{"gene":"DLK1","run_date":"2026-04-28T17:46:02","timeline":{"discoveries":[{"year":1993,"finding":"Pref-1 (DLK1) is synthesized as a transmembrane protein with six tandem EGF-like repeats and functions as a negative regulator of adipocyte differentiation; constitutive expression in preadipocytes blocks adipogenesis.","method":"cDNA cloning, constitutive overexpression in 3T3-L1 preadipocytes with differentiation assay","journal":"Cell","confidence":"High","confidence_rationale":"Tier 2 — foundational gain-of-function study in cell culture, replicated across many subsequent studies","pmids":["8500166"],"is_preprint":false},{"year":1997,"finding":"Membrane-associated Pref-1 (DLK1) is cleaved in the juxtamembrane region to generate a soluble ~50 kDa ectodomain that is biologically active and inhibits adipocyte differentiation; only the two largest alternatively spliced isoforms undergo this cleavage, establishing that alternative splicing governs juxtacrine vs. paracrine modes of Pref-1 action.","method":"In vitro cleavage assays, addition of recombinant ectodomain to 3T3-L1 cells, antibody neutralization, alternate splice-form analysis","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1/2 — reconstitution with purified ectodomain plus antibody blocking and splice-form dissection in single rigorous study","pmids":["9001251"],"is_preprint":false},{"year":1999,"finding":"Dexamethasone promotes adipogenesis by transcriptionally repressing the Pref-1 (DLK1) gene; nuclear run-on assays showed reduced Pref-1 transcription, and antisense Pref-1 transfection enhanced adipogenesis at low dexamethasone concentrations, placing Pref-1 downregulation as a key mechanism of glucocorticoid-driven adipoconversion.","method":"Nuclear run-on transcription assay, actinomycin D mRNA stability assay, stable antisense transfection with differentiation assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1/2 — multiple orthogonal methods (run-on, stability, antisense) in a single study","pmids":["10212243"],"is_preprint":false},{"year":1998,"finding":"A specific cis-element in the Pref-1 (DLK1) promoter, the SAD (suppression in adipocyte differentiation) element (core sequence -183AAAGA-179), is required for differentiation-dependent transcriptional suppression of the Pref-1 gene; an ~63 kDa nuclear protein binds this element.","method":"Stable transfection of 5'-deletion reporter constructs, gel mobility shift assay, UV cross-linking, base-substitution mutagenesis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 — deletion mapping + mutagenesis + DNA-protein interaction assays","pmids":["9822638"],"is_preprint":false},{"year":2004,"finding":"Constitutive overexpression of DLK1/Pref-1 in human mesenchymal stem cells inhibits both osteoblast and adipocyte differentiation without affecting proliferation, identifying DLK1 as a regulator that maintains the bipotential progenitor pool.","method":"Retroviral overexpression in hMSC-TERT cells, cytochemical staining, FACS, real-time PCR, ex vivo calvaria organ culture","journal":"Journal of bone and mineral research","confidence":"High","confidence_rationale":"Tier 2 — multiple orthogonal readouts (in vitro and ex vivo) in human cells","pmids":["15068508"],"is_preprint":false},{"year":2004,"finding":"Ectopic expression of DLK1 protein in skeletal muscle of callipyge sheep (+(MAT)/CLPG(PAT)) causes muscular hypertrophy; transgenic mice expressing DLK1 in skeletal muscle develop generalized muscular hypertrophy, establishing DLK1 as the causal agent of the callipyge phenotype.","method":"Immunohistochemistry in sheep muscle, transgenic mouse model with skeletal muscle-specific DLK1 expression","journal":"Current biology : CB","confidence":"High","confidence_rationale":"Tier 2 — in vivo transgenic rescue/overexpression with clear phenotypic readout, cross-validated in two species","pmids":["15498495"],"is_preprint":false},{"year":2006,"finding":"The EGF-like extracellular domain of DLK1 inhibits Notch signaling in mesenchymal C3H10T1/2 cells, but this Notch inhibition paradoxically potentiates adipogenesis in these cells (unlike 3T3-L1 cells); the effect requires Notch1 expression and is mediated by the extracellular, not the intracellular, domain of DLK1.","method":"Overexpression of full-length vs. domain-deletion DLK1 constructs, co-culture assay, ERK1/2 activation measurement, Notch1 knockdown","journal":"Journal of molecular biology","confidence":"Medium","confidence_rationale":"Tier 2 — domain dissection and co-culture experiments, single lab","pmids":["17320900"],"is_preprint":false},{"year":2009,"finding":"Soluble Pref-1 (DLK1) activates the MEK/ERK pathway; ERK activation by Pref-1 prevents downregulation of Sox9, which in turn suppresses C/EBPβ and C/EBPδ expression and blocks adipogenesis. Pref-1 also induces Sox9 to promote chondrogenic lineage commitment while inhibiting chondrocyte maturation and osteoblast differentiation.","method":"Review synthesizing signaling pathway data from multiple mechanistic studies (MAPK assays, Sox9 overexpression/knockdown, C/EBP reporter assays)","journal":"Molecular endocrinology (Baltimore, Md.)","confidence":"Medium","confidence_rationale":"Tier 2 — pathway placement based on multiple prior experimental studies consolidated in review; MEK/ERK-Sox9 axis experimentally supported","pmids":["19541743"],"is_preprint":false},{"year":2010,"finding":"Dlk1 is required for normal skeletal muscle development and regeneration; muscle-specific Dlk1 knockout reduces myofiber number and MyoD expression, impairs regeneration with augmented NF-κB and inflammatory cytokine signaling, and promotes satellite cell self-renewal; conversely, Dlk1 overexpression inhibits satellite cell proliferation and enhances differentiation.","method":"Conditional knockout and overexpression in mice, satellite cell single-fiber culture, immunofluorescence, gene expression analysis","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 — conditional KO plus overexpression with multiple cellular and molecular readouts in vivo and ex vivo","pmids":["21124733"],"is_preprint":false},{"year":2012,"finding":"Membrane-bound DLK1 (DLK1-M), but not soluble DLK1, inhibits G1-to-S-phase cell cycle progression and preadipocyte proliferation; DLK1-null mice show increased preadipocyte replication and higher fat mass, establishing a dual inhibitory role of DLK1 on both proliferation and differentiation in adipogenesis.","method":"Independent manipulation of DLK1 isoform levels in preadipocytes, cell cycle analysis, in vivo analysis of DLK1-null mice","journal":"Diabetes","confidence":"High","confidence_rationale":"Tier 2 — isoform-specific dissection in vitro and KO in vivo with clear mechanistic readouts","pmids":["22891218"],"is_preprint":false},{"year":2012,"finding":"The membrane-bound form of Dlk1, but not soluble Dlk1, is required for its negative regulatory effect on hematopoietic stem and progenitor cell activity in the aorta-gonad-mesonephros region; this was demonstrated in Dlk1 KO and overexpressing mice and recapitulated in co-cultures with stromal cells expressing different Dlk1 levels.","method":"Dlk1 knockout and overexpression mouse models, co-culture experiments, FACS-based hematopoietic stem cell quantification","journal":"Haematologica","confidence":"High","confidence_rationale":"Tier 2 — KO and OE with isoform dissection in vivo and in vitro","pmids":["22801971"],"is_preprint":false},{"year":2012,"finding":"DLK1 is expressed predominantly as a soluble protein (cleaved extracellular domain) in adult mouse hypothalamus and localizes to soma and dendrites of arginine-vasopressin and oxytocin neurons in multiple hypothalamic nuclei, suggesting a neuroendocrine role.","method":"Western blot of hypothalamic fractions, immunohistochemistry, in situ hybridization","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2/3 — direct localization by fractionation and immunohistochemistry, single lab","pmids":["22563444"],"is_preprint":false},{"year":2014,"finding":"DLK1 promotes lung cancer cell invasion by upregulating MMP9 expression through activation of Notch signaling (NOTCH1/NICD nuclear translocation, HES1); γ-secretase inhibitor (GSI) blocked DLK1-induced MMP9 upregulation.","method":"DLK1 overexpression/knockdown in lung cancer cell lines, transwell invasion assay, Western blot, gelatin zymography, GSI pharmacological inhibition","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 — mechanistic pathway dissection with pharmacological inhibition, single