medicine","url":"https://pubmed.ncbi.nlm.nih.gov/29986678","citation_count":27,"is_preprint":false},{"pmid":"29113787","id":"PMC_29113787","title":"Nox4 genetic inhibition in experimental hypertension and metabolic syndrome.","date":"2017","source":"Archives of cardiovascular diseases","url":"https://pubmed.ncbi.nlm.nih.gov/29113787","citation_count":27,"is_preprint":false},{"pmid":"24719867","id":"PMC_24719867","title":"Nuclear Nox4-derived reactive oxygen species in myelodysplastic syndromes.","date":"2014","source":"BioMed research international","url":"https://pubmed.ncbi.nlm.nih.gov/24719867","citation_count":27,"is_preprint":false},{"pmid":"38354631","id":"PMC_38354631","title":"DRD4 alleviates acute kidney injury by suppressing ISG15/NOX4 axis-associated oxidative stress.","date":"2024","source":"Redox biology","url":"https://pubmed.ncbi.nlm.nih.gov/38354631","citation_count":26,"is_preprint":false},{"pmid":"32780450","id":"PMC_32780450","title":"The role of NOX4 in pulmonary diseases.","date":"2020","source":"Journal of cellular physiology","url":"https://pubmed.ncbi.nlm.nih.gov/32780450","citation_count":26,"is_preprint":false},{"pmid":"27110716","id":"PMC_27110716","title":"Nox4 and Duox1/2 Mediate Redox Activation of Mesenchymal Cell Migration by PDGF.","date":"2016","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/27110716","citation_count":26,"is_preprint":false},{"pmid":"38483540","id":"PMC_38483540","title":"Myricetin Induces Ferroptosis and Inhibits Gastric Cancer Progression by Targeting NOX4.","date":"2024","source":"Journal of agricultural and food chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/38483540","citation_count":26,"is_preprint":false},{"pmid":"25410908","id":"PMC_25410908","title":"Nox4-mediated cell signaling regulates differentiation and survival of neural crest stem cells.","date":"2014","source":"Molecules and cells","url":"https://pubmed.ncbi.nlm.nih.gov/25410908","citation_count":26,"is_preprint":false},{"pmid":"37017270","id":"PMC_37017270","title":"Ginsenoside Rg1 treatment alleviates renal fibrosis by inhibiting the NOX4-MAPK pathway in T2DM mice.","date":"2023","source":"Renal failure","url":"https://pubmed.ncbi.nlm.nih.gov/37017270","citation_count":26,"is_preprint":false},{"pmid":"33836590","id":"PMC_33836590","title":"Genetic deletion of Nox4 enhances cancerogen-induced formation of solid tumors.","date":"2021","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/33836590","citation_count":25,"is_preprint":false},{"pmid":"24868315","id":"PMC_24868315","title":"Lentivirus-mediated Nox4 shRNA invasion and angiogenesis and enhances radiosensitivity in human glioblastoma.","date":"2014","source":"Oxidative medicine and cellular longevity","url":"https://pubmed.ncbi.nlm.nih.gov/24868315","citation_count":24,"is_preprint":false},{"pmid":"30637800","id":"PMC_30637800","title":"Downregulated NOX4 underlies a novel inhibitory role of microRNA-137 in prostate cancer.","date":"2019","source":"Journal of cellular biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/30637800","citation_count":24,"is_preprint":false},{"pmid":"28099519","id":"PMC_28099519","title":"CRISPR-Cas9 Mediated NOX4 Knockout Inhibits Cell Proliferation and Invasion in HeLa Cells.","date":"2017","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/28099519","citation_count":22,"is_preprint":false},{"pmid":"37119283","id":"PMC_37119283","title":"TRIM-containing 44 aggravates cardiac hypertrophy via TLR4/NOX4-induced ferroptosis.","date":"2023","source":"Journal of molecular medicine (Berlin, Germany)","url":"https://pubmed.ncbi.nlm.nih.gov/37119283","citation_count":22,"is_preprint":false},{"pmid":"8262519","id":"PMC_8262519","title":"Chromosomal localization of 9 KOX zinc finger genes: physical linkages suggest clustering of KOX genes on chromosomes 12, 16, and 19.","date":"1993","source":"Human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/8262519","citation_count":22,"is_preprint":false},{"pmid":"33493901","id":"PMC_33493901","title":"NADPH oxidase 4 (Nox4) deletion accelerates liver regeneration in mice.","date":"2020","source":"Redox biology","url":"https://pubmed.ncbi.nlm.nih.gov/33493901","citation_count":22,"is_preprint":false},{"pmid":"31419440","id":"PMC_31419440","title":"Tachycardia-induced CD44/NOX4 signaling is involved in the development of atrial remodeling.","date":"2019","source":"Journal of molecular and cellular cardiology","url":"https://pubmed.ncbi.nlm.nih.gov/31419440","citation_count":22,"is_preprint":false},{"pmid":"32818796","id":"PMC_32818796","title":"Oxidation of HDAC4 by Nox4-derived H2O2 maintains tube formation by endothelial cells.","date":"2020","source":"Redox biology","url":"https://pubmed.ncbi.nlm.nih.gov/32818796","citation_count":22,"is_preprint":false},{"pmid":"35711428","id":"PMC_35711428","title":"MicroRNA-182-5p Attenuates Asthmatic Airway Inflammation by Targeting NOX4.","date":"2022","source":"Frontiers in immunology","url":"https://pubmed.ncbi.nlm.nih.gov/35711428","citation_count":21,"is_preprint":false},{"pmid":"34656824","id":"PMC_34656824","title":"Cooperation between CYB5R3 and NOX4 via coenzyme Q mitigates endothelial inflammation.","date":"2021","source":"Redox biology","url":"https://pubmed.ncbi.nlm.nih.gov/34656824","citation_count":21,"is_preprint":false},{"pmid":"23832701","id":"PMC_23832701","title":"Nox2 and Nox4 influence neonatal c-kit(+) cardiac precursor cell status and differentiation.","date":"2013","source":"American journal of physiology. Heart and circulatory physiology","url":"https://pubmed.ncbi.nlm.nih.gov/23832701","citation_count":21,"is_preprint":false},{"pmid":"34326272","id":"PMC_34326272","title":"Obesity increases neuropathic pain via the AMPK-ERK-NOX4 pathway in rats.","date":"2021","source":"Aging","url":"https://pubmed.ncbi.nlm.nih.gov/34326272","citation_count":21,"is_preprint":false},{"pmid":"36302499","id":"PMC_36302499","title":"Astragaloside IV ameliorates sepsis-induced myocardial dysfunction by regulating NOX4/JNK/BAX pathway.","date":"2022","source":"Life sciences","url":"https://pubmed.ncbi.nlm.nih.gov/36302499","citation_count":21,"is_preprint":false},{"pmid":"37432297","id":"PMC_37432297","title":"Diosgenin Inhibits ROS Generation by Modulating NOX4 and Mitochondrial Respiratory Chain and Suppresses Apoptosis in Diabetic Nephropathy.","date":"2023","source":"Nutrients","url":"https://pubmed.ncbi.nlm.nih.gov/37432297","citation_count":21,"is_preprint":false},{"pmid":"36040503","id":"PMC_36040503","title":"Intestinal microbiota regulates diabetes and cancer progression by IL-1β and NOX4 dependent signaling cascades.","date":"2022","source":"Cellular and molecular life sciences : CMLS","url":"https://pubmed.ncbi.nlm.nih.gov/36040503","citation_count":21,"is_preprint":false},{"pmid":"38993139","id":"PMC_38993139","title":"NOX4 exacerbates Parkinson's disease pathology by promoting neuronal ferroptosis and neuroinflammation.","date":"2024","source":"Neural regeneration research","url":"https://pubmed.ncbi.nlm.nih.gov/38993139","citation_count":20,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":53883,"output_tokens":9738,"usd":0.15386,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":20016,"output_tokens":7928,"usd":0.14914,"stage2_stop_reason":"end_turn"},"total_usd":0.303,"stage1_batch_id":"msgbatch_013a8chtv3AWyTqv4z4sEF4C","stage2_batch_id":"msgbatch_01AHsuW63vinsvuTgzmQ3n62","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2001,\n      \"finding\": \"NOX4 encodes a ~65 kDa protein homologous to gp91phox (the catalytic subunit of the phagocyte NADPH oxidase), containing 5-6 conserved predicted transmembrane alpha-helices with putative heme-binding regions plus a flavoprotein homology domain with predicted FAD and NADPH binding sites, establishing it as an NADPH-dependent ROS-generating enzyme.