{"gene":"HIF1AN","run_date":"2026-06-10T01:55:22","timeline":{"discoveries":[{"year":2002,"finding":"FIH-1 is an asparaginyl hydroxylase (Fe(II)-dependent, O2-dependent dioxygenase) that hydroxylates an asparagine residue within the C-terminal transactivation domain (C-TAD) of HIF-1α, blocking its association with transcriptional coactivators and suppressing HIF transcriptional activity under normoxia.","method":"In vitro hydroxylase assay, Fe(II)-dependence tests, molecular O2 substrate assays","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct in vitro enzymatic assay establishing catalytic mechanism, independently corroborated by multiple subsequent studies","pmids":["12080085"],"is_preprint":false},{"year":2001,"finding":"FIH-1 physically binds to HIF-1α and inhibits its transactivation function; FIH-1 also binds to VHL, and VHL recruits histone deacetylases to further repress HIF-1α transactivation, providing a dual mechanism for O2-dependent regulation of HIF-1α activity.","method":"Co-immunoprecipitation, mammalian two-hybrid, transactivation reporter assays","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP and functional reporter assays, replicated across subsequent studies","pmids":["11641274"],"is_preprint":false},{"year":2008,"finding":"FIH-1 undergoes uncoupled O2-activation in the absence of its HIF-1α substrate, resulting in auto-hydroxylation of Trp296 (forming an Fe(III)-O-Trp296 chromophore) and oxidation of Met275; the enzyme active site contains high-spin Fe(III) in both outcomes, with distorted octahedral geometry as shown by XAS and EPR. Co(II)-substituted FIH-1 exists in a mixture of 5- and 6-coordinate forms, indicating the resting enzyme can readily undergo uncoupled O2-activation by loss of an H2O ligand.","method":"UV-Vis spectroscopy, EPR, XAS, in vitro uncoupling assay, Co(II) substitution","journal":"Journal of inorganic biochemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — multiple spectroscopic and biochemical methods in a single study establishing mechanistic intermediates","pmids":["18805587"],"is_preprint":false},{"year":2010,"finding":"Crystal structures of human FIH-1 in complex with quinol inhibitors (Clioquinol and 8-Hydroxyquinoline) show that these compounds bind the FIH-1 active site by coordinating the Fe(II) ion, thereby blocking binding of the co-substrate 2-oxoglutarate (2OG) and inhibiting hydroxylase activity.","method":"X-ray crystallography (crystal structures of FIH-1:inhibitor complexes)","journal":"Molecules and cells","confidence":"High","confidence_rationale":"Tier 1 / Moderate — crystal structures with two inhibitors provide direct structural mechanism for inhibition","pmids":["20396966"],"is_preprint":false},{"year":2021,"finding":"Kinetic isotope effect (KIE) studies with deuterated peptide substrate demonstrate that FIH performs canonical hydrogen atom transfer (HAT) on the asparagine substrate (Dkcat = 10 ± 1), that this step is partially rate-limiting on kcat, and that the deuterated substrate causes uncoupled O2-activation allowing spectroscopic observation of the ferryl intermediate. The close energy barrier between αKG decarboxylation and HAT is identified as the mechanistic basis for FIH's role as an O2-sensing enzyme.","method":"Steady-state kinetics with deuterated substrate, KIE measurements, spectroscopic detection of ferryl intermediate","journal":"Biochemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — mechanistic probe with KIE, ferryl intermediate detection, and internal competition analysis in a single rigorous study","pmids":["34714626"],"is_preprint":false},{"year":2007,"finding":"FIH-1 transcription in renal cell carcinoma is repressed by the homeodomain protein CDP/Cut, which binds a defined site on the FIH-1 promoter in a PKCζ-dependent manner (with Ser987 phosphorylation of CDP as the regulatory phosphosite); this repression reduces FIH-1 levels and permits increased HIF-1 activity and downstream target gene (VEGF, GLUT-1) expression.","method":"Chromatin immunoprecipitation, luciferase promoter assay, PKCζ inhibition, site-directed mutagenesis (CDP S987)","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — ChIP, promoter reporter, and mutagenesis in a single study with multiple orthogonal methods","pmids":["17682059"],"is_preprint":false},{"year":2013,"finding":"FIH-1 hydroxylates the ankyrin repeat domain (ARD)-containing protein ASPP2 at asparagine 986 (N986); this hydroxylation promotes binding of ASPP2 to the cell polarity regulator Par-3. FIH-1 depletion or hydroxylase inhibition (DMOG) causes ASPP2 to relocate from cell-cell contacts to the cytosol without affecting ASPP2 stability, p53 interaction, apoptosis, or proliferation.","method":"Mass spectrometry substrate identification, Co-IP, in vitro hydroxylation assay, immunofluorescence localization, DMOG pharmacological inhibition","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — in vitro hydroxylation, MS identification of hydroxylation site, Co-IP, and localization in one study","pmids":["23606740"],"is_preprint":false},{"year":2013,"finding":"Gankyrin binds to and sequesters FIH-1, reducing FIH-1/HIF-1α interaction, thereby increasing HIF-1 transcriptional activity and VEGF production, and promoting vascular tumor formation in transgenic mouse livers.","method":"Co-immunoprecipitation, transgenic mouse model, cell-based HIF activity assays","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus in vivo transgenic model, single lab","pmids":["23376718"],"is_preprint":false},{"year":2014,"finding":"FIH-1 positively regulates keratinocyte migration by binding to LRRK1 (a regulator of EGFR endosomal trafficking), preventing EGFR/LRRK1 complex formation, thereby enhancing EGFR signaling and downstream ERK1/2 activation. FIH-1 null mice show delayed wound healing.","method":"In vitro scratch wound assays, Co-IP (FIH-1/LRRK1 interaction), FIH-1 knockout mice, EGFR/MAPK pathway analysis","journal":"The American journal of pathology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, KO mouse phenotype, pathway analysis; single lab","pmids":["25455687"],"is_preprint":false},{"year":2012,"finding":"FIH-1 negatively regulates glycogen metabolism in corneal epithelial cells through the AKT/GSK-3β/glycogen synthase pathway in a hydroxylase-independent manner (enzyme-dead FIH-1 fails to reduce glycogen), and