lab","pmids":["24621612"],"is_preprint":false},{"year":2014,"finding":"Membrane-bound Dlk1 promotes myogenic hypertrophy and fusion in C2C12 cells, whereas soluble Dlk1 inhibits myotube formation, demonstrating isoform-specific opposing roles in myogenesis.","method":"C2C12 cell lines stably expressing membrane-bound vs. soluble Dlk1, fusion index quantification, gene expression analysis","journal":"FEBS letters","confidence":"Medium","confidence_rationale":"Tier 2 — direct isoform-specific functional comparison, single lab","pmids":["24582655"],"is_preprint":false},{"year":2014,"finding":"DLK1/Pref-1 overexpression causes reduced fat stores, pituitary IGF1 resistance, impaired GH feedback regulation, and increased circulating GH, which shifts whole-body fuel metabolism toward peripheral lipid oxidation and reduces hepatic steatosis; these effects were demonstrated by overexpressing Dlk1 from endogenous control elements in mice.","method":"Transgenic mouse overexpression from endogenous elements, metabolic phenotyping, hormone measurements, glucose/insulin tolerance tests","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 — in vivo transgenic model with multiple orthogonal metabolic and endocrine readouts","pmids":["25349437"],"is_preprint":false},{"year":2014,"finding":"Pref-1+ cells are very early mesenchymal adipose precursors that precede Zfp423+ and PPARγ+ cells in the adipogenic hierarchy; ablation of Pref-1-marked cells prevents embryonic white adipose tissue development and adult adipose expansion upon high-fat feeding.","method":"Transgenic mouse lineage tracing with Pref-1 promoter-driven fluorescent labels and inducible cell ablation","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 — in vivo lineage tracing and genetic cell ablation with clear developmental and physiological phenotypes","pmids":["25088414"],"is_preprint":false},{"year":2015,"finding":"Exogenous soluble DLK1 (Fc-DLK1 fusion) reduces hepatic steatosis and hyperglycemia in db/db mice by activating AMPK and ACC phosphorylation, suppressing SREBP-1c, lowering PEPCK/G6Pase expression, and inducing AKT-mediated cytosolic translocation of FOXO1; the gluconeogenic effect was blocked by an AMPK inhibitor.","method":"Recombinant soluble DLK1 treatment of db/db mice and primary hepatocytes, AMPK inhibitor co-administration, Western blot, glucose production assay","journal":"International journal of obesity (2005)","confidence":"Medium","confidence_rationale":"Tier 2 — in vivo and in vitro with pharmacological inhibitor validation, single lab","pmids":["26315841"],"is_preprint":false},{"year":2016,"finding":"DLK1 directly interacts with NOTCH1 in mammals; the interaction occurs between EGF domains 5 and 6 of DLK1 and EGF domains 10-15 of NOTCH1, demonstrated by mammalian two-hybrid system; DLK1 reduces NOTCH1 activation in Dlk1+/+ vs Dlk1-/- embryos and in siRNA-manipulated cell lines.","method":"Mammalian two-hybrid system, domain deletion mapping, comparison of NOTCH1 activation in Dlk1+/+ vs Dlk1-/- E16.5 mouse tissues, siRNA knockdown in mammalian cell line","journal":"Cellular signalling","confidence":"High","confidence_rationale":"Tier 2 — direct protein-protein interaction mapped to specific EGF domains, validated in vivo and in vitro","pmids":["26791579"],"is_preprint":false},{"year":2017,"finding":"Loss-of-function genomic defects in DLK1 (paternally inherited) cause familial central precocious puberty; affected individuals have undetectable serum DLK1 and Dlk1 is expressed in mouse hypothalamus and kisspeptin neuron-derived cell lines, implicating DLK1 in the neuroendocrine control of pubertal timing.","method":"Linkage analysis, whole-genome sequencing, serum DLK1 ELISA, in situ hybridization in mouse hypothalamus, Dlk1 expression in kisspeptin cell lines","journal":"The Journal of clinical endocrinology and metabolism","confidence":"High","confidence_rationale":"Tier 2 — human loss-of-function genetics with direct in vivo expression data and serum protein measurement","pmids":["28324015"],"is_preprint":false},{"year":2018,"finding":"Soluble DLK1 (extracellular domain) stimulates angiogenesis by activating Notch1/Akt/eNOS/Hes-1 signaling in endothelial cells; γ-secretase inhibitor or Notch1 knockdown/antibody neutralization reversed DLK1-EC-induced endothelial migration and angiogenesis.","method":"Recombinant DLK1-EC treatment of endothelial cells and aorta ring sprouting assay, corneal neovascularization in rats, Western blot/luciferase reporter for Notch1 pathway, pharmacological and siRNA inhibition of Notch1","journal":"Angiogenesis","confidence":"Medium","confidence_rationale":"Tier 2 — in vitro and in vivo assays with pathway inhibitor validation, single lab","pmids":["29383634"],"is_preprint":false},{"year":2018,"finding":"Sox9 inactivation in Pref-1+ cells converts them to PDGFRα+ cells expressing early adipogenic markers, demonstrating that Sox9 maintains the Pref-1+ early precursor state and that Sox9 inactivation is required for white adipose tissue expansion; Pref-1+ cells precede PDGFRα+ cells in the adipogenic hierarchy.","method":"Pref-1 promoter-rtTA system for inducible Sox9 inactivation in mice, lineage tracing, FACS, gene expression","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 — inducible in vivo genetic inactivation with clear precursor hierarchy placement","pmids":["30355480"],"is_preprint":false},{"year":2020,"finding":"Hypoxia induces ADAM17-dependent cleavage of DLK1 and release of an intracellular fragment that translocates to the nucleus in glioma cells; HIF-1α/HIF-2α are required for this cleavage. Expression of cleavable DLK1 promotes colony formation, stem cell marker expression, PI3K-pathway-mediated metabolic shift, and invasion; non-cleavable DLK1 does not replicate these effects.","method":"Immunofluorescence of DLK1 nuclear translocation, ADAM17 inhibitors and HIF siRNA to block cleavage, cleavable vs. non-cleavable DLK1 constructs, PDGFB-induced glioma mouse model","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 2 — mechanistic dissection with pharmacological/genetic inhibition and cleavage-defective mutant, validated in vivo","pmids":["32205867"],"is_preprint":false},{"year":2021,"finding":"Dlk1 is expressed biallelically (loss of canonical imprinting) in the subgranular zone of the hippocampus; both parental alleles are required for normal adult neurogenesis and cognitive function (spatial discrimination), demonstrated by allele-specific Dlk1 mutant mice.","method":"Allele-specific expression analysis, maternal/paternal/biallelic Dlk1 mutant mouse behavioral battery, adult neurogenesis quantification","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 — allele-specific genetic manipulation with behavioral and cellular readouts, single study","pmids":["33712542"],"is_preprint":false},{"year":2005,"finding":"Intracellular domain interactions of DLK1 are critical for inhibiting differentiation and proliferation of hematopoietic HL-60 cells; unlike in preadipocytes, proteolytic release of the extracellular domain is not required for DLK1 function in hematopoietic cells.","method":"Ectopic expression of DLK1 with domain mutations in HL-60 cells, differentiation and proliferation assays","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 — domain-specific dissection demonstrating differential mechanism vs. adipocytes, single lab","pmids":["15806146"],"is_preprint":false},{"year":2014,"finding":"DLK1 loss-of-function in neuroblastoma xenografts enhances tumor cell differentiation likely by increasing basal MEK/ERK phosphorylation; DLK1+ cells are preferentially located in hypoxic tumor regions.","method":"Loss-of-function DLK1 mutants in neuroblastoma xenografts, MEK/ERK phosphorylation analysis, hypoxia immunostaining","journal":"Cancer letters","confidence":"Medium","confidence_rationale":"Tier 2 — in vivo xenograft KD with signaling readout, single lab","pmids":["22142700"],"is_preprint":false},{"year":2014,"finding":"DLK1 inhibits branching morphogenesis of submandibular salivary glands and parasympathetic innervation of epithelial end buds through inhibition of NOTCH signaling; soluble DLK1 and the γ-secretase inhibitor DAPT phenocopied each other in organotypic cultures, reducing branching, increasing epithelial apoptosis, and impairing parasympathetic ganglion outgrowth.","method":"Organotypic submandibular gland cultures with soluble DLK1 or DAPT treatment, rescue with carbachol, immunohistochemistry","journal":"Biology of the cell","confidence":"Medium","confidence_rationale":"Tier 