{"gene":"ELANE","run_date":"2026-06-09T23:54:42","timeline":{"discoveries":[{"year":1999,"finding":"Heterozygous single-base substitutions in ELA2 (encoding neutrophil elastase, a serine protease of neutrophil and monocyte granules) were identified in all 13 families with cyclic haematopoiesis, establishing that mutations in this gene cause the 21-day oscillation in blood-cell production.","method":"Genome-wide linkage screen and positional cloning, followed by direct sequencing of ELA2 in affected families","journal":"Nature genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — positional cloning with direct sequencing replicated across 13 independent families plus a sporadic case","pmids":["10581030"],"is_preprint":false},{"year":2003,"finding":"GFI1, a transcriptional repressor, directly represses ELA2 expression; dominant-negative GFI1 zinc-finger mutations in neutropenic patients elevate ELA2 expression in vivo, placing GFI1 upstream of ELA2 in a common myeloid differentiation pathway.","method":"Chromatin immunoprecipitation, electrophoretic mobility shift assay, reporter assays, and measurement of elevated ELA2 expression in patient bone marrow","journal":"Nature genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (ChIP, EMSA, reporter, in vivo expression) in a single rigorous study","pmids":["12778173"],"is_preprint":false},{"year":2007,"finding":"Expression of mutant (but not wild-type) NE in granulocytic precursors strongly induces BiP/GRP78 mRNA and XBP1 mRNA splicing (markers of the unfolded protein response), and activates CHOP/apoptosis in a protease-independent fashion; UPR activation and decreased NE protein were confirmed in primary SCN patient granulocytic precursors.","method":"Transient expression of mutant vs. wild-type NE in cell lines and primary human granulocytic precursors; quantitative RT-PCR for UPR markers; apoptosis assays","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — orthogonal cell-based assays plus primary patient cells; replicated across multiple mutants with phenotype-severity correlation","pmids":["17761833"],"is_preprint":false},{"year":2003,"finding":"LEF-1 and core-binding factor alpha (CBFalpha/AML1) co-activate the human ELA2 promoter; the high-mobility-group domain of LEF-1 physically interacts with the runt DNA-binding and proline/serine/threonine-rich activation domains of CBFalpha; promoter substitutions in SCN patients within the LEF-1 motif correlate with elevated ELA2 transcript levels.","method":"In vitro protein-protein interaction assay, promoter reporter assays, bone marrow mRNA quantification from SCN and familial platelet disorder/AML patients","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Moderate — in vitro interaction mapping plus reporter assays plus patient-derived mRNA evidence; multiple orthogonal methods in one study","pmids":["14594802"],"is_preprint":false},{"year":2006,"finding":"ELA2 expression is regulated by the hematopoietic transcription factors AML1, C/EBPalpha, PU.1, and c-Myb, as shown by chromatin immunoprecipitation and siRNA knockdown; the leukemic fusion protein AML1-ETO binds the ELA2 promoter but does not significantly repress ELA2 expression.","method":"Chromatin immunoprecipitation, siRNA knockdown, promoter reporter assays in myeloid cell lines","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP and siRNA in cell lines; single lab, two orthogonal methods","pmids":["16247445"],"is_preprint":false},{"year":2009,"finding":"LEF-1 directly activates the ELA2 promoter; LEF-1 transduction of hematopoietic cells upregulates ELA2/NE synthesis, while LEF-1 shRNA knockdown markedly reduces ELA2/NE levels; LEF-1 rescue of CD34+ cells from CN patients restores granulocytic differentiation with concomitant increase in functionally active NE.","method":"Promoter binding assays, lentiviral overexpression and shRNA knockdown of LEF-1 in hematopoietic cells, primary patient CD34+ cell rescue experiments","journal":"Blood","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — promoter assays plus gain-/loss-of-function in hematopoietic cells; single lab","pmids":["19620402"],"is_preprint":false},{"year":2007,"finding":"ELA2 mutations that cause cyclic neutropenia versus SCN have different and partly opposing effects on proteolytic activity, subcellular trafficking of NE, and induction of the unfolded protein response; pairs of cis mutations act approximately additively, and disturbed subcellular localization was identified as a potential disease mechanism distinct from the UPR.","method":"Functional characterization of mutant NE in transfected cells: protease activity assays, immunofluorescence for subcellular localization, UPR marker induction","journal":"Human mutation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal assays (activity, localization, UPR); single lab","pmids":["17436313"],"is_preprint":false},{"year":2015,"finding":"ELANE point mutations cause promyelocyte death and differentiation arrest associated with NE mislocalization away from primary granules and activation of UPR/ER stress; sivelestat (an NE-specific inhibitor) corrected dysgranulopoiesis by restoring normal NE localization to primary granules, ameliorating UPR/ER stress, and promoting promyelocyte survival and differentiation via C/EBPalpha (not C/EBPbeta).","method":"SCN patient-derived iPSCs with genome editing and granulopoiesis differentiation protocols; immunofluorescence for NE localization; UPR markers; pharmacological inhibition with sivelestat","journal":"The Journal of clinical investigation","confidence":"High","confidence_rationale":"Tier 2 / Strong — iPSC disease model with genome editing, multiple orthogonal functional readouts, pharmacological rescue; single rigorous study","pmids":["26193632"],"is_preprint":false},{"year":2013,"finding":"ELANE mutations that disrupt the translational start site force translation from downstream in-frame ATGs, producing amino-terminally truncated NE isoforms lacking the ER-localizing (pre) and zymogen-maintaining (pro) sequences but retaining catalytic residues; an internal ribosome entry site (IRES) within ELANE was defined; expression of these truncated isoforms reduces myeloid cell clonogenic capacity.","method":"Patient-derived iPSCs recapitulating the hematopoietic phenotype; identification of downstream translation initiation codons; IRES mapping; clonogenic assays in myeloid cells","journal":"Blood","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — iPSC phenotyping, molecular characterization of novel isoforms, IRES mapping; single lab with multiple orthogonal methods","pmids":["24184683"],"is_preprint":false},{"year":2021,"finding":"ELANE encodes neutrophil elastase which, together with cathepsin G and proteinase 3, catalyzes proteolytic cleavage of histone H3 at the amino terminus (H3ΔN) in human peripheral blood monocytes; this histone mark is repressed as monocytes differentiate into macrophages, and simultaneous NSP depletion causes H3ΔN loss and increased chromatin accessibility, priming chromatin for gene expression reprogramming during macrophage development.","method":"Integrative epigenomic analysis; simultaneous siRNA depletion of neutrophil serine proteases in monocytic cells; ATAC-seq for chromatin accessibility; primary monocyte and macrophage analysis","journal":"Nature