independently of HIF-1α.","method":"Retroviral transduction of enzyme-dead FIH-1 mutant, antagomir knockdown, glycogen assay, AKT/GSK-3β western blot, HIF-1α reporter (negative result for HIF-1α dependence)","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — enzyme-dead mutant plus pathway rescue experiments; single lab","pmids":["22532441"],"is_preprint":false},{"year":2014,"finding":"FIH-1 silencing in human vascular endothelial cells (HUVECs) leads to increased Notch2 activity, enhanced Hey-1 expression, and selective repression of survivin, resulting in apoptosis and growth arrest. FIH-1 is proposed to repress Notch2 activity, controlling endothelial cell survival.","method":"siRNA knockdown, western blot (Notch2, Hey-1, survivin), apoptosis assays","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — knockdown with defined pathway readout (Notch2→Hey-1→survivin) but single lab, no direct FIH-1/Notch2 binding demonstrated","pmids":["25837583"],"is_preprint":false},{"year":2010,"finding":"FIH-1 directly interacts with the pro-apoptotic protein Bax and functions as a cytosolic retention factor for Bax, preventing its translocation to mitochondria in response to apoptotic stimuli; FIH-1 overexpression suppresses Bax-mediated apoptosis, while FIH-1 deficiency accelerates it.","method":"Co-immunoprecipitation, GST pull-down, apoptosis assays, subcellular fractionation","journal":"Molecular and cellular biochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP, GST pull-down, and functional apoptosis assays; single lab","pmids":["21069436"],"is_preprint":false},{"year":2017,"finding":"KANK3 is a substrate for HIF1AN asparaginyl hydroxylation; in vitro hydroxylation assay confirmed hydroxylation, and mass spectrometry identified three hydroxylated asparagine residues within the ankyrin repeat domain of KANK3. KANK3 tumor-suppressive effects on cell migration/invasion were not observed under hypoxic conditions, consistent with oxygen-dependent HIF1AN activity.","method":"In vitro hydroxylation assay, mass spectrometry, cell migration/invasion assays under normoxia vs. hypoxia","journal":"Cell biology international","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro assay and MS identification of hydroxylation sites; single lab, no mutagenesis validation of individual sites","pmids":["29047187"],"is_preprint":false},{"year":2014,"finding":"FIH-1 inhibits HIF-1-mediated transcription of GLUT1 and VEGF-A even under hypoxic conditions in glioblastoma cells, and FIH-1 is more potent than PTEN in inhibiting HIF function in this context.","method":"Retroviral FIH-1 expression, siRNA knockdown, quantitative RT-PCR for HIF target genes (GLUT1, VEGF-A) under hypoxia","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — gain- and loss-of-function with defined transcriptional readouts; single lab","pmids":["24465898"],"is_preprint":false},{"year":2011,"finding":"In VHL-defective CCRCC cells expressing both HIF-1α and HIF-2α, FIH-1 knockdown increases HIF target gene expression and apoptosis. In 786-O cells expressing only HIF-2α, FIH-1 knockdown has no effect; re-introduction of HIF-1α restores sensitivity to FIH-1 knockdown. This establishes FIH-1's substrate preference for HIF-1α over HIF-2α in modulating both target gene expression and cell viability.","method":"siRNA knockdown, retroviral gene expression, qRT-PCR, Annexin V/propidium iodide apoptosis assay","journal":"British journal of cancer","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic complementation (HIF-1α rescue) plus matched isogenic cell lines; single lab","pmids":["21386837"],"is_preprint":false},{"year":2019,"finding":"FIH-1 interacts with Plectin1 and STAT1 (novel binding partners identified by BioID proteomics), as well as known partners ASPP2 and HDAC1, to positively regulate ΔNp63α expression in keratinocytes. ASPP2 knockdown upregulates ΔNp63α; FIH-1 regulates GADD45α (a negative regulator of ΔNp63α) via HDAC1 interaction.","method":"BioID proteomics, Co-IP, siRNA knockdown, FIH-1 conditional transgenic and knockout mouse","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — BioID proximity labeling, Co-IP, and in vivo mouse model; single lab","pmids":["31914679"],"is_preprint":false},{"year":2018,"finding":"ANKDD1A directly interacts with FIH1 and upregulates FIH1 activity, leading to decreased HIF1α transcriptional activity and reduced HIF1α protein half-life. This decreases glucose uptake, lactate production, and autophagy while inducing apoptosis in glioblastoma cells under hypoxia.","method":"Co-immunoprecipitation (ANKDD1A-FIH1 interaction), HIF1α stability assay, glucose/lactate measurement, autophagy and apoptosis assays","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — Co-IP plus functional readouts; single lab","pmids":["30082910"],"is_preprint":false},{"year":2021,"finding":"The N-terminal disordered region of Mint3 (amino acids 78–88 as core binding site) binds FIH-1 with a large enthalpy and entropy change consistent with disorder-to-order transition upon binding; thermodynamic mapping by ITC with truncated Mint3 constructs identified the core binding site and distinguished affinity-contributing from enthalpy-only-contributing regions.","method":"Circular dichroism, NMR, hydrogen/deuterium exchange-MS, isothermal titration calorimetry (ITC) with truncation series","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — multiple orthogonal biophysical methods (CD, NMR, HDX-MS, ITC) with systematic truncation analysis in one rigorous study","pmids":["34655613"],"is_preprint":false},{"year":2022,"finding":"FIH-1 knockout in intestinal epithelial cells (FIH-1 silencing in HIEC-6 cells) activates the PI3K-AKT signaling pathway and cell cycle redistribution into S-phase, conferring radioprotection by mitigating radiation-induced cell damage.","method":"shRNA knockdown, colony formation assay, cell cycle analysis, apoptosis assay, RNA-Seq, western blot","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — RNA-Seq plus western blot pathway analysis with functional radiosensitivity assays; single lab","pmids":["40762438"],"is_preprint":false},{"year":2023,"finding":"ASB9 (E3 ubiquitin ligase) directly interacts with HIF1AN and promotes its ubiquitination and degradation; ASB9 overexpression inhibits human SSC line proliferation and increases apoptosis, effects reversed by