\",\n      \"method\": \"cDNA cloning, sequence analysis, tissue expression (Northern blot/RT-PCR)\",\n      \"journal\": \"Gene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — structural inference from sequence homology confirmed across multiple tissues; functional enzymatic activity not yet directly reconstituted in this paper but domain architecture well established\",\n      \"pmids\": [\"11376945\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Nox4-based NADPH oxidase, regulated upstream by Rac1 and arachidonic acid, mediates angiotensin II-induced ROS generation and downstream Akt/PKB activation and protein synthesis in mesangial cells. Antisense Nox4 knockdown abolished ANG II-induced NADPH oxidase activity and Akt/PKB activation, and dominant-negative Rac1 blocked Nox4-dependent ROS.\",\n      \"method\": \"Antisense oligonucleotide knockdown, dominant-negative Rac1 transfection, NADPH oxidase activity assay, ROS measurement, Akt phosphorylation assays, protein synthesis assay\",\n      \"journal\": \"American journal of physiology. Renal physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods (AS knockdown, DN-Rac1, functional ROS/kinase assays) in single lab\",\n      \"pmids\": [\"12842860\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Nox4 is the major source of NADPH-dependent ROS in the diabetic kidney; antisense-mediated Nox4 knockdown reduced NADPH oxidase activity in renal cortical/glomerular homogenates, blocked glucose-induced ROS in isolated glomeruli, and reduced downstream Akt/PKB and ERK1/2 activation, renal hypertrophy, and fibronectin expression.\",\n      \"method\": \"Antisense oligonucleotide administration in vivo (osmotic minipump), NADPH oxidase activity assay, ROS measurement from intact glomeruli, immunoblotting for Akt/ERK, histology for hypertrophy, fibronectin immunostaining\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods in vivo and in vitro, single lab\",\n      \"pmids\": [\"16135519\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"NOX4-GFP fusion protein localizes to the endoplasmic reticulum in human endothelial cells (HUVECs), as shown by co-staining with an ER marker; distribution did not overlap with lysosomes, Weibel-Palade bodies, or mitochondria.\",\n      \"method\": \"Fluorescence confocal microscopy with ER, lysosomal, and mitochondrial markers; NOX4-GFP overexpression\",\n      \"journal\": \"Antioxidants & redox signaling\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — direct localization by confocal with multiple organelle markers, single lab\",\n      \"pmids\": [\"15706079\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"NOX2 and NOX4 co-localize with the ER marker calreticulin in endothelial cells and interact with p22phox, as demonstrated by bimolecular fluorescence complementation; both NOX2 and NOX4 contribute equally to endothelial ROS production and proliferation.\",\n      \"method\": \"Bimolecular fluorescence complementation (BiFC) for NOX4-p22phox interaction, co-localization with calreticulin by immunofluorescence, siRNA knockdown, ROS measurement\",\n      \"journal\": \"Antioxidants & redox signaling\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — protein–protein interaction confirmed by BiFC plus functional ROS/proliferation readouts, single lab\",\n      \"pmids\": [\"16987004\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Nox4 controls the switch between insulin-induced proliferation and differentiation in preadipocytes by regulating MAP kinase phosphatase-1 (MKP-1) expression; Nox4 siRNA reduced ROS and MKP-1, de-repressed ERK1/2, which phosphorylated IRS-1 at Ser612 to block differentiation and promote proliferation.\",\n      \"method\": \"siRNA knockdown, Nox4 overexpression, ERK1/2 phosphorylation assays, MKP-1 expression analysis, IRS-1 phosphorylation, proliferation/differentiation assays\",\n      \"journal\": \"Arteriosclerosis, thrombosis, and vascular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal siRNA/overexpression with mechanistic pathway dissection, single lab\",\n      \"pmids\": [\"19057021\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Nox4 localizes to mitochondria in mesangial cells and kidney cortex; siRNA-mediated knockdown of Nox4 significantly reduces NADPH oxidase activity in purified mitochondria and blocks glucose-induced mitochondrial superoxide generation. Mitochondrial Nox4 expression is increased in diabetic kidney cortex.\",\n      \"method\": \"Subcellular fractionation, immunoblotting of mitochondrial fractions, immunofluorescence confocal with Mitotracker, MitoProt prediction, siRNA knockdown, NADPH oxidase activity assay in purified mitochondria, in vivo diabetic rat model\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal localization methods (fractionation, confocal, computational) plus functional knockdown in purified organelle, single lab\",\n      \"pmids\": [\"19706525\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Structural elements in Nox4 determine both its subcellular localization to the ER and the type of ROS released (H2O2 extracellularly rather than O2-). The cytosolic tail of Nox4 confers constitutive activity (independent of cytosolic subunits), the N-terminal region determines ER localization, and the N-terminal part of Nox1 (but not Nox4) is cleaved. Replacing the Nox1 N-terminus with the Nox4 signal peptide redirected Nox1 from plasma membrane to vesicular structures and switched ROS from O2- to H2O2.\",\n      \"method\": \"Chimeric Nox1/Nox4 constructs expressed in HEK293 cells, TIRF microscopy, ROS type measurement, Myc-tagged Nox constructs, co-expression studies\",\n      \"journal\": \"Antioxidants & redox signaling\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — reconstitution with chimeric constructs and domain-swap mutagenesis directly mapping structural determinants of localization and ROS type, multiple orthogonal readouts\",\n      \"pmids\": [\"19061439\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"NOX4 is required for PDGF-induced cell cycle entry in normal human fibroblasts; NOX4 knockdown did not block cyclin D1 upregulation but reduced ERK1 phosphorylation hours after stimulation and increased p53 and p21 levels. Co-knockdown of NOX4 with p53 or p21 rescued Rb phosphorylation, indicating NOX4 promotes cell cycle entry by suppressing a p53/p21 checkpoint.