immunology","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct enzymatic activity established for H3 cleavage, loss-of-function with defined epigenomic phenotype, primary human cells; single rigorous study with multiple orthogonal methods","pmids":["34017121"],"is_preprint":false},{"year":2021,"finding":"ELANE-mutant (misfolding) SCN patient-derived iPSC hematopoietic progenitors display elevated reactive oxygen species and increased numbers of PML nuclear bodies (markers of acute oxidative stress); PML controls the metabolic state of these progenitors and the expression of ELANE itself; both PML deletion and ELANE mutation correction restored normal G-CSF responses.","method":"iPSC-derived hematopoietic progenitor cells from SCN patients; ROS measurement; PML nuclear body quantification; PML genetic deletion; ELANE mutation correction; CSF3 response assays","journal":"Blood advances","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — iPSC model with genetic rescue experiments; single lab, multiple functional readouts","pmids":["33560392"],"is_preprint":false},{"year":2021,"finding":"CRISPR-based pooled screen in human hematopoietic stem and progenitor cells established that only -1 frameshift indels in late ELANE exons (mimicking SCN-associated mutations) impede neutrophil maturation, while -2 frame late-exon indels repress translation and support maturation; early exon disruption elicits nonsense-mediated decay, overcomes maturation arrest, and preserves hematopoietic engraftment.","method":"Pooled CRISPR screen in human HSPCs; nonsense-mediated decay assays; in vitro neutrophil differentiation; in vivo engraftment in humanized mice","journal":"Cell stem cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome editing screen with multiple allele types, in vitro and in vivo validation; single rigorous study with orthogonal methods","pmids":["33513358"],"is_preprint":false},{"year":2020,"finding":"The SCN-associated ELANE mutation p.G185R diminishes NE enzymatic activity and impairs granulocytic differentiation without significantly inducing the unfolded protein response or causing apoptosis in myeloblast cell lines, challenging the prevailing UPR model; impaired differentiation was associated with decreased expression of Gfi1, Cebpd, Cebpe, Spi1, Csf3r, and neutrophil granule protein genes.","method":"Inducible expression system for mutant ELANE in murine 32D and human NB4 cells; enzymatic activity assays; UPR marker measurement; gene expression profiling","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — inducible expression with enzymatic assays and transcriptomic readouts; single lab; contradicts UPR model reported by other labs","pmids":["32299910"],"is_preprint":false},{"year":2015,"finding":"Different ELANE mutants activate distinct UPR pathways: the CN-specific p.C71R mutant induces ATF6 and its target genes (PPP1R15A, DDIT3, HSPA5), while other mutants do not; secretory leukocyte protease inhibitor (SLPI), a natural NE inhibitor whose levels are reduced in CN but not CyN patients, modulates the extent of ELANE-triggered UPR when co-depleted with the p.S126L mutant.","method":"Transduction of hematopoietic cells with specific ELANE mutants; quantitative RT-PCR for UPR markers; shRNA knockdown of SLPI combined with mutant ELANE transduction","journal":"British journal of haematology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional cell assays with specific mutants plus SLPI knockdown; single lab","pmids":["26567890"],"is_preprint":false},{"year":2017,"finding":"Cell-permeable beta-lactam inhibitors of NE (particularly MK0339) promote cell survival and increase formation of mature neutrophils in three cellular models expressing mutant ELANE (patient iPSCs, HL60 transient expression, HL60 inducible expression), suggesting that inhibiting NE enzymatic activity or acting as a chaperone can rescue the differentiation defect.","method":"Patient-derived iPSCs, HL60 transient and inducible mutant NE expression models; annexin V/FACS viability assay; neutrophil differentiation assessment with NE inhibitor treatment","journal":"Journal of leukocyte biology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — pharmacological rescue in three cell models; mechanism (direct inhibition vs. chaperone) not definitively distinguished","pmids":["28754797"],"is_preprint":false},{"year":1998,"finding":"Mouse proteinase-3 (Prtn3) and neutrophil elastase (Ela2) genes are located within 2.2 kb of each other on mouse chromosome 10C2; each gene has five exons and four introns conserving the typical granule-associated serine proteinase gene structure; the catalytic triad and its placement are conserved; mouse Ela2 promoter contains functional c-myb and ets transcriptional elements.","method":"Molecular cloning, gene structure characterization, FISH chromosomal localization, sequence homology analysis, promoter element identification","journal":"Cytogenetics and cell genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct molecular characterization of gene structure and promoter elements; single study","pmids":["9925946"],"is_preprint":false},{"year":2017,"finding":"A novel ELANE mutation was found to decrease NE enzymatic activity, impair NETosis (neutrophil extracellular trap formation), and alter neutrophil cytokine production (IL-12, IL-8), without causing neutropenia; 3D structural modeling suggested the mutation alters protein folding and surface charge distribution, potentially perturbing protein trafficking.","method":"Scanning electron microscopy of NET formation; flow cytometry for neutrophil activation; fluorescent substrate-based NE activity assay; multiplex cytokine assay; 3D structural modeling","journal":"Arthritis & rheumatology (Hoboken, N.J.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct functional assays on patient neutrophils with multiple orthogonal methods; single case report with functional follow-up","pmids":["28881492"],"is_preprint":false},{"year":2019,"finding":"miR-608 directly targets ELANE mRNA: cotransfection of a miR-608 mimic into HEK293T cells inhibited ELANE 3'UTR reporter activity, and mutation of the miRNA seed sequences abolished this repression; decreased miR-608 in monocytes leads to ELANE overexpression at both mRNA and protein levels.","method":"Luciferase reporter assay with wild-type and mutant ELANE 3'UTR; lentiviral miR-608 inhibition in U937 cells; cDNA microarray; qRT-PCR and Western blot","journal":"Journal of clinical immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct reporter assay with mutagenesis confirms miR-608 targets ELANE 3'UTR; single lab","pmids":["31749032"],"is_preprint":false},{"year":2025,"finding":"ELANE (neutrophil elastase) promotes ferroptosis in hepatocytes in the context of MAFLD by enhancing KEAP1 protein stability: recombinant ELANE treatment weakened the binding between P62 and KEAP1, inhibiting lysosomal degradation of KEAP1, thereby increasing KEAP1 levels, enhancing NRF2 ubiquitination, and suppressing the NRF2/GPX4 anti-ferroptosis pathway; ELANE-KO mice showed opposite effects.","method":"Elane knockout mice on HFD; primary hepatocyte treatment with recombinant ELANE; Co-IP to assess P62-KEAP1 interaction; Western blot for KEAP1, NRF2, GPX4; ubiquitination assays; lysosomal degradation inhibition experiments","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, KO