HIF1AN re-expression.","method":"Co-immunoprecipitation, western blot (HIF1AN protein level), functional proliferation/apoptosis assays with rescue","journal":"Biological research","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — Co-IP demonstrating direct interaction, functional rescue; single lab","pmids":["36683111"],"is_preprint":false},{"year":2022,"finding":"FIH-1 knockout in colon epithelial cells (Vil1-Cre/FIH+f/+f mice) attenuates chronic colitis severity and reduces infiltrating macrophages and immune-response gene expression, but does not alter colorectal tumor occurrence in an AOM/DSS model.","method":"Conditional knockout mouse (Vil1-Cre), AOM/DSS colitis-cancer model, RNA-seq of colon tissue, histology, immunohistochemistry","journal":"Journal of immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo conditional KO with transcriptomics and histology; single lab","pmids":["35121641"],"is_preprint":false},{"year":2025,"finding":"FIH-1 does not interact with IKKα/β or IκBα (negative result by immunoprecipitation), and FIH-1 overexpression does not affect IKKα/β or p65 protein levels, indicating FIH-1 does not regulate the classical NF-κB pathway.","method":"Immunoprecipitation, western blot, FIH-1 overexpression","journal":"The Journal of toxicological sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — negative result from Co-IP and overexpression; single lab, single method per assay","pmids":["40024754"],"is_preprint":false},{"year":2014,"finding":"FIH-1 (HIF1AN) physically interacts with Mindbomb (Mib) E3 ubiquitin ligase in zebrafish and cultured cells (co-localization confirmed); depletion of fih-1 in zebrafish embryos induces ectopic VEGF-A expression and ectopic intersegmental vessel sprouting, while fih-1 overexpression attenuates ISV formation in a VEGF-A-rescuable manner, establishing FIH-1 as an anti-angiogenic regulator of VEGF-A signaling.","method":"Yeast two-hybrid (Mib interaction), co-localization in cultured cells, zebrafish morpholino knockdown, fih-1 mRNA overexpression, ISV imaging","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — yeast two-hybrid plus in vivo zebrafish gain- and loss-of-function with rescue; single lab","pmids":["25347788"],"is_preprint":false},{"year":2025,"finding":"PML (promyelocytic leukemia protein) directly binds HIF1AN and promotes its ubiquitination-mediated degradation; PML knockdown or HIF1AN upregulation suppresses osteogenic differentiation of BMSCs; HIF1α directly binds the SOD3 promoter; PML overexpression promotes osteogenesis via HIF1AN/HIF1α/SOD3 axis and PI3K/AKT pathway.","method":"Co-immunoprecipitation, immunofluorescence, ChIP, dual-luciferase reporter, western blot, ALP/Alizarin red staining","journal":"International journal of stem cells","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and ChIP with functional differentiation readouts; single lab","pmids":["40059075"],"is_preprint":false}],"current_model":"HIF1AN (FIH-1) is an Fe(II)- and O2-dependent asparaginyl hydroxylase that hydroxylates Asn803 of HIF-1α's C-terminal transactivation domain, blocking CBP/p300 co-activator recruitment and suppressing HIF-1 transcriptional activity under normoxia; it also hydroxylates ankyrin repeat domain (ARD)-containing substrates (including ASPP2, KANK3, and Notch) to modulate their protein–protein interactions, and performs HIF-1α-independent functions including regulation of AKT/GSK-3β-dependent glycogen metabolism, cytosolic retention of Bax to suppress apoptosis, and EGFR/ERK-mediated keratinocyte migration via LRRK1 binding, with its own expression controlled transcriptionally by the CDP/Cut–PKCζ axis and its protein stability regulated by ASB9 and PML-mediated ubiquitination."},"narrative":{"mechanistic_narrative":"HIF1AN (FIH-1) is an Fe(II)- and O2-dependent asparaginyl hydroxylase that functions as the principal oxygen sensor restraining HIF transcriptional activity under normoxia by hydroxylating an asparagine in the C-terminal transactivation domain of HIF-1α, blocking coactivator recruitment and binding both HIF-1α and VHL to achieve dual O2-dependent repression [PMID:12080085, PMID:11641274]. Its catalytic cycle proceeds through canonical hydrogen-atom transfer on the asparagine via a ferryl intermediate, and the near-equal energy barrier between αKG decarboxylation and HAT is the mechanistic basis for its O2-sensing role; in the absence of substrate the enzyme undergoes uncoupled O2-activation with auto-hydroxylation of Trp296, and quinol inhibitors block activity by coordinating the active-site Fe and excluding 2-oxoglutarate [PMID:18805587, PMID:20396966, PMID:34714626]. FIH-1 shows substrate preference for HIF-1α over HIF-2α and suppresses HIF target genes such as GLUT1 and VEGF-A even under hypoxia [PMID:24465898, PMID:21386837]. Beyond HIF, FIH-1 hydroxylates ankyrin-repeat-domain substrates including ASPP2 at N986—promoting ASPP2/Par-3 binding and junctional localization—and KANK3, coupling its enzymatic activity to protein-protein interactions [PMID:23606740, PMID:29047187]. It also performs HIF-1α-independent functions: hydroxylase-independent suppression of glycogen metabolism via the AKT/GSK-3β axis, cytosolic retention of pro-apoptotic Bax to suppress apoptosis, and regulation of keratinocyte migration and ΔNp63α expression through binding partners including LRRK1, Plectin1, STAT1, ASPP2 and HDAC1 [PMID:22532441, PMID:21069436, PMID:25455687, PMID:31914679]. FIH-1 abundance is set transcriptionally by CDP/Cut–PKCζ repression and post-translationally by ASB9- and PML-mediated ubiquitination and degradation, and its activity is modulated by interacting proteins that either sequester it (Gankyrin) or stimulate it (ANKDD1A) to tune HIF output [PMID:17682059, PMID:36683111, PMID:40059075, PMID:23376718, PMID:30082910].","teleology":[{"year":2001,"claim":"Established that FIH-1 is a physical inhibitor of HIF-1α transactivation, defining a new oxygen-regulatory node distinct from the VHL/proteolysis axis.","evidence":"Co-IP, mammalian two-hybrid, and transactivation reporter assays showing FIH-1 binds HIF-1α and VHL","pmids":["11641274"],"confidence":"High","gaps":["Did not define the enzymatic basis of inhibition","Mechanism of VHL-mediated HDAC recruitment not resolved"]},{"year":2002,"claim":"Identified the molecular mechanism: FIH-1 is an Fe(II)/O2-dependent asparaginyl