2 — pharmacological phenocopy between DLK1 and Notch inhibitor with organotypic culture, single lab","pmids":["24828459"],"is_preprint":false},{"year":2024,"finding":"DLK1 is robustly expressed on the cell surface of neuroblastoma cells; shRNA-mediated silencing of DLK1 in neuroblastoma cells increases cellular differentiation; a DLK1-targeting antibody-drug conjugate (ADCT-701) shows potent cytotoxicity in DLK1-expressing neuroblastoma xenograft models.","method":"Proteogenomic analysis, immunofluorescence, flow cytometry, IHC, shRNA silencing with differentiation assay, ADC xenograft efficacy study","journal":"Cancer cell","confidence":"Medium","confidence_rationale":"Tier 2 — KD with functional readout and in vivo therapeutic validation, single study","pmids":["39454577"],"is_preprint":false},{"year":2010,"finding":"An insulator element 18 kb upstream of DLK1, bound by CTCF, regulates monoallelic (imprinted) DLK1 expression; hypermethylation of this region on both alleles in AML disrupts CTCF binding and results in biallelic DLK1 overexpression.","method":"SNP allele-specific expression analysis, quantitative methylation analysis, allele-specific methylation analysis, chromatin immunoprecipitation for CTCF","journal":"Blood","confidence":"Medium","confidence_rationale":"Tier 2 — ChIP demonstrating CTCF binding with allele-specific methylation correlation, single lab","pmids":["20089961"],"is_preprint":false}],"current_model":"DLK1 (Pref-1) is a paternally imprinted transmembrane protein with six EGF-like repeats that is proteolytically cleaved (by TACE/ADAM17) to release a biologically active soluble ectodomain; membrane-bound and soluble isoforms have distinct functions — the soluble form inhibits adipogenic differentiation by activating MEK/ERK to maintain Sox9 expression (suppressing C/EBPβ/δ), while the membrane-bound form additionally inhibits preadipocyte and hematopoietic stem cell proliferation; DLK1 acts as a non-canonical inhibitory ligand of NOTCH1 through direct interaction between its EGF domains 5–6 and NOTCH1 EGF domains 10–15; in hypoxic contexts, ADAM17/HIF-driven cleavage releases an intracellular DLK1 fragment that translocates to the nucleus to promote stem-like and invasive phenotypes; loss-of-function DLK1 mutations cause central precocious puberty and metabolic dysregulation in humans, and DLK1 is required for normal skeletal muscle development, hippocampal neurogenesis, and B-cell development in mice."},"narrative":{"teleology":[{"year":1993,"claim":"Identification of DLK1 as an EGF-repeat transmembrane protein that blocks adipogenesis established the first functional role for this gene as a negative regulator of differentiation.","evidence":"cDNA cloning and constitutive overexpression in 3T3-L1 preadipocytes with differentiation assay","pmids":["8500166"],"confidence":"High","gaps":["receptor or binding partner unknown","mechanism of differentiation inhibition uncharacterized","in vivo relevance not tested"]},{"year":1997,"claim":"Demonstration that DLK1 is proteolytically cleaved to release a biologically active soluble ectodomain, and that alternative splicing governs whether cleavage occurs, resolved how DLK1 can function in both juxtacrine and paracrine modes.","evidence":"In vitro cleavage assays, recombinant ectodomain addition and antibody neutralization in 3T3-L1 cells, splice-form analysis","pmids":["9001251"],"confidence":"High","gaps":["identity of the protease performing cleavage unknown","whether membrane-bound vs. soluble forms have distinct downstream signaling not yet tested"]},{"year":1998,"claim":"Mapping of a cis-regulatory SAD element required for transcriptional silencing of DLK1 during differentiation revealed how DLK1 expression is shut off to permit adipogenesis.","evidence":"5'-deletion reporter constructs, gel shift, UV cross-linking, and mutagenesis in 3T3-L1 cells","pmids":["9822638"],"confidence":"High","gaps":["identity of the ~63 kDa SAD-binding protein unknown","whether this element operates in non-adipose lineages untested"]},{"year":1999,"claim":"Demonstration that dexamethasone promotes adipogenesis by transcriptionally repressing DLK1 connected glucocorticoid signaling to the DLK1 regulatory axis.","evidence":"Nuclear run-on, mRNA stability assays, and antisense transfection in 3T3-L1 cells","pmids":["10212243"],"confidence":"High","gaps":["glucocorticoid receptor binding at DLK1 promoter not directly shown","relevance to in vivo glucocorticoid-induced obesity not tested"]},{"year":2004,"claim":"DLK1 was shown to maintain bipotential mesenchymal progenitor identity by inhibiting both osteoblast and adipocyte differentiation, broadening its role beyond adipogenesis; separately, DLK1 was established as the causal gene for callipyge muscular hypertrophy.","evidence":"Retroviral overexpression in human MSCs with cytochemistry, FACS, and ex vivo calvaria culture; transgenic DLK1 expression in mouse skeletal muscle and IHC in callipyge sheep","pmids":["15068508","15498495"],"confidence":"High","gaps":["molecular mechanism of osteoblast inhibition not dissected","downstream myogenic signaling targets unknown"]},{"year":2006,"claim":"Showing that DLK1's extracellular domain inhibits Notch signaling in mesenchymal cells provided the first direct link between DLK1 and the Notch pathway, though the outcome (pro- vs. anti-adipogenic) proved cell-type-dependent.","evidence":"Domain-deletion constructs, co-culture, ERK1/2 activation, and Notch1 knockdown in C3H10T1/2 cells","pmids":["17320900"],"confidence":"Medium","gaps":["direct physical interaction between DLK1 and Notch1 not yet demonstrated","basis for cell-type-specific divergent outcomes unclear"]},{"year":2009,"claim":"Consolidation of the MEK/ERK–Sox9–C/EBPβ/δ signaling axis downstream of soluble DLK1 provided a coherent molecular mechanism for adipogenesis inhibition.","evidence":"Synthesis of MAPK assays, Sox9 overexpression/knockdown, and C/EBP reporter assays from multiple studies","pmids":["19541743"],"confidence":"Medium","gaps":["direct receptor for soluble DLK1 upstream of MEK/ERK not identified","pathway validated mainly in 3T3-L1 model"]},{"year":2010,"claim":"Conditional knockout revealed that Dlk1 is required for normal skeletal muscle development and satellite cell regulation, and that CTCF-bound insulator elements control imprinted DLK1 expression with loss of imprinting in AML.","evidence":"Muscle-specific Dlk1 KO/OE in mice with satellite cell culture; allele-specific expression, methylation, and CTCF ChIP in AML samples","pmids":["21124733","20089961"],"confidence":"High","gaps":["downstream targets of Dlk1 in satellite cells beyond NF-κB not defined","functional consequence of biallelic expression in AML not directly tested"]},{"year":2012,"claim":"Isoform-specific dissection revealed that membrane-bound DLK1 uniquely inhibits preadipocyte and hematopoietic stem cell proliferation, whereas soluble DLK1 primarily inhibits differentiation, establishing that cleavage status dictates functional mode.","evidence":"Independent isoform manipulation in preadipocytes with cell cycle analysis and DLK1-null mice; Dlk1 KO/OE mice and stromal co-cultures for hematopoietic readouts","pmids":["22891218","22801971"],"confidence":"High","gaps":["mechanism by which membrane-bound DLK1 arrests G1/S progression unknown","whether isoform-specific effects extend to other tissues untested"]},{"year":2014,"claim":"Multiple studies mapped DLK1 function in diverse contexts: Pref-1+ cells were placed as the earliest mesenchymal adipose precursors preceding Zfp423+ and PPARγ+ cells; DLK1 overexpression was shown to cause pituitary IGF1 resistance and shift metabolism toward lipid oxidation; and DLK1 was linked to Notch inhibition in salivary gland organogenesis.","evidence":"Lineage tracing with Pref-1 promoter and inducible cell ablation in mice; transgenic DLK1 overexpression with metabolic and endocrine phenotyping; organotypic gland cultures with DLK1 and DAPT","pmids":["25088414","25349437","24828459"],"confidence":"High","gaps":["receptor mediating DLK1 metabolic effects in liver/pituitary not identified","how DLK1 interacts with GH axis mechanistically unclear"]},{"year":2016,"claim":"Direct physical interaction between DLK1 EGF domains 5–6 and NOTCH1 EGF domains 10–15 was mapped, providing the structural basis for DLK1-mediated Notch inhibition.","evidence":"Mammalian two-hybrid domain deletion mapping, Notch1 activation comparison in Dlk1+/+ vs Dlk1−/− E16.5 tissues, siRNA in cell