mice, recombinant protein treatment with multiple readouts; single lab, mechanistic pathway defined","pmids":["40204709"],"is_preprint":false},{"year":2022,"finding":"ELANE promotes M2 macrophage polarization by down-regulating PTEN expression in THP-1-derived macrophages, leading to increased M2 markers (CD206, CCL22, IL-10, CCL18); ELANE-polarized M2 macrophage conditioned medium enhanced lung cancer cell proliferation, migration, and invasion in vitro and tumor growth in vivo.","method":"THP-1 monocyte-to-macrophage differentiation; ELANE overexpression; qRT-PCR, ELISA, flow cytometry for M2 markers; Western blot for PTEN; co-culture conditioned medium assays; xenograft mouse model","journal":"Immunological investigations","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, limited mechanistic detail on how ELANE reduces PTEN; no direct enzymatic or binding experiment linking ELANE to PTEN","pmids":["36102787"],"is_preprint":false}],"current_model":"ELANE encodes neutrophil elastase (NE), a serine protease stored in neutrophil primary granules whose expression is transcriptionally activated by LEF-1, CBFalpha/AML1, C/EBPalpha, PU.1, and c-Myb and repressed post-transcriptionally by miR-608; dominant heterozygous ELANE mutations cause cyclic and severe congenital neutropenia primarily by producing misfolded NE that mislocalizes away from primary granules, triggers unfolded protein response/ER stress, and induces apoptosis of granulocytic precursors (with some mutations additionally generating truncated isoforms via alternative translation or disrupting NETosis), while in differentiated monocytes wild-type NE catalyzes proteolytic cleavage of histone H3 at the amino terminus to regulate chromatin accessibility during macrophage development, and in the liver context NE promotes ferroptosis by stabilizing KEAP1 and suppressing the NRF2/GPX4 pathway."},"narrative":{"mechanistic_narrative":"ELANE encodes neutrophil elastase (NE), a granule-associated serine protease whose dominant heterozygous mutations cause cyclic and severe congenital neutropenia [PMID:10581030]. Its transcription is driven by a network of hematopoietic factors — LEF-1, CBFalpha/AML1, C/EBPalpha, PU.1, and c-Myb — with LEF-1 directly activating the promoter and rescuing granulocytic differentiation in patient CD34+ cells [PMID:14594802, PMID:16247445, PMID:19620402], while the repressor GFI1 directly limits ELANE expression and dominant-negative GFI1 mutations elevate it in vivo [PMID:12778173]; post-transcriptionally, miR-608 targets the ELANE 3'UTR to restrain expression [PMID:31749032]. The dominant disease mechanism centers on protein misfolding: mutant NE mislocalizes away from primary granules and triggers the unfolded protein response and ER stress in a protease-independent manner, driving CHOP-dependent apoptosis and differentiation arrest of granulocytic precursors [PMID:17761833, PMID:26193632], with distinct mutations engaging distinct UPR arms and modulators such as ATF6 and SLPI [PMID:26567890] and an accompanying oxidative/PML-mediated stress state [PMID:33560392]. Pharmacological correction by NE inhibitors (sivelestat, beta-lactam MK0339) restores granule localization, relieves ER stress, and rescues differentiation via C/EBPalpha [PMID:26193632, PMID:28754797], and CRISPR mapping shows that only late-exon frameshifts mimicking SCN alleles impede maturation whereas early-exon disruption triggers nonsense-mediated decay and rescues the phenotype [PMID:33513358]. Beyond hematopoiesis, NE has enzymatic substrate roles: in monocytes it cleaves the histone H3 amino terminus (H3ΔN) to regulate chromatin accessibility during macrophage differentiation [PMID:34017121], and in hepatocytes it promotes ferroptosis by stabilizing KEAP1 and suppressing the NRF2/GPX4 pathway [PMID:40204709].","teleology":[{"year":1999,"claim":"Established the genetic cause of cyclic hematopoiesis, transforming an idiopathic oscillatory blood disorder into a defined single-gene disease.","evidence":"Genome-wide linkage and positional cloning with direct ELA2 sequencing across 13 affected families","pmids":["10581030"],"confidence":"High","gaps":["Did not explain how a protease mutation produces a 21-day oscillation","Functional consequence of the mutations on NE protein unresolved"]},{"year":2003,"claim":"Defined the transcriptional control of ELANE by placing the repressor GFI1 and the activators LEF-1/CBFalpha upstream within myeloid differentiation, connecting ELANE dysregulation to neutropenia beyond coding mutations.","evidence":"ChIP, EMSA, reporter assays, in vitro interaction mapping, and patient bone marrow expression measurements","pmids":["12778173","14594802"],"confidence":"High","gaps":["Whether elevated ELANE alone is sufficient to cause neutropenia not established","Combinatorial logic among the activators not resolved"]},{"year":2006,"claim":"Expanded the activating transcription factor set (AML1, C/EBPalpha, PU.1, c-Myb) regulating ELANE in myeloid cells.","evidence":"ChIP, siRNA knockdown, and promoter reporter assays in myeloid cell lines","pmids":["16247445"],"confidence":"Medium","gaps":["Relative contribution of each factor in vivo not quantified","Cell-line context may not reflect primary precursors"]},{"year":2007,"claim":"Identified protein misfolding–driven UPR/ER stress and apoptosis as a protease-independent disease mechanism, shifting the model from loss of enzymatic function to a toxic gain-of-function.","evidence":"Mutant vs. wild-type NE expression in cell lines and primary granulocytic precursors with UPR markers and apoptosis assays; functional characterization distinguishing CyN from SCN mutations","pmids":["17761833","17436313"],"confidence":"High","gaps":["Why specific mutations partition into cyclic versus severe phenotypes incompletely defined","Link between mislocalization and UPR not fully separated"]},{"year":2013,"claim":"Revealed an alternative translation mechanism in which start-site-disrupting mutations use downstream ATGs and an internal ribosome entry site to make truncated NE isoforms, broadening the molecular consequences of ELANE mutations.","evidence":"Patient-derived iPSCs, downstream initiation codon and IRES mapping, clonogenic assays","pmids":["24184683"],"confidence":"Medium","gaps":["In vivo relevance of truncated isoforms to neutropenia unclear","Single-lab characterization"]},{"year":2015,"claim":"Demonstrated that restoring NE localization to primary granules with the inhibitor sivelestat relieves ER stress and rescues differentiation via C/EBPalpha, establishing mislocalization as a therapeutically tractable driver, and that distinct mutants engage distinct UPR arms.","evidence":"Genome-edited SCN patient iPSC granulopoiesis with immunofluorescence, UPR markers, pharmacological rescue; mutant-specific ATF6/SLPI UPR analysis","pmids":["26193632","26567890"],"confidence":"High","gaps":["Whether localization rescue fully accounts for clinical response not established","Role of SLPI in patient outcomes unconfirmed"]},{"year":2020,"claim":"Challenged the universality of the UPR model by showing that the p.G185R mutant impairs differentiation through reduced enzymatic activity and altered myeloid transcription without inducing UPR or apoptosis.","evidence":"Inducible mutant ELANE expression