hydroxylase that modifies the HIF-1α C-TAD asparagine to block coactivator binding, explaining O2-dependent HIF suppression.","evidence":"In vitro hydroxylase assays with Fe(II)- and molecular O2-dependence tests","pmids":["12080085"],"confidence":"High","gaps":["Catalytic intermediates and rate-limiting steps not characterized","Non-HIF substrates not yet known"]},{"year":2007,"claim":"Showed FIH-1 expression itself is controlled, with CDP/Cut repressing the FIH-1 promoter in a PKCζ-dependent manner to derepress HIF activity in renal carcinoma.","evidence":"ChIP, luciferase promoter assays, PKCζ inhibition, and CDP S987 mutagenesis","pmids":["17682059"],"confidence":"High","gaps":["Generality of CDP/Cut control beyond RCC unknown","Upstream signals activating PKCζ not defined"]},{"year":2008,"claim":"Defined the active-site chemistry of substrate-free FIH-1, revealing uncoupled O2-activation, Trp296 auto-hydroxylation, and high-spin Fe(III) geometry.","evidence":"UV-Vis, EPR, XAS, Co(II) substitution, and in vitro uncoupling assays","pmids":["18805587"],"confidence":"High","gaps":["Physiological consequence of auto-hydroxylation unclear","Did not capture the productive ferryl intermediate"]},{"year":2010,"claim":"Provided structural mechanism of inhibition, showing quinol compounds coordinate the active-site Fe and exclude 2OG, and revealed FIH-1 acts as a cytosolic Bax retention factor independent of its HIF role.","evidence":"X-ray crystallography of FIH-1:inhibitor complexes; Co-IP, GST pull-down, fractionation, and apoptosis assays for Bax","pmids":["20396966","21069436"],"confidence":"High","gaps":["Whether Bax retention requires hydroxylase activity not established","Structural basis of Bax interaction unknown"]},{"year":2011,"claim":"Demonstrated FIH-1 substrate preference for HIF-1α over HIF-2α in modulating target genes and cell viability, using isogenic CCRCC lines and HIF-1α complementation.","evidence":"siRNA knockdown, retroviral HIF-1α re-introduction, qRT-PCR, and apoptosis assays","pmids":["21386837"],"confidence":"Medium","gaps":["Structural basis of isoform discrimination not addressed","Single lab"]},{"year":2012,"claim":"Revealed a hydroxylase-independent, HIF-1α-independent function in glycogen metabolism via the AKT/GSK-3β pathway, broadening FIH-1 biology beyond catalysis.","evidence":"Enzyme-dead FIH-1 mutant, antagomir knockdown, glycogen assays, and pathway western blots","pmids":["22532441"],"confidence":"Medium","gaps":["Molecular target linking FIH-1 to AKT not identified","Generality beyond corneal epithelium unknown"]},{"year":2013,"claim":"Extended FIH-1 substrate range to ARD proteins, showing ASPP2 N986 hydroxylation controls ASPP2/Par-3 binding and junctional localization, and identifying Gankyrin as a sequestering regulator that boosts HIF activity in vivo.","evidence":"MS site identification, in vitro hydroxylation, Co-IP, immunofluorescence (ASPP2); Co-IP and transgenic mouse (Gankyrin)","pmids":["23606740","23376718"],"confidence":"Medium","gaps":["Functional consequence of ASPP2 mislocalization on tissue physiology unclear","Whether Gankyrin affects FIH-1 catalysis directly not shown"]},{"year":2014,"claim":"Expanded FIH-1 into angiogenesis, endothelial survival, keratinocyte migration and continued HIF suppression under hypoxia, linking it to Notch2, LRRK1/EGFR, and Mindbomb.","evidence":"siRNA/Notch readouts (HUVEC); scratch assays, Co-IP, KO mice (LRRK1); zebrafish morpholino, Y2H, ISV imaging (Mib); retroviral/siRNA qRT-PCR in glioblastoma","pmids":["25837583","25455687","25347788","24465898"],"confidence":"Medium","gaps":["Direct FIH-1/Notch2 binding not demonstrated","Whether LRRK1 and Mib effects depend on hydroxylation unresolved"]},{"year":2017,"claim":"Confirmed KANK3 as an ARD hydroxylation substrate with three hydroxylated asparagines, reinforcing FIH-1's role as a regulator of ARD-containing tumor suppressors under oxygen control.","evidence":"In vitro hydroxylation, mass spectrometry, and normoxia-vs-hypoxia migration/invasion assays","pmids":["29047187"],"confidence":"Medium","gaps":["Individual sites not validated by mutagenesis","Functional impact of each hydroxylation site undefined"]},{"year":2018,"claim":"Identified ANKDD1A as a direct activator that upregulates FIH-1 to lower HIF1α stability and activity, suppressing glycolysis and autophagy in glioblastoma.","evidence":"Co-IP, HIF1α stability assays, glucose/lactate measurement, autophagy/apoptosis assays","pmids":["30082910"],"confidence":"Medium","gaps":["Mechanism by which ANKDD1A enhances catalysis unknown","Single lab"]},{"year":2019,"claim":"Mapped an interactome (Plectin1, STAT1, ASPP2, HDAC1) through which FIH-1 positively regulates ΔNp63α in keratinocytes, integrating it into epidermal gene regulation.","evidence":"BioID proteomics, Co-IP, siRNA, and conditional transgenic/knockout mice","pmids":["31914679"],"confidence":"Medium","gaps":["Whether these interactions require hydroxylase activity unclear","Direct hydroxylation of new partners not tested"]},{"year":2021,"claim":"Resolved the catalytic basis of O2 sensing—partially rate-limiting HAT through a ferryl intermediate with a close barrier to αKG decarboxylation—and thermodynamically characterized disordered-substrate (Mint3) binding via disorder-to-order transition.","evidence":"KIE with deuterated substrate and ferryl detection (kinetics); CD, NMR, HDX-MS, ITC with truncations (Mint3)","pmids":["34714626","34655613"],"confidence":"High","gaps":["In-cell relevance of the kinetic barrier not tested","Structure of the FIH-1/Mint3 complex not determined"]},{"year":2023,"claim":"Established post-translational control of FIH-1 abundance, with ASB9 directly binding and targeting HIF1AN for ubiquitin-mediated degradation in spermatogonial stem cells.","evidence":"Co-IP, HIF1AN protein-level western blots, and proliferation/apoptosis rescue assays","pmids":["36683111"],"confidence":"Medium","gaps":["Ubiquitination sites not mapped","Generality beyond SSC lines unknown"]},{"year":2025,"claim":"Refined the regulatory and signaling boundaries of FIH-1: PML drives its ubiquitin-mediated degradation to promote osteogenesis, while FIH-1 was shown not to regulate the classical NF-κB pathway.","evidence":"Co-IP, ChIP, luciferase, differentiation assays (PML); IP and overexpression negative results (NF-κB)","pmids":["40059075","40024754"],"confidence":"Medium","gaps":["PML and ASB9 relationship to each other unresolved","Negative NF-κB result is single-lab"]},{"year":null,"claim":"How FIH-1 selects among its many catalytic (HIF-1α, ASPP2, KANK3) and non-catalytic (Bax, LRRK1, AKT) functions in different tissues, and which depend on hydroxylase activity versus scaffolding, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified rule distinguishing hydroxylation-dependent from independent functions","Tissue-specific substrate hierarchy undefined","Structural models for most non-HIF substrates lacking"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016491","term_label":"oxidoreductase activity","supporting_discovery_ids":[0,2,4,6,12]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,6,12]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[1,11,13]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[6,11]}],"pathway":[{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[0,1,13,14]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[1,13,15]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[11,10]}],"complexes":[],"partners":["HIF1A","VHL","ASPP2","KANK3","LRRK1","BAX","HDAC1","ANKDD1A"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9NWT6","full_name":"Hypoxia-inducible factor 1-alpha inhibitor","aliases":["Factor inhibiting HIF-1","FIH-1","Hypoxia-inducible factor asparagine hydroxylase"],"length_aa":349,"mass_kda":40.3,"function":"Hydroxylates HIF-1 alpha at 'Asn-803' in the C-terminal transactivation domain (CAD). Functions as an oxygen sensor and, under normoxic conditions, the hydroxylation prevents interaction of HIF-1 with transcriptional coactivators including Cbp/p300-interacting transactivator. Involved in transcriptional repression through interaction with HIF1A, VHL and histone deacetylases. Hydroxylates specific Asn residues within ankyrin repeat domains (ARD) of NFKB1, NFKBIA, NOTCH1, ASB4, PPP1R12A and several other ARD-containing proteins. Also hydroxylates Asp and His residues within ARDs of ANK1 and TNKS2, respectively. Negatively regulates NOTCH1 activity, accelerating myogenic differentiation. Positively regulates ASB4 activity, promoting vascular differentiation","subcellular_location":"Nucleus; Cytoplasm; Cytoplasm, perinuclear region","url":"https://www.uniprot.org/uniprotkb/Q9NWT6/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/HIF1AN","classification":"Not Classified","n_dependent_lines":75,"n_total_lines":1208,"dependency_fraction":0.062086092715231786},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/HIF1AN","total_profiled":1310},"omim":[{"mim_id":"617310","title":"ANKYRIN REPEAT AND STERILE ALPHA MOTIF DOMAINS-CONTAINING PROTEIN 3; ANKS3","url":"https://www.omim.org/entry/617310"},{"mim_id":"615370","title":"ANKYRIN REPEAT AND STERILE ALPHA MOTIF DOMAINS-CONTAINING PROTEIN 6; ANKS6","url":"https://www.omim.org/entry/615370"},{"mim_id":"609799","title":"NIMA-RELATED KINASE 8; NEK8","url":"https://www.omim.org/entry/609799"},{"mim_id":"608537","title":"VON HIPPEL-LINDAU TUMOR SUPPRESSOR; VHL","url":"https://www.omim.org/entry/608537"},{"mim_id":"608002","title":"NEPHROCYSTIN 3; NPHP3","url":"https://www.omim.org/entry/608002"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"skeletal muscle","ntpm":38.9},{"tissue":"tongue","ntpm":38.1}],"url":"https://www.proteinatlas.org/search/HIF1AN"},"hgnc":{"alias_symbol":["FLJ20615","DKFZp762F1811","FLJ22027","FIH1"],"prev_symbol":[]},"alphafold":{"accession":"Q9NWT6","domains":[{"cath_id":"2.60.120.10","chopping":"28-297","consensus_level":"high","plddt":96.8458,"start":28,"end":297},{"cath_id":"1.10.287.1010","chopping":"311-343","consensus_level":"high","plddt":93.8097,"start":311,"end":343}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NWT6","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NWT6-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NWT6-F1-predicted_aligned_error_v6.png","plddt_mean":91.38},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=HIF1AN","jax_strain_url":"https://www.jax.org/strain/search?query=HIF1AN"},"sequence":{"accession":"Q9NWT6","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9NWT6.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9NWT6/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NWT6"}},"corpus_meta":[{"pmid":"12080085","id":"PMC_12080085","title":"FIH-1 is an asparaginyl hydroxylase enzyme that regulates the transcriptional activity of hypoxia-inducible factor.","date":"2002","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/12080085","citation_count":1258,"is_preprint":false},{"pmid":"11641274","id":"PMC_11641274","title":"FIH-1: a novel protein that interacts with HIF-1alpha and VHL to mediate repression of HIF-1 transcriptional activity.","date":"2001","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/11641274","citation_count":1174,"is_preprint":false},{"pmid":"26303948","id":"PMC_26303948","title":"MicroRNA-455 regulates brown adipogenesis via a novel HIF1an-AMPK-PGC1α signaling network.","date":"2015","source":"EMBO reports","url":"https://pubmed.ncbi.nlm.nih.gov/26303948","citation_count":133,"is_preprint":false},{"pmid":"31410089","id":"PMC_31410089","title":"MiR-135-5p promotes osteoblast differentiation by targeting HIF1AN in MC3T3-E1 cells.","date":"2019","source":"Cellular & molecular biology letters","url":"https://pubmed.ncbi.nlm.nih.gov/31410089","citation_count":98,"is_preprint":false},{"pmid":"26205124","id":"PMC_26205124","title":"Hypoxia-inducible miR-182 enhances HIF1α signaling via targeting PHD2 and FIH1 in prostate cancer.","date":"2015","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/26205124","citation_count":87,"is_preprint":false},{"pmid":"34474073","id":"PMC_34474073","title":"Exosomes from adipose-derived stem cells alleviate myocardial infarction via microRNA-31/FIH1/HIF-1α pathway.","date":"2021","source":"Journal of molecular and cellular cardiology","url":"https://pubmed.ncbi.nlm.nih.gov/34474073","citation_count":68,"is_preprint":false},{"pmid":"24384398","id":"PMC_24384398","title":"Transcriptional regulation of hypoxia inducible factors alpha (HIF-α) and their inhibiting factor (FIH-1) of channel catfish (Ictalurus punctatus) under hypoxia.","date":"2013","source":"Comparative biochemistry and physiology. Part B, Biochemistry & molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/24384398","citation_count":67,"is_preprint":false},{"pmid":"15220362","id":"PMC_15220362","title":"Molecular genetic analysis of FIH-1, FH, and SDHB candidate tumour suppressor genes in renal cell carcinoma.","date":"2004","source":"Journal of clinical pathology","url":"https://pubmed.ncbi.nlm.nih.gov/15220362","citation_count":57,"is_preprint":false},{"pmid":"24521875","id":"PMC_24521875","title":"MicroRNA-31 contributes to colorectal cancer development by targeting factor inhibiting HIF-1α (FIH-1).","date":"2014","source":"Cancer biology & therapy","url":"https://pubmed.ncbi.nlm.nih.gov/24521875","citation_count":54,"is_preprint":false},{"pmid":"22532441","id":"PMC_22532441","title":"MicroRNA-31 targets FIH-1 to positively regulate corneal epithelial glycogen metabolism.","date":"2012","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/22532441","citation_count":53,"is_preprint":false},{"pmid":"38030054","id":"PMC_38030054","title":"M2 macrophage-derived exosomes induce angiogenesis and increase skin flap survival through HIF1AN/HIF-1α/VEGFA control.","date":"2023","source":"Archives of biochemistry and biophysics","url":"https://pubmed.ncbi.nlm.nih.gov/38030054","citation_count":51,"is_preprint":false},{"pmid":"25907805","id":"PMC_25907805","title":"MicroRNA-135b regulates ERα, AR and HIF1AN and affects breast and prostate cancer cell growth.","date":"2015","source":"Molecular oncology","url":"https://pubmed.ncbi.nlm.nih.gov/25907805","citation_count":50,"is_preprint":false},{"pmid":"25728779","id":"PMC_25728779","title":"The role of the miR-31/FIH1 pathway in TGF-β-induced liver fibrosis.","date":"2015","source":"Clinical science (London, England : 1979)","url":"https://pubmed.ncbi.nlm.nih.gov/25728779","citation_count":49,"is_preprint":false},{"pmid":"23606740","id":"PMC_23606740","title":"Factor inhibiting HIF-1 (FIH-1) modulates protein interactions of apoptosis-stimulating p53 binding protein 2 (ASPP2).","date":"2013","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/23606740","citation_count":45,"is_preprint":false},{"pmid":"35592530","id":"PMC_35592530","title":"Regulation of Transactivation at C-TAD Domain of HIF-1α by Factor-Inhibiting HIF-1α (FIH-1): A Potential Target for Therapeutic Intervention in Cancer.","date":"2022","source":"Oxidative medicine and cellular longevity","url":"https://pubmed.ncbi.nlm.nih.gov/35592530","citation_count":43,"is_preprint":false},{"pmid":"24465898","id":"PMC_24465898","title":"The role of factor inhibiting HIF (FIH-1) in inhibiting HIF-1 transcriptional activity in glioblastoma multiforme.","date":"2014","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/24465898","citation_count":43,"is_preprint":false},{"pmid":"30082910","id":"PMC_30082910","title":"Hypermethylated gene ANKDD1A is a candidate tumor suppressor that interacts with FIH1 and decreases HIF1α stability to inhibit cell autophagy in the glioblastoma multiforme hypoxia microenvironment.","date":"2018","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/30082910","citation_count":40,"is_preprint":false},{"pmid":"27074047","id":"PMC_27074047","title":"MicroRNA-125a-5p Contributes to Hepatic Stellate Cell Activation through Targeting FIH1.","date":"2016","source":"Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/27074047","citation_count":35,"is_preprint":false},{"pmid":"21386837","id":"PMC_21386837","title":"Factor inhibiting HIF (FIH-1) promotes renal cancer cell survival by protecting cells from HIF-1α-mediated apoptosis.","date":"2011","source":"British journal of cancer","url":"https://pubmed.ncbi.nlm.nih.gov/21386837","citation_count":35,"is_preprint":false},{"pmid":"25602156","id":"PMC_25602156","title":"Factor inhibiting HIF1α (FIH-1) functions as a tumor suppressor in human colorectal cancer by repressing HIF1α pathway.","date":"2015","source":"Cancer biology & therapy","url":"https://pubmed.ncbi.nlm.nih.gov/25602156","citation_count":34,"is_preprint":false},{"pmid":"25837583","id":"PMC_25837583","title":"Factor-inhibiting HIF-1 (FIH-1) is required for human vascular endothelial cell survival.","date":"2015","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/25837583","citation_count":27,"is_preprint":false},{"pmid":"25347788","id":"PMC_25347788","title":"FIH-1, a novel interactor of mindbomb, functions as an essential anti-angiogenic factor during zebrafish vascular development.","date":"2014","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/25347788","citation_count":26,"is_preprint":false},{"pmid":"29088812","id":"PMC_29088812","title":"Downregulation of microRNA-31 inhibits proliferation and induces apoptosis by targeting HIF1AN in human keloid.","date":"2017","source":"Oncotarget","url":"https://pubmed.ncbi.nlm.nih.gov/29088812","citation_count":25,"is_preprint":false},{"pmid":"25277131","id":"PMC_25277131","title":"Upregulation of miR-184 enhances the malignant biological behavior of human glioma cell line A172 by targeting FIH-1.","date":"2014","source":"Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/25277131","citation_count":24,"is_preprint":false},{"pmid":"18805587","id":"PMC_18805587","title":"Coordination changes and auto-hydroxylation of FIH-1: uncoupled O2-activation in a human hypoxia sensor.","date":"2008","source":"Journal of inorganic biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/18805587","citation_count":24,"is_preprint":false},{"pmid":"25431923","id":"PMC_25431923","title":"Variants of the low oxygen sensors EGLN1 and HIF-1AN associated with acute mountain sickness.","date":"2014","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/25431923","citation_count":23,"is_preprint":false},{"pmid":"34046994","id":"PMC_34046994","title":"TMEM161B-AS1 suppresses proliferation, invasion and glycolysis by targeting miR-23a-3p/HIF1AN signal axis in oesophageal squamous cell carcinoma.","date":"2021","source":"Journal of cellular and molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/34046994","citation_count":22,"is_preprint":false},{"pmid":"16705796","id":"PMC_16705796","title":"Down-regulation of the expression of the FIH-1 and ARD-1 genes at the transcriptional level by nickel and cobalt in the human lung adenocarcinoma A549 cell line.","date":"2005","source":"International journal of environmental research and public