lines","pmids":["26791579"],"confidence":"High","gaps":["no biophysical or structural characterization of the interaction","whether DLK1 competes with canonical Notch ligands at these domains not directly tested"]},{"year":2017,"claim":"Human genetics established that paternally inherited DLK1 loss-of-function causes familial central precocious puberty with undetectable serum DLK1, linking DLK1 to neuroendocrine pubertal timing control.","evidence":"Linkage analysis, whole-genome sequencing of affected families, serum DLK1 ELISA, DLK1 expression in mouse hypothalamus and kisspeptin neuron cell lines","pmids":["28324015"],"confidence":"High","gaps":["molecular mechanism by which DLK1 regulates GnRH/kisspeptin axis unknown","whether Notch inhibition mediates the pubertal phenotype untested"]},{"year":2020,"claim":"Discovery of a hypoxia-driven, ADAM17/HIF-dependent cleavage event that releases a DLK1 intracellular fragment translocating to the nucleus revealed a non-canonical signaling mode promoting stemness and invasion in glioma.","evidence":"ADAM17 inhibitors, HIF siRNA, cleavable vs. non-cleavable DLK1 constructs, nuclear immunofluorescence, PDGFB-induced glioma mouse model","pmids":["32205867"],"confidence":"High","gaps":["nuclear targets of the DLK1 intracellular fragment not identified","whether this mechanism operates outside glioma unknown","structural determinants of intracellular fragment release vs. ectodomain shedding unclear"]},{"year":2021,"claim":"Biallelic expression of Dlk1 in the hippocampal subgranular zone, escaping canonical imprinting, was shown to be required for normal adult neurogenesis and spatial discrimination, demonstrating tissue-specific regulation of imprinting.","evidence":"Allele-specific expression analysis, maternal/paternal/biallelic Dlk1 mutant mice with behavioral and neurogenesis quantification","pmids":["33712542"],"confidence":"Medium","gaps":["mechanism of imprint escape in hippocampus not defined","downstream neurogenic targets of Dlk1 in subgranular zone unknown"]},{"year":null,"claim":"The identity of the cell-surface receptor that transduces soluble DLK1 signaling to MEK/ERK and AMPK pathways remains unknown, and the nuclear targets of the DLK1 intracellular fragment have not been identified.","evidence":"","pmids":[],"confidence":"High","gaps":["no receptor for soluble DLK1 identified","nuclear binding partners/targets of DLK1 ICD unknown","structural basis of DLK1–NOTCH1 interaction not resolved at atomic level"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,6,17,25]},{"term_id":"GO:0048018","term_label":"receptor ligand activity","supporting_discovery_ids":[1,7,16,19]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[0,1,9,10,26]},{"term_id":"GO:0005576","term_label":"extracellular region","supporting_discovery_ids":[1,7,11,16,19]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[21]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[6,7,12,17,19,25]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[5,8,15,20]},{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[14,16]}],"complexes":[],"partners":["NOTCH1","ADAM17","SOX9","CTCF"],"other_free_text":[]},"mechanistic_narrative":"DLK1 (Pref-1) is a paternally expressed, imprinted transmembrane protein containing six EGF-like repeats that functions as a gatekeeper of progenitor cell identity, inhibiting terminal differentiation of adipocytes, osteoblasts, myoblasts, and hematopoietic cells while regulating proliferation in a context- and isoform-dependent manner [PMID:8500166, PMID:15068508, PMID:22801971, PMID:22891218]. Proteolytic cleavage releases a soluble ectodomain that inhibits adipogenesis by activating MEK/ERK signaling to sustain Sox9 expression, thereby suppressing C/EBPβ/δ [PMID:9001251, PMID:19541743, PMID:30355480], whereas the membrane-bound form additionally inhibits preadipocyte and hematopoietic stem cell proliferation through cell-autonomous mechanisms [PMID:22891218, PMID:22801971]; DLK1 directly interacts with NOTCH1 via its EGF domains 5–6 to modulate Notch signaling outputs in a cell-type-specific manner [PMID:26791579, PMID:17320900]. Under hypoxia, ADAM17/HIF-dependent cleavage generates a nuclear-translocating intracellular fragment that promotes stem-like and invasive phenotypes in glioma [PMID:32205867]. Paternally inherited loss-of-function mutations in DLK1 cause familial central precocious puberty with undetectable serum DLK1 [PMID:28324015]."},"prefetch_data":{"uniprot":{"accession":"P80370","full_name":"Protein delta homolog 1","aliases":["pG2"],"length_aa":383,"mass_kda":41.3,"function":"May have a role in neuroendocrine differentiation","subcellular_location":"Membrane; Cytoplasm","url":"https://www.uniprot.org/uniprotkb/P80370/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/DLK1","classification":"Not Classified","n_dependent_lines":16,"n_total_lines":1208,"dependency_fraction":0.013245033112582781},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/DLK1","total_profiled":1310},"omim":[{"mim_id":"621120","title":"DELTA-LIKE NONCANONICAL NOTCH LIGAND 2; 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cell","url":"https://pubmed.ncbi.nlm.nih.gov/24828459","citation_count":25,"is_preprint":false},{"pmid":"9888532","id":"PMC_9888532","title":"Implication of ZOG protein (zona glomerulosa-specific protein) in zone development of the adrenal cortex.","date":"1998","source":"Endocrine research","url":"https://pubmed.ncbi.nlm.nih.gov/9888532","citation_count":24,"is_preprint":false},{"pmid":"28715922","id":"PMC_28715922","title":"DLK1-DIO3 imprinted locus deregulation in development, respiratory disease, and cancer.","date":"2017","source":"Expert review of respiratory medicine","url":"https://pubmed.ncbi.nlm.nih.gov/28715922","citation_count":24,"is_preprint":false},{"pmid":"34710357","id":"PMC_34710357","title":"A bipartite element with allele-specific functions safeguards DNA methylation imprints at the Dlk1-Dio3 locus.","date":"2021","source":"Developmental cell","url":"https://pubmed.ncbi.nlm.nih.gov/34710357","citation_count":24,"is_preprint":false},{"pmid":"32924531","id":"PMC_32924531","title":"Astragalus Polysaccharide (PG2) Suppresses Macrophage Migration Inhibitory Factor and Aggressiveness of Lung Adenocarcinoma Cells.","date":"2020","source":"The American journal of Chinese medicine","url":"https://pubmed.ncbi.nlm.nih.gov/32924531","citation_count":23,"is_preprint":false},{"pmid":"20058200","id":"PMC_20058200","title":"Isolation and differentiation of chondrocytic cells derived from human embryonic stem cells using dlk1/FA1 as a novel surface marker.","date":"2009","source":"Stem cell reviews and reports","url":"https://pubmed.ncbi.nlm.nih.gov/20058200","citation_count":22,"is_preprint":false},{"pmid":"17451526","id":"PMC_17451526","title":"Expression of ksdD gene encoding 3-ketosteroid-Delta1-dehydrogenase from Arthrobacter simplex in Bacillus subtilis.","date":"2007","source":"Letters in applied microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/17451526","citation_count":21,"is_preprint":false},{"pmid":"28549249","id":"PMC_28549249","title":"CD146 (MCAM) in human cs-DLK1-/cs-CD34+ adipose stromal/progenitor cells.","date":"2017","source":"Stem cell research","url":"https://pubmed.ncbi.nlm.nih.gov/28549249","citation_count":21,"is_preprint":false},{"pmid":"25551289","id":"PMC_25551289","title":"Antagonistic roles in fetal development and adult physiology for the oppositely imprinted Grb10 and Dlk1 genes.","date":"2014","source":"BMC biology","url":"https://pubmed.ncbi.nlm.nih.gov/25551289","citation_count":21,"is_preprint":false},{"pmid":"31118676","id":"PMC_31118676","title":"MicroRNA-143-3p suppresses tumorigenesis by targeting catenin-δ1 in colorectal cancer.","date":"2019","source":"OncoTargets and therapy","url":"https://pubmed.ncbi.nlm.nih.gov/31118676","citation_count":21,"is_preprint":false},{"pmid":"24582655","id":"PMC_24582655","title":"Membrane-bound delta-like 1 homolog (Dlk1) promotes while soluble Dlk1 inhibits myogenesis in C2C12 cells.","date":"2014","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/24582655","citation_count":20,"is_preprint":false},{"pmid":"30697070","id":"PMC_30697070","title":"Dysregulation of ncRNAs located at the DLK1‑DIO3 imprinted domain: involvement in urological cancers.","date":"2019","source":"Cancer management and research","url":"https://pubmed.ncbi.nlm.nih.gov/30697070","citation_count":20,"is_preprint":false},{"pmid":"39454577","id":"PMC_39454577","title":"A proteogenomic surfaceome study identifies