in murine 32D and human NB4 cells with enzymatic assays and gene expression profiling","pmids":["32299910"],"confidence":"Medium","gaps":["Contradicts UPR findings from other labs; reconciliation across mutant classes unresolved","Cell-line model may not capture primary precursor biology"]},{"year":2021,"claim":"Defined the allele-specific genetic logic of ELANE-driven maturation arrest and identified an oxidative/PML stress axis, clarifying which lesions cause disease and pointing to gene-editing rescue strategies.","evidence":"Pooled CRISPR screen in human HSPCs with NMD assays and humanized-mouse engraftment; iPSC progenitors with ROS/PML quantification and genetic rescue","pmids":["33513358","33560392"],"confidence":"High","gaps":["Mechanistic link between PML/ROS state and ELANE expression incomplete","Therapeutic translation of early-exon editing not yet clinical"]},{"year":2021,"claim":"Assigned a physiological enzymatic substrate role to NE beyond neutropenia: cleavage of histone H3 to regulate chromatin accessibility during monocyte-to-macrophage development.","evidence":"Integrative epigenomics, simultaneous NSP siRNA depletion, and ATAC-seq in primary monocytes/macrophages","pmids":["34017121"],"confidence":"High","gaps":["Specific contribution of NE versus cathepsin G and proteinase 3 not separated","Functional consequences for macrophage gene programs broadly defined"]},{"year":2025,"claim":"Extended NE function to liver disease by showing it promotes hepatocyte ferroptosis through KEAP1 stabilization and NRF2/GPX4 suppression, implicating ELANE in metabolic-associated fatty liver disease.","evidence":"Elane-KO mice, recombinant ELANE on primary hepatocytes, Co-IP for P62-KEAP1, ubiquitination and lysosomal degradation assays","pmids":["40204709"],"confidence":"Medium","gaps":["Whether the effect requires NE catalytic activity not established","Single-lab mechanistic model"]},{"year":null,"claim":"How a single protease reconciles its toxic misfolding role in granulocyte precursors with its physiological substrate-cleavage roles in chromatin and ferroptosis, and which mutant class predominates in patients, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model linking UPR-dependent and enzymatic-activity-dependent disease mechanisms","Physiological substrate repertoire of NE incompletely mapped"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[9,18]},{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[0,6,16]},{"term_id":"GO:0140097","term_label":"catalytic activity, acting on DNA","supporting_discovery_ids":[9]}],"localization":[{"term_id":"GO:0031410","term_label":"cytoplasmic vesicle","supporting_discovery_ids":[7]},{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[2,7]}],"pathway":[{"term_id":"GO:0140097","term_label":"catalytic activity, acting on DNA","supporting_discovery_ids":[9]},{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[2,7]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[2,18]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[9,11]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[9,16]}],"complexes":[],"partners":["KEAP1","SLPI","GFI1","LEF1","PML"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P08246","full_name":"Neutrophil elastase","aliases":["Bone marrow serine protease","Elastase-2","Human leukocyte elastase","HLE","Medullasin","PMN elastase"],"length_aa":267,"mass_kda":28.5,"function":"Serine protease that modifies the functions of natural killer cells, monocytes and granulocytes. Inhibits C5a-dependent neutrophil enzyme release and chemotaxis (PubMed:15140022). Promotes cleavage of GSDMB, thereby inhibiting pyroptosis (PubMed:36899106). Promotes blood coagulation (PubMed:20676107). Through the activation of the platelet fibrinogen receptor integrin alpha-IIb/beta-3, potentiates platelet aggregation induced by a threshold concentration of cathepsin G (CTSG) (PubMed:25211214, PubMed:9111081). Cleaves and thus inactivates tissue factor pathway inhibitor (TFPI) (PubMed:20676107, PubMed:25211214). Capable of killing E.coli but not S.aureus in vitro; digests outer membrane protein A (ompA) in E.coli and K.pneumoniae (PubMed:10947984)","subcellular_location":"Cytoplasmic vesicle, phagosome","url":"https://www.uniprot.org/uniprotkb/P08246/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ELANE","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/ELANE","total_profiled":1310},"omim":[{"mim_id":"621351","title":"SERINE PROTEASE 57; PRSS57","url":"https://www.omim.org/entry/621351"},{"mim_id":"618023","title":"NOTCH2 N-TERMINAL-LIKE A; NOTCH2NLA","url":"https://www.omim.org/entry/618023"},{"mim_id":"615788","title":"NEDD4-BINDING PROTEIN 2-LIKE 2; N4BP2L2","url":"https://www.omim.org/entry/615788"},{"mim_id":"614161","title":"PR DOMAIN-CONTAINING PROTEIN 5; PRDM5","url":"https://www.omim.org/entry/614161"},{"mim_id":"613107","title":"NEUTROPENIA, SEVERE CONGENITAL, 2, AUTOSOMAL DOMINANT; SCN2","url":"https://www.omim.org/entry/613107"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Vesicles","reliability":"Supported"},{"location":"Golgi apparatus","reliability":"Additional"}],"tissue_specificity":"Tissue enriched","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"bone marrow","ntpm":3672.0}],"url":"https://www.proteinatlas.org/search/ELANE"},"hgnc":{"alias_symbol":["NE","HNE","HLE","PMN-E"],"prev_symbol":["ELA2"]},"alphafold":{"accession":"P08246","domains":[{"cath_id":"2.40.10.10","chopping":"31-251","consensus_level":"medium","plddt":95.8352,"start":31,"end":251}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P08246","model_url":"https://alphafold.ebi.ac.uk/files/AF-P08246-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P08246-F1-predicted_aligned_error_v6.png","plddt_mean":88.19},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ELANE","jax_strain_url":"https://www.jax.org/strain/search?query=ELANE"},"sequence":{"accession":"P08246","fasta_url":"https://rest.uniprot.org/uniprotkb/P08246.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P08246/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P08246"}},"corpus_meta":[{"pmid":"24096871","id":"PMC_24096871","title":"Cell death and diseases related to oxidative stress: 4-hydroxynonenal (HNE) in the balance.","date":"2013","source":"Cell death and differentiation","url":"https://pubmed.ncbi.nlm.nih.gov/24096871","citation_count":489,"is_preprint":false},{"pmid":"25598486","id":"PMC_25598486","title":"Role of lipid peroxidation derived 4-hydroxynonenal (4-HNE) in cancer: focusing on mitochondria.","date":"2014","source":"Redox biology","url":"https://pubmed.ncbi.nlm.nih.gov/25598486","citation_count":414,"is_preprint":false},{"pmid":"10581030","id":"PMC_10581030","title":"Mutations in ELA2, encoding neutrophil elastase, define a 21-day biological clock in cyclic haematopoiesis.","date":"1999","source":"Nature genetics","url":"https://pubmed.ncbi.nlm.nih.gov/10581030","citation_count":335,"is_preprint":false},{"pmid":"12778173","id":"PMC_12778173","title":"Mutations in proto-oncogene GFI1 cause human neutropenia and target ELA2.","date":"2003","source":"Nature 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reports","url":"https://pubmed.ncbi.nlm.nih.gov/33846419","citation_count":15,"is_preprint":false},{"pmid":"38556793","id":"PMC_38556793","title":"CRISPR-Cas9n-mediated ELANE promoter