health","url":"https://pubmed.ncbi.nlm.nih.gov/16705796","citation_count":22,"is_preprint":false},{"pmid":"23376718","id":"PMC_23376718","title":"Overexpression of gankyrin in mouse hepatocytes induces hemangioma by suppressing factor inhibiting hypoxia-inducible factor-1 (FIH-1) and activating hypoxia-inducible factor-1.","date":"2013","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/23376718","citation_count":22,"is_preprint":false},{"pmid":"36683111","id":"PMC_36683111","title":"Ubiquitin protein E3 ligase ASB9 suppresses proliferation and promotes apoptosis in human spermatogonial stem cell line by inducing HIF1AN degradation.","date":"2023","source":"Biological research","url":"https://pubmed.ncbi.nlm.nih.gov/36683111","citation_count":20,"is_preprint":false},{"pmid":"21732131","id":"PMC_21732131","title":"Subcellular FIH-1 expression patterns in invasive breast cancer in relation to HIF-1α expression.","date":"2011","source":"Cellular oncology (Dordrecht, Netherlands)","url":"https://pubmed.ncbi.nlm.nih.gov/21732131","citation_count":19,"is_preprint":false},{"pmid":"30536131","id":"PMC_30536131","title":"The miRNA-184 drives renal fibrosis by targeting HIF1AN in vitro and in vivo.","date":"2018","source":"International urology and nephrology","url":"https://pubmed.ncbi.nlm.nih.gov/30536131","citation_count":18,"is_preprint":false},{"pmid":"36074324","id":"PMC_36074324","title":"METTL3 m6A-dependently promotes miR-21-5p maturation to accelerate choriocarcinoma progression via the HIF1AN-induced inactivation of the HIF1A/VEGF pathway.","date":"2022","source":"Genes & genomics","url":"https://pubmed.ncbi.nlm.nih.gov/36074324","citation_count":18,"is_preprint":false},{"pmid":"17682059","id":"PMC_17682059","title":"Protein kinase C-mediated modulation of FIH-1 expression by the homeodomain protein CDP/Cut/Cux.","date":"2007","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/17682059","citation_count":15,"is_preprint":false},{"pmid":"25903459","id":"PMC_25903459","title":"microRNA-98 mediated microvascular hyperpermeability during burn shock phase via inhibiting FIH-1.","date":"2015","source":"European journal of medical research","url":"https://pubmed.ncbi.nlm.nih.gov/25903459","citation_count":15,"is_preprint":false},{"pmid":"20396966","id":"PMC_20396966","title":"Crystal structures of human FIH-1 in complex with quinol family inhibitors.","date":"2010","source":"Molecules and cells","url":"https://pubmed.ncbi.nlm.nih.gov/20396966","citation_count":14,"is_preprint":false},{"pmid":"33791813","id":"PMC_33791813","title":"Silencing of SNHG6 alleviates hypoxia/reoxygenation-induced cardiomyocyte apoptosis by modulating miR-135a-5p/HIF1AN to activate Shh/Gli1 signalling pathway.","date":"2021","source":"The Journal of pharmacy and pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/33791813","citation_count":13,"is_preprint":false},{"pmid":"29047187","id":"PMC_29047187","title":"A novel HIF1AN substrate KANK3 plays a tumor-suppressive role in hepatocellular carcinoma.","date":"2017","source":"Cell biology international","url":"https://pubmed.ncbi.nlm.nih.gov/29047187","citation_count":13,"is_preprint":false},{"pmid":"33619902","id":"PMC_33619902","title":"CircCDR1as Suppresses Bone Microvascular Endothelial Cell Activity and Angiogenesis Through Targeting miR-135b/ FIH-1 Axis.","date":"2021","source":"Orthopaedic surgery","url":"https://pubmed.ncbi.nlm.nih.gov/33619902","citation_count":13,"is_preprint":false},{"pmid":"32387587","id":"PMC_32387587","title":"MicroRNA-1-3p enhances osteoblast differentiation of MC3T3-E1 cells by interacting with hypoxia-inducible factor 1 α inhibitor (HIF1AN).","date":"2020","source":"Mechanisms of development","url":"https://pubmed.ncbi.nlm.nih.gov/32387587","citation_count":12,"is_preprint":false},{"pmid":"33336500","id":"PMC_33336500","title":"CASC2 inhibits the growth, migration, and invasion of thyroid cancer cells through sponging miR-18a-5p/FIH1 axis.","date":"2020","source":"The Kaohsiung journal of medical sciences","url":"https://pubmed.ncbi.nlm.nih.gov/33336500","citation_count":12,"is_preprint":false},{"pmid":"25455687","id":"PMC_25455687","title":"FIH-1 disrupts an LRRK1/EGFR complex to positively regulate keratinocyte migration.","date":"2014","source":"The American journal of pathology","url":"https://pubmed.ncbi.nlm.nih.gov/25455687","citation_count":11,"is_preprint":false},{"pmid":"36334804","id":"PMC_36334804","title":"Enhanced BPGM/2,3-DPG pathway activity suppresses glycolysis in hypoxic astrocytes via FIH-1 and TET2.","date":"2022","source":"Brain research bulletin","url":"https://pubmed.ncbi.nlm.nih.gov/36334804","citation_count":10,"is_preprint":false},{"pmid":"30607683","id":"PMC_30607683","title":"Molecular characterization and expression regulation of the factor-inhibiting HIF-1 (FIH-1) gene under hypoxic stress in bighead carp (Aristichthys nobilis).","date":"2019","source":"Fish physiology and biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/30607683","citation_count":10,"is_preprint":false},{"pmid":"31914679","id":"PMC_31914679","title":"FIH-1 engages novel binding partners to positively influence epithelial proliferation via p63.","date":"2019","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/31914679","citation_count":10,"is_preprint":false},{"pmid":"31157259","id":"PMC_31157259","title":"Polymorphisms in microRNA let-7 binding sites of the HIF1AN and CLDN12 genes can predict pathologic complete response to taxane- and platinum-based neoadjuvant chemotherapy in breast cancer.","date":"2019","source":"Annals of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/31157259","citation_count":9,"is_preprint":false},{"pmid":"26749281","id":"PMC_26749281","title":"Investigation of FIH-1 and SOCS3 expression in KRAS mutant and wild-type patients with colorectal cancer.","date":"2016","source":"Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine","url":"https://pubmed.ncbi.nlm.nih.gov/26749281","citation_count":9,"is_preprint":false},{"pmid":"38163053","id":"PMC_38163053","title":"Hepatitis B Virus-Encoded MicroRNA (HBV-miR-3) Inhibits FIH-1 Expression to Promote Tumor Angiogenesis in HBV-Related Hepatocellular Carcinoma.","date":"2023","source":"Journal of hepatocellular carcinoma","url":"https://pubmed.ncbi.nlm.nih.gov/38163053","citation_count":9,"is_preprint":false},{"pmid":"35610079","id":"PMC_35610079","title":"CircSLC8A1 targets miR-181a-5p/HIF1AN