DLK1 as an immunotherapeutic target in neuroblastoma.","date":"2024","source":"Cancer cell","url":"https://pubmed.ncbi.nlm.nih.gov/39454577","citation_count":19,"is_preprint":false},{"pmid":"21637593","id":"PMC_21637593","title":"Expression of DLK1 and MEG3 genes in porcine tissues during postnatal development.","date":"2010","source":"Genetics and molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/21637593","citation_count":18,"is_preprint":false},{"pmid":"30315226","id":"PMC_30315226","title":"Glutamate delta-1 receptor regulates cocaine-induced plasticity in the nucleus accumbens.","date":"2018","source":"Translational psychiatry","url":"https://pubmed.ncbi.nlm.nih.gov/30315226","citation_count":18,"is_preprint":false},{"pmid":"17471553","id":"PMC_17471553","title":"Expansion and characterization of ventral mesencephalic precursor cells: effect of mitogens and investigation of FA1 as a potential dopaminergic marker.","date":"2007","source":"Journal of neuroscience research","url":"https://pubmed.ncbi.nlm.nih.gov/17471553","citation_count":18,"is_preprint":false},{"pmid":"18344616","id":"PMC_18344616","title":"Synergistic role of Igf2 and Dlk1 in fetal liver development and hematopoiesis in bi-maternal mice.","date":"2008","source":"The Journal of reproduction and development","url":"https://pubmed.ncbi.nlm.nih.gov/18344616","citation_count":18,"is_preprint":false},{"pmid":"30268817","id":"PMC_30268817","title":"Deletion of Dlk1 increases the vulnerability to developing anxiety-like behaviors and ethanol consumption in mice.","date":"2018","source":"Biochemical pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/30268817","citation_count":18,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":48640,"output_tokens":6638,"usd":0.122745},"stage2":{"model":"claude-opus-4-6","input_tokens":10326,"output_tokens":3573,"usd":0.211432},"total_usd":0.334177,"stage1_batch_id":"msgbatch_0118RScg3WGaGkaZRtWZbtce","stage2_batch_id":"msgbatch_01Cpfzvtbj42aqp5oYZ5Te2s","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1993,\n      \"finding\": \"Pref-1 (DLK1) is synthesized as a transmembrane protein with six tandem EGF-like repeats and functions as a negative regulator of adipocyte differentiation; constitutive expression in preadipocytes blocks adipogenesis.\",\n      \"method\": \"cDNA cloning, constitutive overexpression in 3T3-L1 preadipocytes with differentiation assay\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — foundational gain-of-function study in cell culture, replicated across many subsequent studies\",\n      \"pmids\": [\"8500166\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"Membrane-associated Pref-1 (DLK1) is cleaved in the juxtamembrane region to generate a soluble ~50 kDa ectodomain that is biologically active and inhibits adipocyte differentiation; only the two largest alternatively spliced isoforms undergo this cleavage, establishing that alternative splicing governs juxtacrine vs. paracrine modes of Pref-1 action.\",\n      \"method\": \"In vitro cleavage assays, addition of recombinant ectodomain to 3T3-L1 cells, antibody neutralization, alternate splice-form analysis\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1/2 — reconstitution with purified ectodomain plus antibody blocking and splice-form dissection in single rigorous study\",\n      \"pmids\": [\"9001251\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Dexamethasone promotes adipogenesis by transcriptionally repressing the Pref-1 (DLK1) gene; nuclear run-on assays showed reduced Pref-1 transcription, and antisense Pref-1 transfection enhanced adipogenesis at low dexamethasone concentrations, placing Pref-1 downregulation as a key mechanism of glucocorticoid-driven adipoconversion.\",\n      \"method\": \"Nuclear run-on transcription assay, actinomycin D mRNA stability assay, stable antisense transfection with differentiation assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1/2 — multiple orthogonal methods (run-on, stability, antisense) in a single study\",\n      \"pmids\": [\"10212243\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"A specific cis-element in the Pref-1 (DLK1) promoter, the SAD (suppression in adipocyte differentiation) element (core sequence -183AAAGA-179), is required for differentiation-dependent transcriptional suppression of the Pref-1 gene; an ~63 kDa nuclear protein binds this element.\",\n      \"method\": \"Stable transfection of 5'-deletion reporter constructs, gel mobility shift assay, UV cross-linking, base-substitution mutagenesis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — deletion mapping + mutagenesis + DNA-protein interaction assays\",\n      \"pmids\": [\"9822638\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Constitutive overexpression of DLK1/Pref-1 in human mesenchymal stem cells inhibits both osteoblast and adipocyte differentiation without affecting proliferation, identifying DLK1 as a regulator that maintains the bipotential progenitor pool.\",\n      \"method\": \"Retroviral overexpression in hMSC-TERT cells, cytochemical staining, FACS, real-time PCR, ex vivo calvaria organ culture\",\n      \"journal\": \"Journal of bone and mineral research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal readouts (in vitro and ex vivo) in human cells\",\n      \"pmids\": [\"15068508\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Ectopic expression of DLK1 protein in skeletal muscle of callipyge sheep (+(MAT)/CLPG(PAT)) causes muscular hypertrophy; transgenic mice expressing DLK1 in skeletal muscle develop generalized muscular hypertrophy, establishing DLK1 as the causal agent of the callipyge phenotype.\",\n      \"method\": \"Immunohistochemistry in sheep muscle, transgenic mouse model with skeletal muscle-specific DLK1 expression\",\n      \"journal\": \"Current biology : CB\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — in vivo transgenic rescue/overexpression with clear phenotypic readout, cross-validated in two species\",\n      \"pmids\": [\"15498495\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"The EGF-like extracellular domain of DLK1 inhibits Notch signaling in mesenchymal C3H10T1/2 cells, but this Notch inhibition paradoxically potentiates adipogenesis in these cells (unlike 3T3-L1 cells); the effect requires Notch1 expression and is mediated by the extracellular, not the intracellular, domain of DLK1.\",\n      \"method\": \"Overexpression of full-length vs. domain-deletion DLK1 constructs, co-culture assay, ERK1/2 activation measurement, Notch1 knockdown\",\n      \"journal\": \"Journal of molecular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — domain dissection and co-culture experiments, single lab\",\n      \"pmids\": [\"17320900\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Soluble Pref-1 (DLK1) activates the MEK/ERK pathway; ERK activation by Pref-1 prevents downregulation of Sox9, which in turn suppresses C/EBPβ and C/EBPδ expression and blocks adipogenesis. Pref-1 also induces Sox9 to promote chondrogenic lineage commitment while inhibiting chondrocyte maturation and osteoblast differentiation.\",\n      \"method\": \"Review synthesizing signaling pathway data from multiple mechanistic studies (MAPK assays, Sox9 overexpression/knockdown, C/EBP reporter assays)\",\n      \"journal\": \"Molecular endocrinology (Baltimore, Md.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — pathway placement based on multiple prior experimental studies consolidated in review; MEK/ERK-Sox9 axis experimentally supported\",\n      \"pmids\": [\"19541743\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Dlk1 is required for normal skeletal muscle development and regeneration; muscle-specific Dlk1 knockout reduces myofiber number and MyoD expression, impairs regeneration with augmented NF-κB and inflammatory cytokine signaling, and promotes satellite cell self-renewal; conversely, Dlk1 overexpression inhibits satellite cell proliferation and enhances differentiation.\",\n      \"method\": \"Conditional knockout and overexpression in mice, satellite cell single-fiber culture, immunofluorescence, gene expression analysis\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — conditional KO plus overexpression with multiple cellular and molecular readouts in vivo and ex vivo\",\n      \"pmids\": [\"21124733\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Membrane-bound DLK1 (DLK1-M), but not soluble DLK1, inhibits G1-to-S-phase cell cycle progression and preadipocyte proliferation; DLK1-null mice show increased preadipocyte replication and higher fat mass, establishing a dual inhibitory role of DLK1 on both proliferation and differentiation in adipogenesis.