editing for gene therapy of severe congenital neutropenia.","date":"2024","source":"Molecular therapy : the journal of the American Society of Gene Therapy","url":"https://pubmed.ncbi.nlm.nih.gov/38556793","citation_count":14,"is_preprint":false},{"pmid":"9925946","id":"PMC_9925946","title":"Characterization and localization of the genes for mouse proteinase-3 (Prtn3) and neutrophil elastase (Ela2).","date":"1998","source":"Cytogenetics and cell genetics","url":"https://pubmed.ncbi.nlm.nih.gov/9925946","citation_count":14,"is_preprint":false},{"pmid":"36102787","id":"PMC_36102787","title":"ELANE Promotes M2 Macrophage Polarization by Down-Regulating PTEN and Participates in the Lung Cancer Progression.","date":"2022","source":"Immunological investigations","url":"https://pubmed.ncbi.nlm.nih.gov/36102787","citation_count":14,"is_preprint":false},{"pmid":"34580954","id":"PMC_34580954","title":"Reduced ELANE and SLPI expression compromises dental pulp cell activity.","date":"2021","source":"Cell proliferation","url":"https://pubmed.ncbi.nlm.nih.gov/34580954","citation_count":13,"is_preprint":false},{"pmid":"35583313","id":"PMC_35583313","title":"Characterization of an Anti-CD70 Half-Life Extended Bispecific T-Cell Engager (HLE-BiTE) and Associated On-Target Toxicity in Cynomolgus Monkeys.","date":"2022","source":"Toxicological sciences : an official journal of the Society of Toxicology","url":"https://pubmed.ncbi.nlm.nih.gov/35583313","citation_count":13,"is_preprint":false},{"pmid":"31749032","id":"PMC_31749032","title":"MiR-608 Exerts Anti-inflammatory Effects by Targeting ELANE in Monocytes.","date":"2019","source":"Journal of clinical immunology","url":"https://pubmed.ncbi.nlm.nih.gov/31749032","citation_count":13,"is_preprint":false},{"pmid":"12469218","id":"PMC_12469218","title":"CYP2E1 overexpression up-regulates both non-specific delta-aminolevulinate synthase and heme oxygenase-1 in the human hepatoma cell line HLE/2E1.","date":"2003","source":"International journal of molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/12469218","citation_count":13,"is_preprint":false},{"pmid":"26830905","id":"PMC_26830905","title":"4-HNE expression in diabetic rat kidneys and the protective effects of probucol.","date":"2016","source":"Journal of endocrinological investigation","url":"https://pubmed.ncbi.nlm.nih.gov/26830905","citation_count":13,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":50500,"output_tokens":5301,"usd":0.115507,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":13661,"output_tokens":3716,"usd":0.080602,"stage2_stop_reason":"end_turn"},"total_usd":0.196109,"stage1_batch_id":"msgbatch_01AsR7g8dGTEiEw9U83bJeWn","stage2_batch_id":"msgbatch_01YV5ZyxtFNUZEjxRcpd9jzb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1999,\n      \"finding\": \"Heterozygous single-base substitutions in ELA2 (encoding neutrophil elastase, a serine protease of neutrophil and monocyte granules) were identified in all 13 families with cyclic haematopoiesis, establishing that mutations in this gene cause the 21-day oscillation in blood-cell production.\",\n      \"method\": \"Genome-wide linkage screen and positional cloning, followed by direct sequencing of ELA2 in affected families\",\n      \"journal\": \"Nature genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — positional cloning with direct sequencing replicated across 13 independent families plus a sporadic case\",\n      \"pmids\": [\"10581030\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"GFI1, a transcriptional repressor, directly represses ELA2 expression; dominant-negative GFI1 zinc-finger mutations in neutropenic patients elevate ELA2 expression in vivo, placing GFI1 upstream of ELA2 in a common myeloid differentiation pathway.\",\n      \"method\": \"Chromatin immunoprecipitation, electrophoretic mobility shift assay, reporter assays, and measurement of elevated ELA2 expression in patient bone marrow\",\n      \"journal\": \"Nature genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (ChIP, EMSA, reporter, in vivo expression) in a single rigorous study\",\n      \"pmids\": [\"12778173\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Expression of mutant (but not wild-type) NE in granulocytic precursors strongly induces BiP/GRP78 mRNA and XBP1 mRNA splicing (markers of the unfolded protein response), and activates CHOP/apoptosis in a protease-independent fashion; UPR activation and decreased NE protein were confirmed in primary SCN patient granulocytic precursors.\",\n      \"method\": \"Transient expression of mutant vs. wild-type NE in cell lines and primary human granulocytic precursors; quantitative RT-PCR for UPR markers; apoptosis assays\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — orthogonal cell-based assays plus primary patient cells; replicated across multiple mutants with phenotype-severity correlation\",\n      \"pmids\": [\"17761833\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"LEF-1 and core-binding factor alpha (CBFalpha/AML1) co-activate the human ELA2 promoter; the high-mobility-group domain of LEF-1 physically interacts with the runt DNA-binding and proline/serine/threonine-rich activation domains of CBFalpha; promoter substitutions in SCN patients within the LEF-1 motif correlate with elevated ELA2 transcript levels.\",\n      \"method\": \"In vitro protein-protein interaction assay, promoter reporter assays, bone marrow mRNA quantification from SCN and familial platelet disorder/AML patients\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro interaction mapping plus reporter assays plus patient-derived mRNA evidence; multiple orthogonal methods in one study\",\n      \"pmids\": [\"14594802\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"ELA2 expression is regulated by the hematopoietic transcription factors AML1, C/EBPalpha, PU.1, and c-Myb, as shown by chromatin immunoprecipitation and siRNA knockdown; the leukemic fusion protein AML1-ETO binds the ELA2 promoter but does not significantly repress ELA2 expression.\",\n      \"method\": \"Chromatin immunoprecipitation, siRNA knockdown, promoter reporter assays in myeloid cell lines\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP and siRNA in cell lines; single lab, two orthogonal methods\",\n      \"pmids\": [\"16247445\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"LEF-1 directly activates the ELA2 promoter; LEF-1 transduction of hematopoietic cells upregulates ELA2/NE synthesis, while LEF-1 shRNA knockdown markedly reduces ELA2/NE levels; LEF-1 rescue of CD34+ cells from CN patients restores granulocytic differentiation with concomitant increase in functionally active NE.\",\n      \"method\": \"Promoter binding assays, lentiviral overexpression and shRNA knockdown of LEF-1 in hematopoietic cells, primary patient CD34+ cell rescue experiments\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — promoter assays plus gain-/loss-of-function in hematopoietic cells; single lab\",\n      \"pmids\": [\"19620402\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"ELA2 mutations that cause cyclic neutropenia versus SCN have different and partly opposing effects on proteolytic activity, subcellular trafficking of NE, and induction of the unfolded protein response; pairs of cis mutations act approximately additively, and disturbed subcellular localization was identified as a potential disease mechanism distinct from the UPR.