\",\n      \"method\": \"siRNA screen, Rb phosphorylation assay, cyclin D1/p53/p21 immunoblotting, co-knockdown epistasis\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — epistasis co-knockdown experiments in human fibroblasts, single lab, multiple pathway markers\",\n      \"pmids\": [\"20531308\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Nox4 overexpression in cardiomyocytes in vivo activates the Nrf2 transcriptional pathway, leading to increased expression of antioxidant/detoxifying genes and elevated GSH and reduced:oxidized GSH ratio; these effects are abolished in Nrf2-null mice, demonstrating that Nox4-derived ROS activates Nrf2-dependent antioxidant defense.\",\n      \"method\": \"Transgenic mouse overexpression, microarray transcriptomics, Q-PCR, glutathione measurement, Nrf2 knockout genetic background\",\n      \"journal\": \"Free radical biology & medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo transgenic plus genetic epistasis with Nrf2-null, single lab\",\n      \"pmids\": [\"21554947\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"NOX4 silencing in VHL-deficient renal carcinoma cells abrogates nuclear accumulation of HIF2α and blocks cell branching, invasion, colony formation, and xenograft growth, demonstrating that NOX4-derived ROS is required for HIF2α nuclear localization and renal tumorigenesis.\",\n      \"method\": \"siRNA knockdown, ROS scavengers (TEMPOL, MnSOD/catalase overexpression), nuclear fractionation for HIF2α, in vitro invasion/colony assays, murine xenograft model\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal ROS manipulation methods (siRNA, scavengers, antioxidant overexpression) with in vivo xenograft validation, single lab\",\n      \"pmids\": [\"24755467\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"NOX4 activity is increased in the ER (but not mitochondria) of cardiomyocytes during energy deprivation; NOX4-derived ROS activates the PERK-eIF2α-ATF4 pathway to induce autophagy, which preserves cellular energy and limits cell death.\",\n      \"method\": \"Subcellular fractionation, NOX4 knockdown/knockout, ER-specific ROS measurement, PERK/eIF2α/ATF4 pathway activation assays, autophagy flux measurement, cardiomyocyte viability\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — organelle-specific activity localization plus pathway knockdown epistasis, single lab\",\n      \"pmids\": [\"24492492\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"FLT3ITD-driven leukemic transformation elevates NOX4 expression via STAT5-mediated activation of the NOX4 promoter; NOX4-derived ROS inactivates protein-tyrosine phosphatase DEP-1/PTPRJ, sustaining FLT3ITD signaling. Nox4 knockout hematopoietic progenitors are refractory to FLT3ITD transformation in vitro, and NOX4 downregulation attenuates myeloproliferative disease in vivo.\",\n      \"method\": \"NOX4 mRNA/protein quantification in FLT3ITD cells, STAT5 ChIP on NOX4 promoter, siRNA/knockout, DEP-1 PTP activity assay, ROS measurement, in vitro transformation assay, in vivo mouse disease models\",\n      \"journal\": \"Leukemia\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — transcriptional mechanism (STAT5→NOX4 promoter), enzymatic PTP activity assay, in vitro and in vivo rescue experiments across multiple labs/models\",\n      \"pmids\": [\"26308771\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Nox4 regulates eIF2α-mediated stress signaling by binding to the PP1-targeting subunit GADD34 at the ER and inhibiting PP1 phosphatase activity through oxidation of its metal center (not thiol oxidation), thereby sustaining eIF2α phosphorylation and ATF4 levels. This is spatially confined to the ER and does not affect PP1 targets at other locations.\",\n      \"method\": \"Co-immunoprecipitation of Nox4 with GADD34, PP1 activity assays with metal center vs. thiol oxidation characterization, eIF2α phosphorylation/ATF4 immunoblotting, ER localization, genetic knockdown/overexpression, in vivo heart ischemia-reperfusion and acute kidney injury models\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct biochemical enzymatic inhibition assay with mechanistic distinction (metal center vs. thiol), protein–protein interaction by Co-IP, in vivo disease models, multiple orthogonal methods\",\n      \"pmids\": [\"26742780\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"FYN tyrosine kinase directly interacts with the C-terminal domain of NOX4 and phosphorylates it at tyrosine 566, negatively regulating NOX4-induced O2- production and apoptosis in cardiomyocytes. FYN and NOX4 co-localize in perinuclear mitochondria, ER, and nuclear fractions. FYN-deficient mice have exacerbated cardiac hypertrophy with increased ROS, rescued by Nox4 deletion.\",\n      \"method\": \"Co-immunoprecipitation, co-localization imaging, site-directed mutagenesis (Y566), NOX4 activity assay, Nox4-/- genetic rescue of FYN-/- cardiac phenotype, transverse aortic constriction model, human failing heart analysis\",\n      \"journal\": \"The Journal of clinical investigation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct phosphorylation site identified by mutagenesis, Co-IP, in vivo genetic epistasis rescue, multiple orthogonal methods\",\n      \"pmids\": [\"27525436\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"In VEGF-stimulated endothelial cells, Nox4-derived H2O2 activates Nox2, which promotes mitochondrial ROS production via S36 phosphorylation of p66Shc; this Nox4/Nox2/pSer36-p66Shc/mtROS feed-forward axis drives sustained VEGFR2 phosphorylation, EC migration, and proliferation (angiogenesis).\",\n      \"method\": \"Cytosol/mitochondria-targeted RoGFP biosensors with real-time imaging, Nox4/Nox2 siRNA, mitochondria-targeted catalase overexpression, Nox4 overexpression, p66Shc(S36A) mutant, VEGFR2 tyrosine phosphorylation assay, EC migration/proliferation assays\",\n      \"journal\": \"American journal of physiology. Cell physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — RoGFP real-time organelle-specific ROS imaging, genetic epistasis (double knockdown, mutant p66Shc), multiple orthogonal functional assays, single lab with rigorous controls\",\n      \"pmids\": [\"28424170\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"NOX4 contains an ATP-binding motif; ATP directly binds and negatively regulates NOX4 activity. NOX4 localizes to the inner mitochondrial membrane, and subcellular redistribution of ATP from mitochondria acts as an allosteric switch to activate NOX4. NOX4-derived ROS inhibits PCAF-dependent acetylation and lysosomal degradation of PKM2.