\",\n      \"method\": \"Independent manipulation of DLK1 isoform levels in preadipocytes, cell cycle analysis, in vivo analysis of DLK1-null mice\",\n      \"journal\": \"Diabetes\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — isoform-specific dissection in vitro and KO in vivo with clear mechanistic readouts\",\n      \"pmids\": [\"22891218\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"The membrane-bound form of Dlk1, but not soluble Dlk1, is required for its negative regulatory effect on hematopoietic stem and progenitor cell activity in the aorta-gonad-mesonephros region; this was demonstrated in Dlk1 KO and overexpressing mice and recapitulated in co-cultures with stromal cells expressing different Dlk1 levels.\",\n      \"method\": \"Dlk1 knockout and overexpression mouse models, co-culture experiments, FACS-based hematopoietic stem cell quantification\",\n      \"journal\": \"Haematologica\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — KO and OE with isoform dissection in vivo and in vitro\",\n      \"pmids\": [\"22801971\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"DLK1 is expressed predominantly as a soluble protein (cleaved extracellular domain) in adult mouse hypothalamus and localizes to soma and dendrites of arginine-vasopressin and oxytocin neurons in multiple hypothalamic nuclei, suggesting a neuroendocrine role.\",\n      \"method\": \"Western blot of hypothalamic fractions, immunohistochemistry, in situ hybridization\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2/3 — direct localization by fractionation and immunohistochemistry, single lab\",\n      \"pmids\": [\"22563444\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"DLK1 promotes lung cancer cell invasion by upregulating MMP9 expression through activation of Notch signaling (NOTCH1/NICD nuclear translocation, HES1); γ-secretase inhibitor (GSI) blocked DLK1-induced MMP9 upregulation.\",\n      \"method\": \"DLK1 overexpression/knockdown in lung cancer cell lines, transwell invasion assay, Western blot, gelatin zymography, GSI pharmacological inhibition\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — mechanistic pathway dissection with pharmacological inhibition, single lab\",\n      \"pmids\": [\"24621612\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Membrane-bound Dlk1 promotes myogenic hypertrophy and fusion in C2C12 cells, whereas soluble Dlk1 inhibits myotube formation, demonstrating isoform-specific opposing roles in myogenesis.\",\n      \"method\": \"C2C12 cell lines stably expressing membrane-bound vs. soluble Dlk1, fusion index quantification, gene expression analysis\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — direct isoform-specific functional comparison, single lab\",\n      \"pmids\": [\"24582655\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"DLK1/Pref-1 overexpression causes reduced fat stores, pituitary IGF1 resistance, impaired GH feedback regulation, and increased circulating GH, which shifts whole-body fuel metabolism toward peripheral lipid oxidation and reduces hepatic steatosis; these effects were demonstrated by overexpressing Dlk1 from endogenous control elements in mice.\",\n      \"method\": \"Transgenic mouse overexpression from endogenous elements, metabolic phenotyping, hormone measurements, glucose/insulin tolerance tests\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — in vivo transgenic model with multiple orthogonal metabolic and endocrine readouts\",\n      \"pmids\": [\"25349437\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Pref-1+ cells are very early mesenchymal adipose precursors that precede Zfp423+ and PPARγ+ cells in the adipogenic hierarchy; ablation of Pref-1-marked cells prevents embryonic white adipose tissue development and adult adipose expansion upon high-fat feeding.\",\n      \"method\": \"Transgenic mouse lineage tracing with Pref-1 promoter-driven fluorescent labels and inducible cell ablation\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — in vivo lineage tracing and genetic cell ablation with clear developmental and physiological phenotypes\",\n      \"pmids\": [\"25088414\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Exogenous soluble DLK1 (Fc-DLK1 fusion) reduces hepatic steatosis and hyperglycemia in db/db mice by activating AMPK and ACC phosphorylation, suppressing SREBP-1c, lowering PEPCK/G6Pase expression, and inducing AKT-mediated cytosolic translocation of FOXO1; the gluconeogenic effect was blocked by an AMPK inhibitor.\",\n      \"method\": \"Recombinant soluble DLK1 treatment of db/db mice and primary hepatocytes, AMPK inhibitor co-administration, Western blot, glucose production assay\",\n      \"journal\": \"International journal of obesity (2005)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — in vivo and in vitro with pharmacological inhibitor validation, single lab\",\n      \"pmids\": [\"26315841\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"DLK1 directly interacts with NOTCH1 in mammals; the interaction occurs between EGF domains 5 and 6 of DLK1 and EGF domains 10-15 of NOTCH1, demonstrated by mammalian two-hybrid system; DLK1 reduces NOTCH1 activation in Dlk1+/+ vs Dlk1-/- embryos and in siRNA-manipulated cell lines.\",\n      \"method\": \"Mammalian two-hybrid system, domain deletion mapping, comparison of NOTCH1 activation in Dlk1+/+ vs Dlk1-/- E16.5 mouse tissues, siRNA knockdown in mammalian cell line\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — direct protein-protein interaction mapped to specific EGF domains, validated in vivo and in vitro\",\n      \"pmids\": [\"26791579\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Loss-of-function genomic defects in DLK1 (paternally inherited) cause familial central precocious puberty; affected individuals have undetectable serum DLK1 and Dlk1 is expressed in mouse hypothalamus and kisspeptin neuron-derived cell lines, implicating DLK1 in the neuroendocrine control of pubertal timing.\",\n      \"method\": \"Linkage analysis, whole-genome sequencing, serum DLK1 ELISA, in situ hybridization in mouse hypothalamus, Dlk1 expression in kisspeptin cell lines\",\n      \"journal\": \"The Journal of clinical endocrinology and metabolism\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — human loss-of-function genetics with direct in vivo expression data and serum protein measurement\",\n      \"pmids\": [\"28324015\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Soluble DLK1 (extracellular domain) stimulates angiogenesis by activating Notch1/Akt/eNOS/Hes-1 signaling in endothelial cells; γ-secretase inhibitor or Notch1 knockdown/antibody neutralization reversed DLK1-EC-induced endothelial migration and angiogenesis.\",\n      \"method\": \"Recombinant DLK1-EC treatment of endothelial cells and aorta ring sprouting assay, corneal neovascularization in rats, Western blot/luciferase reporter for Notch1 pathway, pharmacological and siRNA inhibition of Notch1\",\n      \"journal\": \"Angiogenesis\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — in vitro and in vivo assays with pathway inhibitor validation, single lab\",\n      \"pmids\": [\"29383634\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Sox9 inactivation in Pref-1+ cells converts them to PDGFRα+ cells expressing early adipogenic markers, demonstrating that Sox9 maintains the Pref-1+ early precursor state and that Sox9 inactivation is required for white adipose tissue expansion; Pref-1+ cells precede PDGFRα+ cells in the adipogenic hierarchy.\",\n      \"method\": \"Pref-1 promoter-rtTA system for inducible Sox9 inactivation in mice, lineage tracing, FACS, gene expression\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — inducible in vivo genetic inactivation with clear precursor hierarchy placement\",\n      \"pmids\": [\"30355480\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Hypoxia induces ADAM17-dependent cleavage of DLK1 and release of an intracellular fragment that translocates to the nucleus in glioma cells; HIF-1α/HIF-2α are required for this cleavage. Expression of cleavable DLK1 promotes colony formation, stem cell marker expression, PI3K-pathway-mediated metabolic shift, and invasion; non-cleavable DLK1 does not replicate these effects.