\",\n      \"method\": \"Functional characterization of mutant NE in transfected cells: protease activity assays, immunofluorescence for subcellular localization, UPR marker induction\",\n      \"journal\": \"Human mutation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal assays (activity, localization, UPR); single lab\",\n      \"pmids\": [\"17436313\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"ELANE point mutations cause promyelocyte death and differentiation arrest associated with NE mislocalization away from primary granules and activation of UPR/ER stress; sivelestat (an NE-specific inhibitor) corrected dysgranulopoiesis by restoring normal NE localization to primary granules, ameliorating UPR/ER stress, and promoting promyelocyte survival and differentiation via C/EBPalpha (not C/EBPbeta).\",\n      \"method\": \"SCN patient-derived iPSCs with genome editing and granulopoiesis differentiation protocols; immunofluorescence for NE localization; UPR markers; pharmacological inhibition with sivelestat\",\n      \"journal\": \"The Journal of clinical investigation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — iPSC disease model with genome editing, multiple orthogonal functional readouts, pharmacological rescue; single rigorous study\",\n      \"pmids\": [\"26193632\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"ELANE mutations that disrupt the translational start site force translation from downstream in-frame ATGs, producing amino-terminally truncated NE isoforms lacking the ER-localizing (pre) and zymogen-maintaining (pro) sequences but retaining catalytic residues; an internal ribosome entry site (IRES) within ELANE was defined; expression of these truncated isoforms reduces myeloid cell clonogenic capacity.\",\n      \"method\": \"Patient-derived iPSCs recapitulating the hematopoietic phenotype; identification of downstream translation initiation codons; IRES mapping; clonogenic assays in myeloid cells\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — iPSC phenotyping, molecular characterization of novel isoforms, IRES mapping; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"24184683\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ELANE encodes neutrophil elastase which, together with cathepsin G and proteinase 3, catalyzes proteolytic cleavage of histone H3 at the amino terminus (H3ΔN) in human peripheral blood monocytes; this histone mark is repressed as monocytes differentiate into macrophages, and simultaneous NSP depletion causes H3ΔN loss and increased chromatin accessibility, priming chromatin for gene expression reprogramming during macrophage development.\",\n      \"method\": \"Integrative epigenomic analysis; simultaneous siRNA depletion of neutrophil serine proteases in monocytic cells; ATAC-seq for chromatin accessibility; primary monocyte and macrophage analysis\",\n      \"journal\": \"Nature immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct enzymatic activity established for H3 cleavage, loss-of-function with defined epigenomic phenotype, primary human cells; single rigorous study with multiple orthogonal methods\",\n      \"pmids\": [\"34017121\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ELANE-mutant (misfolding) SCN patient-derived iPSC hematopoietic progenitors display elevated reactive oxygen species and increased numbers of PML nuclear bodies (markers of acute oxidative stress); PML controls the metabolic state of these progenitors and the expression of ELANE itself; both PML deletion and ELANE mutation correction restored normal G-CSF responses.\",\n      \"method\": \"iPSC-derived hematopoietic progenitor cells from SCN patients; ROS measurement; PML nuclear body quantification; PML genetic deletion; ELANE mutation correction; CSF3 response assays\",\n      \"journal\": \"Blood advances\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — iPSC model with genetic rescue experiments; single lab, multiple functional readouts\",\n      \"pmids\": [\"33560392\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"CRISPR-based pooled screen in human hematopoietic stem and progenitor cells established that only -1 frameshift indels in late ELANE exons (mimicking SCN-associated mutations) impede neutrophil maturation, while -2 frame late-exon indels repress translation and support maturation; early exon disruption elicits nonsense-mediated decay, overcomes maturation arrest, and preserves hematopoietic engraftment.\",\n      \"method\": \"Pooled CRISPR screen in human HSPCs; nonsense-mediated decay assays; in vitro neutrophil differentiation; in vivo engraftment in humanized mice\",\n      \"journal\": \"Cell stem cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome editing screen with multiple allele types, in vitro and in vivo validation; single rigorous study with orthogonal methods\",\n      \"pmids\": [\"33513358\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"The SCN-associated ELANE mutation p.G185R diminishes NE enzymatic activity and impairs granulocytic differentiation without significantly inducing the unfolded protein response or causing apoptosis in myeloblast cell lines, challenging the prevailing UPR model; impaired differentiation was associated with decreased expression of Gfi1, Cebpd, Cebpe, Spi1, Csf3r, and neutrophil granule protein genes.\",\n      \"method\": \"Inducible expression system for mutant ELANE in murine 32D and human NB4 cells; enzymatic activity assays; UPR marker measurement; gene expression profiling\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — inducible expression with enzymatic assays and transcriptomic readouts; single lab; contradicts UPR model reported by other labs\",\n      \"pmids\": [\"32299910\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Different ELANE mutants activate distinct UPR pathways: the CN-specific p.C71R mutant induces ATF6 and its target genes (PPP1R15A, DDIT3, HSPA5), while other mutants do not; secretory leukocyte protease inhibitor (SLPI), a natural NE inhibitor whose levels are reduced in CN but not CyN patients, modulates the extent of ELANE-triggered UPR when co-depleted with the p.S126L mutant.\",\n      \"method\": \"Transduction of hematopoietic cells with specific ELANE mutants; quantitative RT-PCR for UPR markers; shRNA knockdown of SLPI combined with mutant ELANE transduction\",\n      \"journal\": \"British journal of haematology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional cell assays with specific mutants plus SLPI knockdown; single lab\",\n      \"pmids\": [\"26567890\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Cell-permeable beta-lactam inhibitors of NE (particularly MK0339) promote cell survival and increase formation of mature neutrophils in three cellular models expressing mutant ELANE (patient iPSCs, HL60 transient expression, HL60 inducible expression), suggesting that inhibiting NE enzymatic activity or acting as a chaperone can rescue the differentiation defect.