\",\n      \"method\": \"ATP-binding assay (identification of ATP-binding motif), subcellular fractionation for inner mitochondrial membrane localization, NOX4 activity assays with ATP titration, PKM2 acetylation/ubiquitination assays, PCAF interaction, NOX4 silencing in xenograft models\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct biochemical binding assay identifying novel allosteric ATP regulation, enzymatic activity measurement, subcellular localization, and mechanistic PKM2 downstream pathway, multiple orthogonal methods\",\n      \"pmids\": [\"29051480\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"NOX4-derived H2O2 in podocytes activates TRPC6-dependent calcium influx, contributing to podocyte damage in diabetic kidney disease; SSNox4-/- rats show lower basal intracellular Ca2+ in podocytes and less DKD-associated damage, and H2O2-stimulated TRPC-dependent calcium influx is blunted in Trpc6-knockout podocytes.\",\n      \"method\": \"Nox4 knockout rat (SSNox4-/-), TRPC6/TRPC5/6 knockout mice, H2O2 stimulation, live calcium imaging, electrophysiology patch-clamp, biosensor measurements, electron microscopy\",\n      \"journal\": \"Journal of the American Society of Nephrology : JASN\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple knockout models, direct calcium imaging, electrophysiology, and electron microscopy across species, multiple orthogonal methods\",\n      \"pmids\": [\"29793963\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"CD44 directly associates with NOX4 in tachypaced atrial myocytes and atrial fibrillation patient tissues; blocking HAS/HA/CD44 signaling attenuates tachypacing-induced NOX4 expression, oxidative stress, and Ca2+-handling abnormalities (ox-CaMKII/p-RyR2).\",\n      \"method\": \"Co-immunoprecipitation of CD44 with NOX4, CD44-/- mice, anti-CD44 blocking antibody, Ca2+ spark measurement, tachypacing model in vitro and ex vivo, AF patient tissue analysis\",\n      \"journal\": \"Journal of molecular and cellular cardiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP demonstrating direct CD44-NOX4 interaction plus genetic/pharmacological epistasis and functional Ca2+ imaging, single lab\",\n      \"pmids\": [\"31419440\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Nox4 upregulated at ER-mitochondria contact sites (MAMs) during stress inhibits InsP3 receptor-mediated Ca2+ transfer from ER to mitochondria by augmenting Akt-dependent phosphorylation of InsP3R, thereby reducing mitochondrial permeability transition and necrosis; in ischemia-reperfusion, Nox4 limits myocardial infarct size through this mechanism.\",\n      \"method\": \"MAM fractionation, Nox4 overexpression/knockout, InsP3R phosphorylation assays, mitochondrial Ca2+ measurement, mPT assay, cardiac ischemia-reperfusion model, cardiomyocyte/neuron stress models\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — mechanistic dissection at MAM with organelle-specific fractionation, InsP3R phosphorylation, Ca2+ flux and mPT measurements, in vivo cardiac model, multiple orthogonal approaches\",\n      \"pmids\": [\"33001475\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Nox4 is required for exercise-induced expression of metabolic genes (Ucp3, Hk2, Pdk4) in skeletal muscle; global and endothelial-specific Nox4 deletion impairs glucose and fatty acid oxidation after acute exercise, revealing an endothelium-to-skeletal muscle cross-talk mediated by Nox4-derived H2O2.\",\n      \"method\": \"Global and endothelial-specific Nox4 KO mice, 14C-labeled substrate oxidation assays ex vivo, qPCR/immunoblotting for metabolic genes, chronic exercise regimen with time-to-exhaustion measurement, catalase transgenic mice\",\n      \"journal\": \"Molecular metabolism\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — cell-type-specific KO models with functional metabolic assays, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"33400973\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"SIRT1 loss in cachectic muscle induces NF-κB signaling that upregulates FOXO transcription factors and NOX4 expression; skeletal muscle-specific Nox4 knockout or pharmacological NOX4 blockade abrogates tumor-induced cachexia in mice, placing NOX4 downstream of the SIRT1/NF-κB/FOXO axis in muscle wasting.\",\n      \"method\": \"RNA-seq, Nox4 muscle-specific KO mice, pharmacological NOX4 inhibition, SIRT1 reconstitution, co-culture and in vivo cancer cachexia models\",\n      \"journal\": \"The Journal of experimental medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — tissue-specific KO with in vivo rescue plus transcriptomic pathway mapping, single lab\",\n      \"pmids\": [\"32441762\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Nox4 overexpression induces oxidation of HDAC4 in HEK293 cells and endothelial cells; Nox4-derived H2O2 increases HDAC4 phosphorylation at Ser632 and disrupts the HDAC4/Mef2A complex, de-repressing Mef2A and enabling proper endothelial tube formation. A redox-insensitive HDAC4 mutant blocks tube formation, while a redox-dead Nox4 mutant fails to rescue it.\",\n      \"method\": \"Tetracycline-inducible Nox4 overexpression in HEK293, HDAC4 oxidation assay, HDAC4/Mef2A co-immunoprecipitation, Ser632 phosphorylation analysis, redox-insensitive HDAC4 mutant, redox-dead Nox4 mutant, endothelial tube formation assay\",\n      \"journal\": \"Redox biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct substrate oxidation assay, protein complex disruption by Co-IP, structure-function mutagenesis of both Nox4 and HDAC4, functional tube formation readout\",\n      \"pmids\": [\"32818796\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Nox4 promotes RANKL-induced autophagy and osteoclastogenesis by stimulating non-mitochondrial ROS production that activates the PERK/eIF-2α/ATF4 unfolded protein response pathway; inhibition of Nox4 or PERK/eIF-2α/ATF4 or ROS scavenging similarly blocks autophagy and osteoclastogenesis.\",\n      \"method\": \"Nox4 inhibitor (5-O-methyl quercetin), Nox4 shRNA knockdown, ROS scavenger (NAC), PERK inhibitor (GSK2606414), autophagy markers, osteoclastogenesis assays, pathway epistasis\",\n      \"journal\": \"Frontiers in pharmacology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic and pharmacological epistasis with multiple nodes of PERK/eIF-2α/ATF4 pathway, single lab\",\n      \"pmids\": [\"34650437\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Neuronal NOX4 promotes pathological tau accumulation by impairing autophagy-lysosomal pathway flux; global Nox4 knockout and neuronal Nox4 knockdown in mice reduced accumulation of hyperphosphorylated tau, improved macroautophagy flux, reduced neurotoxicity, and prevented cognitive decline in a tauopathy model.