\",\n      \"method\": \"Immunofluorescence of DLK1 nuclear translocation, ADAM17 inhibitors and HIF siRNA to block cleavage, cleavable vs. non-cleavable DLK1 constructs, PDGFB-induced glioma mouse model\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — mechanistic dissection with pharmacological/genetic inhibition and cleavage-defective mutant, validated in vivo\",\n      \"pmids\": [\"32205867\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Dlk1 is expressed biallelically (loss of canonical imprinting) in the subgranular zone of the hippocampus; both parental alleles are required for normal adult neurogenesis and cognitive function (spatial discrimination), demonstrated by allele-specific Dlk1 mutant mice.\",\n      \"method\": \"Allele-specific expression analysis, maternal/paternal/biallelic Dlk1 mutant mouse behavioral battery, adult neurogenesis quantification\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — allele-specific genetic manipulation with behavioral and cellular readouts, single study\",\n      \"pmids\": [\"33712542\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Intracellular domain interactions of DLK1 are critical for inhibiting differentiation and proliferation of hematopoietic HL-60 cells; unlike in preadipocytes, proteolytic release of the extracellular domain is not required for DLK1 function in hematopoietic cells.\",\n      \"method\": \"Ectopic expression of DLK1 with domain mutations in HL-60 cells, differentiation and proliferation assays\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — domain-specific dissection demonstrating differential mechanism vs. adipocytes, single lab\",\n      \"pmids\": [\"15806146\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"DLK1 loss-of-function in neuroblastoma xenografts enhances tumor cell differentiation likely by increasing basal MEK/ERK phosphorylation; DLK1+ cells are preferentially located in hypoxic tumor regions.\",\n      \"method\": \"Loss-of-function DLK1 mutants in neuroblastoma xenografts, MEK/ERK phosphorylation analysis, hypoxia immunostaining\",\n      \"journal\": \"Cancer letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — in vivo xenograft KD with signaling readout, single lab\",\n      \"pmids\": [\"22142700\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"DLK1 inhibits branching morphogenesis of submandibular salivary glands and parasympathetic innervation of epithelial end buds through inhibition of NOTCH signaling; soluble DLK1 and the γ-secretase inhibitor DAPT phenocopied each other in organotypic cultures, reducing branching, increasing epithelial apoptosis, and impairing parasympathetic ganglion outgrowth.\",\n      \"method\": \"Organotypic submandibular gland cultures with soluble DLK1 or DAPT treatment, rescue with carbachol, immunohistochemistry\",\n      \"journal\": \"Biology of the cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — pharmacological phenocopy between DLK1 and Notch inhibitor with organotypic culture, single lab\",\n      \"pmids\": [\"24828459\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"DLK1 is robustly expressed on the cell surface of neuroblastoma cells; shRNA-mediated silencing of DLK1 in neuroblastoma cells increases cellular differentiation; a DLK1-targeting antibody-drug conjugate (ADCT-701) shows potent cytotoxicity in DLK1-expressing neuroblastoma xenograft models.\",\n      \"method\": \"Proteogenomic analysis, immunofluorescence, flow cytometry, IHC, shRNA silencing with differentiation assay, ADC xenograft efficacy study\",\n      \"journal\": \"Cancer cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — KD with functional readout and in vivo therapeutic validation, single study\",\n      \"pmids\": [\"39454577\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"An insulator element 18 kb upstream of DLK1, bound by CTCF, regulates monoallelic (imprinted) DLK1 expression; hypermethylation of this region on both alleles in AML disrupts CTCF binding and results in biallelic DLK1 overexpression.\",\n      \"method\": \"SNP allele-specific expression analysis, quantitative methylation analysis, allele-specific methylation analysis, chromatin immunoprecipitation for CTCF\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — ChIP demonstrating CTCF binding with allele-specific methylation correlation, single lab\",\n      \"pmids\": [\"20089961\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"DLK1 (Pref-1) is a paternally imprinted transmembrane protein with six EGF-like repeats that is proteolytically cleaved (by TACE/ADAM17) to release a biologically active soluble ectodomain; membrane-bound and soluble isoforms have distinct functions — the soluble form inhibits adipogenic differentiation by activating MEK/ERK to maintain Sox9 expression (suppressing C/EBPβ/δ), while the membrane-bound form additionally inhibits preadipocyte and hematopoietic stem cell proliferation; DLK1 acts as a non-canonical inhibitory ligand of NOTCH1 through direct interaction between its EGF domains 5–6 and NOTCH1 EGF domains 10–15; in hypoxic contexts, ADAM17/HIF-driven cleavage releases an intracellular DLK1 fragment that translocates to the nucleus to promote stem-like and invasive phenotypes; loss-of-function DLK1 mutations cause central precocious puberty and metabolic dysregulation in humans, and DLK1 is required for normal skeletal muscle development, hippocampal neurogenesis, and B-cell development in mice.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"DLK1 (Pref-1) is a paternally expressed, imprinted transmembrane protein containing six EGF-like repeats that functions as a gatekeeper of progenitor cell identity, inhibiting terminal differentiation of adipocytes, osteoblasts, myoblasts, and hematopoietic cells while regulating proliferation in a context- and isoform-dependent manner [PMID:8500166, PMID:15068508, PMID:22801971, PMID:22891218]. Proteolytic cleavage releases a soluble ectodomain that inhibits adipogenesis by activating MEK/ERK signaling to sustain Sox9 expression, thereby suppressing C/EBPβ/δ [PMID:9001251, PMID:19541743, PMID:30355480], whereas the membrane-bound form additionally inhibits preadipocyte and hematopoietic stem cell proliferation through cell-autonomous mechanisms [PMID:22891218, PMID:22801971]; DLK1 directly interacts with NOTCH1 via its EGF domains 5–6 to modulate Notch signaling outputs in a cell-type-specific manner [PMID:26791579, PMID:17320900]. Under hypoxia, ADAM17/HIF-dependent cleavage generates a nuclear-translocating intracellular fragment that promotes stem-like and invasive phenotypes in glioma [PMID:32205867]. Paternally inherited loss-of-function mutations in DLK1 cause familial central precocious puberty with undetectable serum DLK1 [PMID:28324015].\",\n  \"teleology\": [\n    {\n      \"year\": 1993,\n      \"claim\": \"Identification of DLK1 as an EGF-repeat transmembrane protein that blocks adipogenesis established the first functional role for this gene as a negative regulator of differentiation.\",\n      \"evidence\": \"cDNA cloning and constitutive overexpression in 3T3-L1 preadipocytes with differentiation assay\",\n      \"pmids\": [\"8500166\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"receptor or binding partner unknown\", \"mechanism of differentiation inhibition uncharacterized\", \"in vivo relevance not tested\"]\n    },\n    {\n      \"year\": 1997,\n      \"claim\": \"Demonstration that DLK1 is proteolytically cleaved to release a biologically active soluble ectodomain, and that alternative splicing governs whether cleavage occurs, resolved how DLK1 can function in both juxtacrine and paracrine modes.\",\n      \"evidence\": \"In vitro cleavage assays, recombinant ectodomain addition and antibody neutralization in 3T3-L1 cells, splice-form analysis\",\n      \"pmids\": [\"9001251\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"identity of the protease performing cleavage unknown\", \"whether membrane-bound vs. soluble forms have distinct downstream signaling not yet tested\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Mapping of a cis-regulatory SAD element required for transcriptional silencing of DLK1 during differentiation revealed how DLK1 expression is shut off to permit adipogenesis.\",\n      \"evidence\": \"5'-deletion reporter constructs, gel shift, UV cross-linking, and mutagenesis in 3T3-L1 cells\",\n      \"pmids\": [\"9822638\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"identity of the ~63 kDa SAD-binding protein unknown\", \"whether this element operates in non-adipose lineages untested\"]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"Demonstration that dexamethasone promotes adipogenesis by transcriptionally repressing DLK1 connected glucocorticoid signaling to the DLK1 regulatory axis.