\",\n      \"method\": \"Patient-derived iPSCs, HL60 transient and inducible mutant NE expression models; annexin V/FACS viability assay; neutrophil differentiation assessment with NE inhibitor treatment\",\n      \"journal\": \"Journal of leukocyte biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — pharmacological rescue in three cell models; mechanism (direct inhibition vs. chaperone) not definitively distinguished\",\n      \"pmids\": [\"28754797\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"Mouse proteinase-3 (Prtn3) and neutrophil elastase (Ela2) genes are located within 2.2 kb of each other on mouse chromosome 10C2; each gene has five exons and four introns conserving the typical granule-associated serine proteinase gene structure; the catalytic triad and its placement are conserved; mouse Ela2 promoter contains functional c-myb and ets transcriptional elements.\",\n      \"method\": \"Molecular cloning, gene structure characterization, FISH chromosomal localization, sequence homology analysis, promoter element identification\",\n      \"journal\": \"Cytogenetics and cell genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct molecular characterization of gene structure and promoter elements; single study\",\n      \"pmids\": [\"9925946\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"A novel ELANE mutation was found to decrease NE enzymatic activity, impair NETosis (neutrophil extracellular trap formation), and alter neutrophil cytokine production (IL-12, IL-8), without causing neutropenia; 3D structural modeling suggested the mutation alters protein folding and surface charge distribution, potentially perturbing protein trafficking.\",\n      \"method\": \"Scanning electron microscopy of NET formation; flow cytometry for neutrophil activation; fluorescent substrate-based NE activity assay; multiplex cytokine assay; 3D structural modeling\",\n      \"journal\": \"Arthritis & rheumatology (Hoboken, N.J.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct functional assays on patient neutrophils with multiple orthogonal methods; single case report with functional follow-up\",\n      \"pmids\": [\"28881492\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"miR-608 directly targets ELANE mRNA: cotransfection of a miR-608 mimic into HEK293T cells inhibited ELANE 3'UTR reporter activity, and mutation of the miRNA seed sequences abolished this repression; decreased miR-608 in monocytes leads to ELANE overexpression at both mRNA and protein levels.\",\n      \"method\": \"Luciferase reporter assay with wild-type and mutant ELANE 3'UTR; lentiviral miR-608 inhibition in U937 cells; cDNA microarray; qRT-PCR and Western blot\",\n      \"journal\": \"Journal of clinical immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct reporter assay with mutagenesis confirms miR-608 targets ELANE 3'UTR; single lab\",\n      \"pmids\": [\"31749032\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"ELANE (neutrophil elastase) promotes ferroptosis in hepatocytes in the context of MAFLD by enhancing KEAP1 protein stability: recombinant ELANE treatment weakened the binding between P62 and KEAP1, inhibiting lysosomal degradation of KEAP1, thereby increasing KEAP1 levels, enhancing NRF2 ubiquitination, and suppressing the NRF2/GPX4 anti-ferroptosis pathway; ELANE-KO mice showed opposite effects.\",\n      \"method\": \"Elane knockout mice on HFD; primary hepatocyte treatment with recombinant ELANE; Co-IP to assess P62-KEAP1 interaction; Western blot for KEAP1, NRF2, GPX4; ubiquitination assays; lysosomal degradation inhibition experiments\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, KO mice, recombinant protein treatment with multiple readouts; single lab, mechanistic pathway defined\",\n      \"pmids\": [\"40204709\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"ELANE promotes M2 macrophage polarization by down-regulating PTEN expression in THP-1-derived macrophages, leading to increased M2 markers (CD206, CCL22, IL-10, CCL18); ELANE-polarized M2 macrophage conditioned medium enhanced lung cancer cell proliferation, migration, and invasion in vitro and tumor growth in vivo.\",\n      \"method\": \"THP-1 monocyte-to-macrophage differentiation; ELANE overexpression; qRT-PCR, ELISA, flow cytometry for M2 markers; Western blot for PTEN; co-culture conditioned medium assays; xenograft mouse model\",\n      \"journal\": \"Immunological investigations\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, limited mechanistic detail on how ELANE reduces PTEN; no direct enzymatic or binding experiment linking ELANE to PTEN\",\n      \"pmids\": [\"36102787\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ELANE encodes neutrophil elastase (NE), a serine protease stored in neutrophil primary granules whose expression is transcriptionally activated by LEF-1, CBFalpha/AML1, C/EBPalpha, PU.1, and c-Myb and repressed post-transcriptionally by miR-608; dominant heterozygous ELANE mutations cause cyclic and severe congenital neutropenia primarily by producing misfolded NE that mislocalizes away from primary granules, triggers unfolded protein response/ER stress, and induces apoptosis of granulocytic precursors (with some mutations additionally generating truncated isoforms via alternative translation or disrupting NETosis), while in differentiated monocytes wild-type NE catalyzes proteolytic cleavage of histone H3 at the amino terminus to regulate chromatin accessibility during macrophage development, and in the liver context NE promotes ferroptosis by stabilizing KEAP1 and suppressing the NRF2/GPX4 pathway.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ELANE encodes neutrophil elastase (NE), a granule-associated serine protease whose dominant heterozygous mutations cause cyclic and severe congenital neutropenia [#0]. Its transcription is driven by a network of hematopoietic factors — LEF-1, CBFalpha/AML1, C/EBPalpha, PU.1, and c-Myb — with LEF-1 directly activating the promoter and rescuing granulocytic differentiation in patient CD34+ cells [#3, #4, #5], while the repressor GFI1 directly limits ELANE expression and dominant-negative GFI1 mutations elevate it in vivo [#1]; post-transcriptionally, miR-608 targets the ELANE 3'UTR to restrain expression [#17]. The dominant disease mechanism centers on protein misfolding: mutant NE mislocalizes away from primary granules and triggers the unfolded protein response and ER stress in a protease-independent manner, driving CHOP-dependent apoptosis and differentiation arrest of granulocytic precursors [#2, #7], with distinct mutations engaging distinct UPR arms and modulators such as ATF6 and SLPI [#13] and an accompanying oxidative/PML-mediated stress state [#10]. Pharmacological correction by NE inhibitors (sivelestat, beta-lactam MK0339) restores granule localization, relieves ER stress, and rescues differentiation via C/EBPalpha [#7, #14], and CRISPR mapping shows that only late-exon frameshifts mimicking SCN alleles impede maturation whereas early-exon disruption triggers nonsense-mediated decay and rescues the phenotype [#11]. Beyond hematopoiesis, NE has enzymatic substrate roles: in monocytes it cleaves the histone H3 amino terminus (H3ΔN) to regulate chromatin accessibility during macrophage differentiation [#9], and in hepatocytes it promotes ferroptosis by stabilizing KEAP1 and suppressing the NRF2/GPX4 pathway [#18].\",\n  \"teleology\": [\n    {\n      \"year\": 1999,\n      \"claim\": \"Established the genetic cause of cyclic hematopoiesis, transforming an idiopathic oscillatory blood disorder into a defined single-gene disease.