\",\n      \"method\": \"Global Nox4 KO mice, neuronal-targeted AAV-mediated Nox4 knockdown, humanized tauopathy mouse model (AAV-TauP301L), tau immunohistochemistry, autophagy flux assays, behavioral testing\",\n      \"journal\": \"Redox biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — neuronal-specific KD with mechanistic autophagy pathway readout and in vivo behavioral rescue, single lab\",\n      \"pmids\": [\"34922273\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"NOX4 deletion in mice promotes cancerogen-induced tumor formation by reducing nuclear PP2A abundance; NOX4-derived H2O2 continuously oxidizes AKT, trapping PP2A in the cytosol, which maintains γH2AX (phospho-H2AX) levels for DNA damage recognition. Without Nox4, PP2A translocates to the nucleus, dephosphorylates γH2AX, impairing DNA damage recognition and simultaneously increasing AKT-driven proliferation.\",\n      \"method\": \"Nox4 KO mice, carcinogen-induced tumor models, AKT oxidation assay, PP2A subcellular fractionation, γH2AX immunostaining/immunoblotting, PP2A activity assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct AKT oxidation assay, PP2A subcellular localization by fractionation, γH2AX functional DNA damage readout, in vivo genetic KO with two carcinogen models\",\n      \"pmids\": [\"33836590\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"CYB5R3 localizes to the mitochondrial outer membrane and directly interacts with NOX4; CYB5R3 activity and membrane translocation are required for optimal NOX4-dependent H2O2 generation via coenzyme Q (CoQ). Cyb5r3 knockdown reduces total H2O2 but increases mitochondrial O2•-, and cells lacking the CoQ-synthesizing enzyme COQ6 show decreased NOX4-derived H2O2.\",\n      \"method\": \"APEX2-based electron microscopy, proximity biotinylation, proximity ligation assay, Co-IP, CYB5R3 activity mutants, COQ6 knockdown, mitochondrial ROS species measurement, endothelium-specific Cyb5r3 KO mice\",\n      \"journal\": \"Redox biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple independent protein interaction methods (APEX2-EM, proximity biotinylation, PLA, Co-IP), organelle localization, structure-function mutants, and CoQ dependency established\",\n      \"pmids\": [\"34656824\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"NOX4 locus contains iron-response element-like (IRE-like) sequences bound and repressed by iron regulatory protein 1 (IRP1); excess iron dissociates IRP1 from these sequences, activating NOX4 transcription, which increases lipid peroxides and causes ferroptosis-associated mitochondrial dysfunction in osteoblasts.\",\n      \"method\": \"IRP1 binding assays on NOX4 promoter (IRE-like sequences), NOX4 expression analysis with iron loading, ferroptosis markers (lipid peroxides, MDA), mitochondrial morphology/function assays, ferroptosis inhibitor ferrostatin-1 and iron chelator DFO in Hepc1-/- mice\",\n      \"journal\": \"Free radical biology & medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — IRP1-IRE-like sequence binding characterization on NOX4 promoter plus in vivo genetic/pharmacological validation, single lab\",\n      \"pmids\": [\"36738798\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"DRD4 reduces NOX4 expression via suppression of ISG15, which ISGylates NOX4 and stabilizes it; when DRD4 is active, ISG15 levels fall, NOX4 ISGylation decreases, ubiquitination of NOX4 increases, and NOX4 is degraded via the ubiquitin-proteasome pathway, reducing oxidative stress in acute kidney injury.\",\n      \"method\": \"Transcriptome sequencing, ISG15 knockdown/overexpression, Co-IP for NOX4 ISGylation and ubiquitination, NOX4 protein stability assays, DRD4 KO/overexpression in AKI models (IRI and cisplatin), oxidative stress measurement\",\n      \"journal\": \"Redox biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biochemical ISGylation/ubiquitination assays for NOX4 stability plus in vivo genetic validation, single lab\",\n      \"pmids\": [\"38354631\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"HOXD10 directly binds to the NOX4 promoter (confirmed by ChIP and dual-luciferase assay) and represses its transcription; HOXD10 overexpression attenuates TGF-β1-induced ferroptosis and renal fibrosis by reducing NOX4 expression and downstream ROS/lipid peroxide accumulation. HOXD10 is epigenetically silenced by hypermethylation in TGF-β1-treated cells.\",\n      \"method\": \"ChIP analysis, dual-luciferase reporter assay, HOXD10 overexpression (AAV in vivo), NOX4 expression assay, ferroptosis markers, bisulfite sequencing PCR for methylation, UUO fibrosis model\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — direct promoter binding confirmed by ChIP and luciferase, in vivo rescue, but single lab\",\n      \"pmids\": [\"38844470\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"NOX4 interacts with activated PKCα (protein kinase C alpha) to promote ferroptosis of dopaminergic neurons; NOX4 inhibition reduced lipid peroxidation, iron accumulation, and astrocytic lipocalin-2 expression (reducing neuroinflammation) in a Parkinson's disease MPTP model. ATF3 transcriptionally increases NOX4 expression in dopaminergic neurons and astrocytes.\",\n      \"method\": \"Co-immunoprecipitation of NOX4 with PKCα, NOX4 inhibitor in MPTP mouse model, lipid peroxidation and iron assays, behavioral tests, lipocalin-2 immunostaining, ATF3 transcriptional analysis\",\n      \"journal\": \"Neural regeneration research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP for NOX4-PKCα interaction plus in vivo NOX4 inhibitor with multiple mechanistic readouts, single lab\",\n      \"pmids\": [\"38993139\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"NOX4 (but not NOX2) activity and protein levels increase specifically in the ER of cardiomyocytes during energy deprivation; this ER-localized NOX4-derived ROS activates the PERK-eIF2α-ATF4 autophagy pathway, which is a critical adaptive response to energy stress.\",\n      \"method\": \"Organelle-specific NOX4 activity measurements (ER vs. mitochondria fractions), NOX4 genetic deletion, PERK/eIF2α/ATF4 pathway immunoblotting, autophagy markers, cardiomyocyte energy deprivation model\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — organelle-resolved biochemistry with genetic KO and pathway epistasis, single lab\",\n      \"pmids\": [\"24492492\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Co-transfection of Nox4 with p22phox in osteoclasts enhances superoxide production and increases expression of cathepsin K and TRAP, with JNK activation and NF-κB inhibition, indicating p22phox is a necessary cofactor for Nox4 activity in osteoclasts.