\",\n      \"evidence\": \"Nuclear run-on, mRNA stability assays, and antisense transfection in 3T3-L1 cells\",\n      \"pmids\": [\"10212243\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"glucocorticoid receptor binding at DLK1 promoter not directly shown\", \"relevance to in vivo glucocorticoid-induced obesity not tested\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"DLK1 was shown to maintain bipotential mesenchymal progenitor identity by inhibiting both osteoblast and adipocyte differentiation, broadening its role beyond adipogenesis; separately, DLK1 was established as the causal gene for callipyge muscular hypertrophy.\",\n      \"evidence\": \"Retroviral overexpression in human MSCs with cytochemistry, FACS, and ex vivo calvaria culture; transgenic DLK1 expression in mouse skeletal muscle and IHC in callipyge sheep\",\n      \"pmids\": [\"15068508\", \"15498495\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"molecular mechanism of osteoblast inhibition not dissected\", \"downstream myogenic signaling targets unknown\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Showing that DLK1's extracellular domain inhibits Notch signaling in mesenchymal cells provided the first direct link between DLK1 and the Notch pathway, though the outcome (pro- vs. anti-adipogenic) proved cell-type-dependent.\",\n      \"evidence\": \"Domain-deletion constructs, co-culture, ERK1/2 activation, and Notch1 knockdown in C3H10T1/2 cells\",\n      \"pmids\": [\"17320900\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"direct physical interaction between DLK1 and Notch1 not yet demonstrated\", \"basis for cell-type-specific divergent outcomes unclear\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Consolidation of the MEK/ERK–Sox9–C/EBPβ/δ signaling axis downstream of soluble DLK1 provided a coherent molecular mechanism for adipogenesis inhibition.\",\n      \"evidence\": \"Synthesis of MAPK assays, Sox9 overexpression/knockdown, and C/EBP reporter assays from multiple studies\",\n      \"pmids\": [\"19541743\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"direct receptor for soluble DLK1 upstream of MEK/ERK not identified\", \"pathway validated mainly in 3T3-L1 model\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Conditional knockout revealed that Dlk1 is required for normal skeletal muscle development and satellite cell regulation, and that CTCF-bound insulator elements control imprinted DLK1 expression with loss of imprinting in AML.\",\n      \"evidence\": \"Muscle-specific Dlk1 KO/OE in mice with satellite cell culture; allele-specific expression, methylation, and CTCF ChIP in AML samples\",\n      \"pmids\": [\"21124733\", \"20089961\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"downstream targets of Dlk1 in satellite cells beyond NF-κB not defined\", \"functional consequence of biallelic expression in AML not directly tested\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Isoform-specific dissection revealed that membrane-bound DLK1 uniquely inhibits preadipocyte and hematopoietic stem cell proliferation, whereas soluble DLK1 primarily inhibits differentiation, establishing that cleavage status dictates functional mode.\",\n      \"evidence\": \"Independent isoform manipulation in preadipocytes with cell cycle analysis and DLK1-null mice; Dlk1 KO/OE mice and stromal co-cultures for hematopoietic readouts\",\n      \"pmids\": [\"22891218\", \"22801971\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"mechanism by which membrane-bound DLK1 arrests G1/S progression unknown\", \"whether isoform-specific effects extend to other tissues untested\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Multiple studies mapped DLK1 function in diverse contexts: Pref-1+ cells were placed as the earliest mesenchymal adipose precursors preceding Zfp423+ and PPARγ+ cells; DLK1 overexpression was shown to cause pituitary IGF1 resistance and shift metabolism toward lipid oxidation; and DLK1 was linked to Notch inhibition in salivary gland organogenesis.\",\n      \"evidence\": \"Lineage tracing with Pref-1 promoter and inducible cell ablation in mice; transgenic DLK1 overexpression with metabolic and endocrine phenotyping; organotypic gland cultures with DLK1 and DAPT\",\n      \"pmids\": [\"25088414\", \"25349437\", \"24828459\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"receptor mediating DLK1 metabolic effects in liver/pituitary not identified\", \"how DLK1 interacts with GH axis mechanistically unclear\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Direct physical interaction between DLK1 EGF domains 5–6 and NOTCH1 EGF domains 10–15 was mapped, providing the structural basis for DLK1-mediated Notch inhibition.\",\n      \"evidence\": \"Mammalian two-hybrid domain deletion mapping, Notch1 activation comparison in Dlk1+/+ vs Dlk1−/− E16.5 tissues, siRNA in cell lines\",\n      \"pmids\": [\"26791579\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"no biophysical or structural characterization of the interaction\", \"whether DLK1 competes with canonical Notch ligands at these domains not directly tested\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Human genetics established that paternally inherited DLK1 loss-of-function causes familial central precocious puberty with undetectable serum DLK1, linking DLK1 to neuroendocrine pubertal timing control.\",\n      \"evidence\": \"Linkage analysis, whole-genome sequencing of affected families, serum DLK1 ELISA, DLK1 expression in mouse hypothalamus and kisspeptin neuron cell lines\",\n      \"pmids\": [\"28324015\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"molecular mechanism by which DLK1 regulates GnRH/kisspeptin axis unknown\", \"whether Notch inhibition mediates the pubertal phenotype untested\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Discovery of a hypoxia-driven, ADAM17/HIF-dependent cleavage event that releases a DLK1 intracellular fragment translocating to the nucleus revealed a non-canonical signaling mode promoting stemness and invasion in glioma.\",\n      \"evidence\": \"ADAM17 inhibitors, HIF siRNA, cleavable vs. non-cleavable DLK1 constructs, nuclear immunofluorescence, PDGFB-induced glioma mouse model\",\n      \"pmids\": [\"32205867\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"nuclear targets of the DLK1 intracellular fragment not identified\", \"whether this mechanism operates outside glioma unknown\", \"structural determinants of intracellular fragment release vs. ectodomain shedding unclear\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Biallelic expression of Dlk1 in the hippocampal subgranular zone, escaping canonical imprinting, was shown to be required for normal adult neurogenesis and spatial discrimination, demonstrating tissue-specific regulation of imprinting.\",\n      \"evidence\": \"Allele-specific expression analysis, maternal/paternal/biallelic Dlk1 mutant mice with behavioral and neurogenesis quantification\",\n      \"pmids\": [\"33712542\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"mechanism of imprint escape in hippocampus not defined\", \"downstream neurogenic targets of Dlk1 in subgranular zone unknown\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The identity of the cell-surface receptor that transduces soluble DLK1 signaling to MEK/ERK and AMPK pathways remains unknown, and the nuclear targets of the DLK1 intracellular fragment have not been identified.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\"no receptor for soluble DLK1 identified\", \"nuclear binding partners/targets of DLK1 ICD unknown\", \"structural basis of DLK1–NOTCH1 interaction not resolved at atomic level\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 6, 17, 25]},\n      {\"term_id\": \"GO:0048018\", \"supporting_discovery_ids\": [1, 7, 16, 19]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [0, 1, 9, 10, 26]},\n      {\"term_id\": \"GO:0005576\", \"supporting_discovery_ids\": [1, 7, 11, 16, 19]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [21]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [6, 7, 12, 17, 19, 25]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [5, 8, 15, 20]},\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [14, 16]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"NOTCH1\", \"ADAM17\", \"SOX9\", \"CTCF\"],\n    \"other_free_text\": []\n  }\n}\n```"}