\",\n      \"evidence\": \"Genome-wide linkage and positional cloning with direct ELA2 sequencing across 13 affected families\",\n      \"pmids\": [\"10581030\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not explain how a protease mutation produces a 21-day oscillation\", \"Functional consequence of the mutations on NE protein unresolved\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Defined the transcriptional control of ELANE by placing the repressor GFI1 and the activators LEF-1/CBFalpha upstream within myeloid differentiation, connecting ELANE dysregulation to neutropenia beyond coding mutations.\",\n      \"evidence\": \"ChIP, EMSA, reporter assays, in vitro interaction mapping, and patient bone marrow expression measurements\",\n      \"pmids\": [\"12778173\", \"14594802\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether elevated ELANE alone is sufficient to cause neutropenia not established\", \"Combinatorial logic among the activators not resolved\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Expanded the activating transcription factor set (AML1, C/EBPalpha, PU.1, c-Myb) regulating ELANE in myeloid cells.\",\n      \"evidence\": \"ChIP, siRNA knockdown, and promoter reporter assays in myeloid cell lines\",\n      \"pmids\": [\"16247445\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Relative contribution of each factor in vivo not quantified\", \"Cell-line context may not reflect primary precursors\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Identified protein misfolding–driven UPR/ER stress and apoptosis as a protease-independent disease mechanism, shifting the model from loss of enzymatic function to a toxic gain-of-function.\",\n      \"evidence\": \"Mutant vs. wild-type NE expression in cell lines and primary granulocytic precursors with UPR markers and apoptosis assays; functional characterization distinguishing CyN from SCN mutations\",\n      \"pmids\": [\"17761833\", \"17436313\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Why specific mutations partition into cyclic versus severe phenotypes incompletely defined\", \"Link between mislocalization and UPR not fully separated\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Revealed an alternative translation mechanism in which start-site-disrupting mutations use downstream ATGs and an internal ribosome entry site to make truncated NE isoforms, broadening the molecular consequences of ELANE mutations.\",\n      \"evidence\": \"Patient-derived iPSCs, downstream initiation codon and IRES mapping, clonogenic assays\",\n      \"pmids\": [\"24184683\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"In vivo relevance of truncated isoforms to neutropenia unclear\", \"Single-lab characterization\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Demonstrated that restoring NE localization to primary granules with the inhibitor sivelestat relieves ER stress and rescues differentiation via C/EBPalpha, establishing mislocalization as a therapeutically tractable driver, and that distinct mutants engage distinct UPR arms.\",\n      \"evidence\": \"Genome-edited SCN patient iPSC granulopoiesis with immunofluorescence, UPR markers, pharmacological rescue; mutant-specific ATF6/SLPI UPR analysis\",\n      \"pmids\": [\"26193632\", \"26567890\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether localization rescue fully accounts for clinical response not established\", \"Role of SLPI in patient outcomes unconfirmed\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Challenged the universality of the UPR model by showing that the p.G185R mutant impairs differentiation through reduced enzymatic activity and altered myeloid transcription without inducing UPR or apoptosis.\",\n      \"evidence\": \"Inducible mutant ELANE expression in murine 32D and human NB4 cells with enzymatic assays and gene expression profiling\",\n      \"pmids\": [\"32299910\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Contradicts UPR findings from other labs; reconciliation across mutant classes unresolved\", \"Cell-line model may not capture primary precursor biology\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Defined the allele-specific genetic logic of ELANE-driven maturation arrest and identified an oxidative/PML stress axis, clarifying which lesions cause disease and pointing to gene-editing rescue strategies.\",\n      \"evidence\": \"Pooled CRISPR screen in human HSPCs with NMD assays and humanized-mouse engraftment; iPSC progenitors with ROS/PML quantification and genetic rescue\",\n      \"pmids\": [\"33513358\", \"33560392\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanistic link between PML/ROS state and ELANE expression incomplete\", \"Therapeutic translation of early-exon editing not yet clinical\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Assigned a physiological enzymatic substrate role to NE beyond neutropenia: cleavage of histone H3 to regulate chromatin accessibility during monocyte-to-macrophage development.\",\n      \"evidence\": \"Integrative epigenomics, simultaneous NSP siRNA depletion, and ATAC-seq in primary monocytes/macrophages\",\n      \"pmids\": [\"34017121\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Specific contribution of NE versus cathepsin G and proteinase 3 not separated\", \"Functional consequences for macrophage gene programs broadly defined\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Extended NE function to liver disease by showing it promotes hepatocyte ferroptosis through KEAP1 stabilization and NRF2/GPX4 suppression, implicating ELANE in metabolic-associated fatty liver disease.\",\n      \"evidence\": \"Elane-KO mice, recombinant ELANE on primary hepatocytes, Co-IP for P62-KEAP1, ubiquitination and lysosomal degradation assays\",\n      \"pmids\": [\"40204709\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether the effect requires NE catalytic activity not established\", \"Single-lab mechanistic model\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How a single protease reconciles its toxic misfolding role in granulocyte precursors with its physiological substrate-cleavage roles in chromatin and ferroptosis, and which mutant class predominates in patients, remains unresolved.\",\n      \"evidence\": null,\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified model linking UPR-dependent and enzymatic-activity-dependent disease mechanisms\", \"Physiological substrate repertoire of NE incompletely mapped\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [9, 18]},\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [0, 6, 16]},\n      {\"term_id\": \"GO:0140097\", \"supporting_discovery_ids\": [9]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0031410\", \"supporting_discovery_ids\": [7]},\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [2, 7]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0140097\", \"supporting_discovery_ids\": [9]},\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [2, 7]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [2, 18]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [9, 11]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [9, 16]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"KEAP1\", \"SLPI\", \"GFI1\", \"LEF1\", \"PML\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}