\",\n      \"method\": \"Nox4/p22phox co-transfection, superoxide assay, cathepsin K/TRAP expression, JNK/NF-κB signaling assays\",\n      \"journal\": \"Journal of cellular biochemistry\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single co-transfection experiment, single lab, no direct Nox4-p22phox interaction assay\",\n      \"pmids\": [\"15108351\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"NOX4 produces H2O2 constitutively rather than superoxide (O2-), making it incapable of scavenging NO; Nox4 knockout mice on an ApoE-/- background develop increased atherosclerosis, and endothelial-specific (but not macrophage-specific) Nox4 deletion increases macrophage adhesion to endothelium, demonstrating an anti-atherosclerotic endothelial function of Nox4.\",\n      \"method\": \"Tamoxifen-inducible Nox4 KO crossed with ApoE-/- mice, partial carotid artery ligation model, atherosclerosis quantification, cell-type-specific KO, macrophage adhesion assay\",\n      \"journal\": \"European heart journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — cell-type-specific genetic KO with mechanistic gene expression analysis, in vivo atherosclerosis models, single lab\",\n      \"pmids\": [\"26385958\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"NOX4 is a constitutively active, NADPH-dependent oxidase that produces H2O2 (rather than superoxide) primarily at the endoplasmic reticulum and ER-mitochondria contact sites (MAMs), where it mediates spatially confined redox signaling: it requires p22phox and, at the mitochondrial outer membrane, CYB5R3/coenzyme Q for full H2O2 output; its activity is allosterically inhibited by direct ATP binding and negatively regulated by FYN kinase-mediated phosphorylation of Y566; structurally, the N-terminal region determines ER localization while the cytosolic tail confers subunit-independent constitutive activity; mechanistically, NOX4-derived H2O2 inactivates PP1 metal centers to sustain eIF2α phosphorylation, oxidizes HDAC4 to de-repress Mef2A, oxidizes AKT to retain PP2A in the cytosol for DNA damage surveillance, activates Nrf2-dependent antioxidant programs, and at the MAM augments Akt-dependent InsP3R phosphorylation to restrict ER-to-mitochondria Ca2+ flux and prevent mitochondrial permeability transition, while upstream its transcription is controlled by STAT5, HOXD10, ATF3, TGF-β/Brd4-p300 epigenetic mechanisms, and IRP1 binding to IRE-like sequences in its promoter.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"NOX4 is a constitutively active, NADPH-dependent oxidase homologous to the phagocyte catalytic subunit gp91phox that generates reactive oxygen species to drive spatially confined redox signaling [#0, #7]. Distinct structural elements set its output and address: the cytosolic tail confers subunit-independent constitutive activity while the N-terminal region directs ER localization and dictates that NOX4 releases H2O2 rather than superoxide [#7], a determinant confirmed by genetic models showing NOX4 produces H2O2 constitutively [#33]. It localizes to the endoplasmic reticulum, mitochondria, and ER-mitochondria contact sites (MAMs) and requires p22phox as a cofactor, with the partner interaction demonstrated directly in endothelial cells [#3, #6, #4]; at mitochondrial membranes, CYB5R3 directly interacts with NOX4 and, through coenzyme Q, is required for optimal H2O2 output [#26]. NOX4 activity is allosterically inhibited by direct ATP binding [#16] and negatively regulated by FYN-mediated phosphorylation at Y566 [#14]. The H2O2 it produces acts on defined targets to control discrete cellular programs: it binds the PP1-targeting subunit GADD34 at the ER and oxidizes the PP1 metal center to sustain eIF2\\u03b1 phosphorylation and ATF4-driven autophagy [#13, #11], oxidizes HDAC4 to disrupt the HDAC4/Mef2A complex and enable endothelial tube formation [#22], oxidizes AKT to retain PP2A in the cytosol and preserve \\u03b3H2AX-dependent DNA damage surveillance [#25], activates Nrf2-dependent antioxidant programs [#9], and at the MAM augments Akt-dependent InsP3R phosphorylation to restrict ER-to-mitochondria Ca2+ flux and limit mitochondrial permeability transition [#19]. Through these outputs NOX4 governs proliferation, angiogenesis, metabolic gene expression, and cell-fate decisions including ferroptosis, and its transcription is controlled by inputs including STAT5, HOXD10, ATF3, and IRP1 binding to IRE-like promoter sequences [#12, #29, #30, #27]. NOX4-derived signaling is implicated across diabetic kidney disease, cardiac remodeling, leukemic transformation, tauopathy, and cancer [#2, #14, #12, #24].\",\n  \"teleology\": [\n    {\n      \"year\": 2001,\n      \"claim\": \"Establishing NOX4 as an NADPH oxidase family member answered whether it could be an enzymatic ROS source by showing it shares the catalytic architecture of gp91phox.\",\n      \"evidence\": \"cDNA cloning and sequence/domain analysis with tissue expression profiling\",\n      \"pmids\": [\"11376945\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Enzymatic activity not directly reconstituted in this work\", \"ROS species (H2O2 vs O2-) not determined\", \"Subcellular site not defined\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Linking NOX4 to angiotensin II-induced ROS and Akt activation placed it in a defined receptor-driven signaling pathway in mesangial cells.\",\n      \"evidence\": \"Antisense knockdown, dominant-negative Rac1, ROS and Akt phosphorylation assays\",\n      \"pmids\": [\"12842860\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Antisense specificity limits, no genetic KO\", \"Direct ROS species not resolved\", \"Mechanism of Rac1/arachidonate input unclear\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"ER localization of NOX4 answered where the enzyme acts, distinguishing it from plasma-membrane oxidases.\",\n      \"evidence\": \"Confocal microscopy of NOX4-GFP with ER, lysosome, and mitochondrial markers in HUVECs\",\n      \"pmids\": [\"15706079\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Based on overexpressed GFP fusion\", \"Does not address mitochondrial pools reported later\", \"No structural basis for targeting\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Demonstrating a direct NOX4-p22phox interaction at the ER identified the obligatory cofactor needed for endothelial ROS production.\",\n      \"evidence\": \"Bimolecular fluorescence complementation, calreticulin co-localization, siRNA, ROS assays\",\n      \"pmids\": [\"16987004\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"BiFC can trap transient interactions\", \"Stoichiometry not defined\", \"Single lab\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Domain-swap mapping established that the N-terminus dictates ER localization and the cytosolic tail confers constitutive, subunit-independent activity and H2O2 output, defining the structural logic of NOX4 specificity.\",\n      \"evidence\": \"Chimeric Nox1/Nox4 constructs, TIRF microscopy, ROS-type measurement in HEK293\",\n      \"pmids\": [\"19061439\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No atomic structure\", \"Mechanism of H2O2 vs O2- determination at the catalytic core unresolved\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Detection of NOX4 in mitochondria broadened its localization beyond the ER and tied it to glucose-induced mitochondrial superoxide in diabetic kidney.\",\n      \"evidence\": \"Subcellular fractionation, Mitotracker confocal, MitoProt prediction, siRNA, organelle NADPH oxidase assays\",\n      \"pmids\": [\"19706525\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Reconciliation with H2O2-only model not addressed here\", \"Import/targeting mechanism unknown\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Showing NOX4-derived ROS activates Nrf2 reframed NOX4 as a driver of antioxidant defense rather than purely oxidative damage.\",\n      \"evidence\": \"Transgenic cardiomyocyte overexpression, transcriptomics, glutathione assays, Nrf2-null epistasis\",\n      \"pmids\": [\"21554947\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Overexpression context\", \"Direct sensor of NOX4 H2O2 in Keap1/Nrf2 axis not identified\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Identifying STAT5-driven NOX4 transcription and NOX4-mediated DEP-1/PTPRJ inactivation explained how NOX4 sustains oncogenic FLT3ITD signaling in leukemia.\",\n      \"evidence\": \"STAT5 ChIP on NOX4 promoter, PTP activity assay, knockout transformation assays, in vivo disease models\",\n      \"pmids\": [\"26308771\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Site of redox-DEP1 encounter not localized\", \"Generalizability beyond FLT3ITD leukemia\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"The GADD34/PP1 work provided a direct molecular target, showing NOX4 H2O2 oxidizes the PP1 metal center to sustain eIF2\\u03b1 phosphorylation in a spatially confined manner at the ER.\",\n      \"evidence\": \"Co-IP with GADD34, PP1 activity assays distinguishing metal-center vs thiol oxidation, in vivo ischemia and AKI models\",\n      \"pmids\": [\"26742780\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How H2O2 is delivered to confined PP1 pool not fully resolved\", \"Other PP1 holoenzymes spared but mechanism of selectivity incomplete\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Identification of FYN-mediated Y566 phosphorylation defined a negative post-translational regulatory input restraining NOX4 in the heart.\",\n      \"evidence\": \"Co-IP, Y566 site-directed mutagenesis, activity assays, Nox4-/- rescue of FYN-/- cardiac phenotype\",\n      \"pmids\": [\"27525436\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural effect of Y566 phosphorylation on catalysis unknown\", \"Upstream control of FYN-NOX4 axis\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Discovery of an ATP-binding motif established direct allosteric inhibition of NOX4 by ATP, providing a metabolic switch coupling energy state to oxidase activity.\",\n      \"evidence\": \"ATP-binding and titration activity assays, inner mitochondrial membrane fractionation, PKM2 downstream pathway, xenografts\",\n      \"pmids\": [\"29051480\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"ATP-binding site not structurally resolved\", \"Inner-membrane localization claim contrasts with ER/MAM models\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"The Nox4/Nox2/p66Shc feed-forward axis showed how NOX4 H2O2 amplifies mitochondrial ROS and VEGFR2 signaling to drive angiogenesis.\",\n      \"evidence\": \"Organelle-targeted RoGFP imaging, double knockdown, p66Shc(S36A) mutant, EC migration/proliferation assays\",\n      \"pmids\": [\"28424170\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct molecular link from NOX4 H2O2 to Nox2 activation not defined\", \"Spatial coupling of the two oxidases unresolved\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"NOX4 H2O2 activation of TRPC6-dependent calcium influx connected the oxidase to podocyte calcium dysregulation in diabetic kidney disease.\",\n      \"evidence\": \"Nox4 KO rat, TRPC6 KO, H2O2 stimulation, calcium imaging, patch-clamp, EM\",\n      \"pmids\": [\"29793963\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct oxidative modification of TRPC6 not demonstrated\", \"Cell-type generality unclear\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Direct CD44-NOX4 association linked HA/CD44 signaling to NOX4-driven Ca2+-handling abnormalities in atrial fibrillation.\",\n      \"evidence\": \"Co-IP, CD44-/- mice, blocking antibody, Ca2+ spark measurement, AF patient tissue\",\n      \"pmids\": [\"31419440\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single Co-IP without reciprocal structural mapping\", \"Whether CD44 regulates NOX4 activity or only expression unclear\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Localizing NOX4 to MAMs and showing it augments Akt-dependent InsP3R phosphorylation defined a cytoprotective role restricting ER-to-mitochondria Ca2+ flux and limiting permeability transition.\",\n      \"evidence\": \"MAM fractionation, overexpression/KO, InsP3R phosphorylation, mitochondrial Ca2+ and mPT assays, cardiac ischemia-reperfusion\",\n      \"pmids\": [\"33001475\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How NOX4 H2O2 promotes Akt activity at MAM not fully resolved\", \"Direct InsP3R oxidation not shown\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"ER-confined NOX4 activation of the PERK-eIF2\\u03b1-ATF4 pathway during energy stress established NOX4 as an inducer of adaptive autophagy.\",\n      \"evidence\": \"Organelle-resolved NOX4 activity, genetic deletion, UPR pathway immunoblotting, autophagy flux in cardiomyocytes\",\n      \"pmids\": [\"24492492\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct PERK oxidation/sensor not identified\", \"Relationship to the GADD34/PP1 mechanism not integrated\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Cell-type-specific KO showed endothelial NOX4-derived H2O2 mediates endothelium-to-muscle cross-talk required for exercise-induced metabolic gene expression.\",\n      \"evidence\": \"Global and endothelial-specific Nox4 KO, 14C substrate oxidation, qPCR, catalase transgenics\",\n      \"pmids\": [\"33400973\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Identity of the cross-talk signal beyond H2O2 unknown\", \"Direct muscle target genes' redox sensors undefined\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Placing NOX4 downstream of SIRT1/NF-\\u03baB/FOXO defined it as an effector in tumor-induced muscle wasting.\",\n      \"evidence\": \"RNA-seq, muscle-specific Nox4 KO, pharmacological inhibition, SIRT1 reconstitution, cachexia models\",\n      \"pmids\": [\"32441762\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Downstream NOX4 targets in wasting muscle not identified\", \"Direct FOXO-NOX4 promoter binding not shown here\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Showing NOX4 oxidizes HDAC4 to disrupt the HDAC4/Mef2A complex provided a direct chromatin-linked substrate enabling endothelial tube formation.\",\n      \"evidence\": \"Inducible overexpression, HDAC4 oxidation assay, HDAC4/Mef2A Co-IP, redox-dead NOX4 and redox-insensitive HDAC4 mutants, tube formation\",\n      \"pmids\": [\"32818796\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Spatial route of H2O2 to nuclear/cytosolic HDAC4 unclear\", \"Overexpression-based system\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"NOX4 was shown to promote cell cycle entry by suppressing a p53/p21 checkpoint downstream of PDGF.\",\n      \"evidence\": \"siRNA, Rb phosphorylation, cyclin D1/p53/p21 immunoblotting, co-knockdown epistasis in fibroblasts\",\n      \"pmids\": [\"20531308\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct redox target upstream of p53/p21 not identified\", \"Single cell type\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"NOX4-derived ROS was found necessary for HIF2\\u03b1 nuclear accumulation and tumorigenesis in VHL-deficient renal carcinoma.\",\n      \"evidence\": \"siRNA, ROS scavengers, MnSOD/catalase, nuclear fractionation, invasion/colony assays, xenografts\",\n      \"pmids\": [\"24755467\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular target linking NOX4 ROS to HIF2\\u03b1 transport not defined\", \"Single tumor model\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"NOX4 oxidation of AKT to retain PP2A in the cytosol revealed a tumor-suppressive role maintaining \\u03b3H2AX-dependent DNA damage surveillance.\",\n      \"evidence\": \"Nox4 KO mice, carcinogen models, AKT oxidation assay, PP2A fractionation, \\u03b3H2AX readouts\",\n      \"pmids\": [\"33836590\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Site of AKT oxidation and its kinetics not fully mapped\", \"How redox state directs PP2A shuttling mechanistically\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identifying CYB5R3 as a direct NOX4 partner at the mitochondrial outer membrane defined a CoQ-dependent requirement for full H2O2 output.\",\n      \"evidence\": \"APEX2-EM, proximity biotinylation, PLA, Co-IP, CYB5R3 activity mutants, COQ6 knockdown, endothelial Cyb5r3 KO mice\",\n      \"pmids\": [\"34656824\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Electron-transfer mechanism between CYB5R3/CoQ and NOX4 not biochemically reconstituted\", \"Reconciliation with ER pool of NOX4\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"NOX4-driven non-mitochondrial ROS was shown to activate PERK/eIF2\\u03b1/ATF4 to promote autophagy and osteoclastogenesis, extending the UPR-autophagy axis to bone.\",\n      \"evidence\": \"Nox4 inhibitor and shRNA, NAC, PERK inhibitor, autophagy and osteoclastogenesis assays\",\n      \"pmids\": [\"34650437\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Pharmacological inhibitor specificity\", \"Direct UPR sensor target not identified\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Neuronal NOX4 was found to impair autophagy-lysosomal flux and drive tau accumulation, linking the oxidase to tauopathy.\",\n      \"evidence\": \"Global KO and neuronal AAV knockdown, tauopathy mouse model, autophagy flux, behavior\",\n      \"pmids\": [\"34922273\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular redox target controlling autophagy flux not identified\", \"Single model\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"IRP1 binding to IRE-like sequences in the NOX4 locus established an iron-sensing transcriptional input coupling iron overload to NOX4-driven ferroptosis.\",\n      \"evidence\": \"IRP1 binding assays on NOX4 promoter, iron loading, ferroptosis markers, Hepc1-/- mice with ferrostatin/DFO\",\n      \"pmids\": [\"36738798\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"IRE-like element function not validated by mutagenesis here\", \"Direct vs indirect IRP1 effect\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"ISG15-mediated ISGylation stabilizing NOX4 against ubiquitin-proteasome degradation, controlled by DRD4, defined a post-translational stability switch in acute kidney injury.\",\n      \"evidence\": \"Transcriptomics, ISG15 manipulation, Co-IP for ISGylation/ubiquitination, stability assays, DRD4 KO/overexpression AKI models\",\n      \"pmids\": [\"38354631\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"ISGylation sites on NOX4 not mapped\", \"E3 ligase mediating NOX4 ubiquitination unidentified\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"HOXD10 was shown to directly bind and repress the NOX4 promoter, and its hypermethylation de-represses NOX4 to drive TGF-\\u03b21-induced ferroptosis and renal fibrosis.\",\n      \"evidence\": \"ChIP, dual-luciferase, HOXD10 AAV overexpression, ferroptosis markers, bisulfite sequencing, UUO model\",\n      \"pmids\": [\"38844470\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab\", \"Interaction with other NOX4 transcriptional regulators not integrated\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"NOX4 interaction with activated PKC\\u03b1 and ATF3-driven transcription was linked to ferroptosis of dopaminergic neurons in a Parkinson's model.\",\n      \"evidence\": \"Co-IP with PKC\\u03b1, NOX4 inhibitor in MPTP mice, lipid peroxidation/iron assays, ATF3 analysis\",\n      \"pmids\": [\"38993139\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct functional consequence of PKC\\u03b1 binding on NOX4 activity unclear\", \"Single model and lab\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How NOX4's multiple subcellular pools (ER, MAM, mitochondrial membranes) are coordinated, and how a single diffusible H2O2 output achieves the documented target selectivity (PP1, AKT, HDAC4, InsP3R), remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No atomic structure of human NOX4 to explain H2O2 vs O2- output\", \"Mechanism conferring spatial confinement and target specificity of NOX4 H2O2 not defined\", \"Reconciliation of ER vs inner/outer mitochondrial membrane localization claims\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016491\", \"supporting_discovery_ids\": [0, 7, 33]},\n      {\"term_id\": \"GO:0140657\", \"supporting_discovery_ids\": [16]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [3, 4, 11, 13, 31]},\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [6, 16, 26]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [7]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [13, 9, 11, 31]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [2, 12, 15, 17, 25]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [27, 29, 30, 19]},\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [11, 23, 24, 31]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"CYBA\", \"FYN\", \"CYB5R3\", \"PPP1R15A\", \"HDAC4\", \"CD44\", \"PRKCA\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}