pathway to inhibit the growth, migration and extracellular matrix deposition of human keloid fibroblasts.","date":"2022","source":"Burns : journal of the International Society for Burn Injuries","url":"https://pubmed.ncbi.nlm.nih.gov/35610079","citation_count":8,"is_preprint":false},{"pmid":"35962614","id":"PMC_35962614","title":"Pinoresinol diglucoside ameliorates H/R-induced injury of cardiomyocytes by regulating miR-142-3p and HIF1AN.","date":"2022","source":"Journal of biochemical and molecular toxicology","url":"https://pubmed.ncbi.nlm.nih.gov/35962614","citation_count":8,"is_preprint":false},{"pmid":"21069436","id":"PMC_21069436","title":"Prevention of apoptosis by the interaction between FIH1 and Bax.","date":"2010","source":"Molecular and cellular biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/21069436","citation_count":8,"is_preprint":false},{"pmid":"34655613","id":"PMC_34655613","title":"Structural and thermodynamical insights into the binding and inhibition of FIH-1 by the N-terminal disordered region of Mint3.","date":"2021","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/34655613","citation_count":8,"is_preprint":false},{"pmid":"29228687","id":"PMC_29228687","title":"Interactions between RASA2, CADM1, HIF1AN gene polymorphisms and body fatness with breast cancer: a population-based case-control study in China.","date":"2017","source":"Oncotarget","url":"https://pubmed.ncbi.nlm.nih.gov/29228687","citation_count":6,"is_preprint":false},{"pmid":"35121641","id":"PMC_35121641","title":"Knockout of Factor-Inhibiting HIF (Hif1an) in Colon Epithelium Attenuates Chronic Colitis but Does Not Reduce Colorectal Cancer in Mice.","date":"2022","source":"Journal of immunology (Baltimore, Md. : 1950)","url":"https://pubmed.ncbi.nlm.nih.gov/35121641","citation_count":6,"is_preprint":false},{"pmid":"27112926","id":"PMC_27112926","title":"Molecular response and association analysis of Megalobrama amblycephala fih-1 with hypoxia.","date":"2016","source":"Molecular genetics and genomics : MGG","url":"https://pubmed.ncbi.nlm.nih.gov/27112926","citation_count":6,"is_preprint":false},{"pmid":"37854282","id":"PMC_37854282","title":"Upregulated microRNA-429 confers endometrial stromal cell dysfunction by targeting HIF1AN and regulating the HIF1A/VEGF pathway.","date":"2023","source":"Open medicine (Warsaw, Poland)","url":"https://pubmed.ncbi.nlm.nih.gov/37854282","citation_count":4,"is_preprint":false},{"pmid":"34626458","id":"PMC_34626458","title":"Withdrawn: Y. Yin, X. Lu, M. Yang, J. Shangguan, Y. Zhang. Inhibition of lncRNA MALAT1 reduces myocardial ischemia-reperfusion injury of rat cardiomyocytes through regulating the miR-135a-5p/HIF1AN axis, published in The Journal of Gene Medicine.","date":"2023","source":"The journal of gene medicine","url":"https://pubmed.ncbi.nlm.nih.gov/34626458","citation_count":4,"is_preprint":false},{"pmid":"36285165","id":"PMC_36285165","title":"Danlou Tablet May Alleviate Vascular Injury Caused by Chronic Intermittent Hypoxia through Regulating FIH-1, HIF-1, and Angptl4.","date":"2022","source":"Evidence-based complementary and alternative medicine : eCAM","url":"https://pubmed.ncbi.nlm.nih.gov/36285165","citation_count":3,"is_preprint":false},{"pmid":"34714626","id":"PMC_34714626","title":"Kinetic Studies of the Hydrogen Atom Transfer in a Hypoxia-Sensing Enzyme, FIH-1: KIE and O2 Reactivity.","date":"2021","source":"Biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/34714626","citation_count":3,"is_preprint":false},{"pmid":"37966352","id":"PMC_37966352","title":"The mechanism of lncRNA SNHG1 in osteogenic differentiation via miR-497-5p/ HIF1AN axis.","date":"2024","source":"Connective tissue research","url":"https://pubmed.ncbi.nlm.nih.gov/37966352","citation_count":2,"is_preprint":false},{"pmid":"38789648","id":"PMC_38789648","title":"Molecular characterization and function of hif1a and fih1 in response to acute thermal stress in American shad (Alosa sapidissima).","date":"2024","source":"Fish physiology and biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/38789648","citation_count":2,"is_preprint":false},{"pmid":"39986587","id":"PMC_39986587","title":"CircPVT1 promotes periodontitis progression by regulating miR-24-3p/HIF1AN pathway.","date":"2025","source":"Journal of stomatology, oral and maxillofacial surgery","url":"https://pubmed.ncbi.nlm.nih.gov/39986587","citation_count":2,"is_preprint":false},{"pmid":"39154583","id":"PMC_39154583","title":"Cancer cell-derived exosomes promote NSCLC progression via the miR-199b-5p/HIF1AN axis.","date":"2024","source":"Molecular immunology","url":"https://pubmed.ncbi.nlm.nih.gov/39154583","citation_count":1,"is_preprint":false},{"pmid":"38823472","id":"PMC_38823472","title":"Rynchopeterine inhibits the formation of hypertrophic scars by regulating the miR-21/HIF1AN axis.","date":"2024","source":"Experimental cell research","url":"https://pubmed.ncbi.nlm.nih.gov/38823472","citation_count":1,"is_preprint":false},{"pmid":"40024754","id":"PMC_40024754","title":"Prolyl hydroxylase domain enzymes (isoforms 1-3, PHD1-3), but not factor-inhibiting HIF-1 (FIH-1), interact with the IKK complex and attenuate LPS-activated NF-kappa-B.","date":"2025","source":"The Journal of toxicological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/40024754","citation_count":1,"is_preprint":false},{"pmid":"40059075","id":"PMC_40059075","title":"PML Regulated HIF1AN Ubiquitination and Activated PI3K/AKT Pathway to Promote Bone Marrow Mesenchymal Stem Cells Osteogenic Differentiation.","date":"2025","source":"International journal of stem cells","url":"https://pubmed.ncbi.nlm.nih.gov/40059075","citation_count":0,"is_preprint":false},{"pmid":"41716025","id":"PMC_41716025","title":"Hypoxia‑induced miR‑135b‑5p promotes neuroendocrine differentiation of prostate cancer cells through HIF1AN‑HIF1α axis.","date":"2026","source":"Oncology reports","url":"https://pubmed.ncbi.nlm.nih.gov/41716025","citation_count":0,"is_preprint":false},{"pmid":"40762438","id":"PMC_40762438","title":"FIH-1 Deletion in Intestinal Epithelial Cells Causes Radioprotection by Activating PI3K-AKT Pathway.","date":"2025","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/40762438","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":38189,"output_tokens":5861,"usd":0.101241,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":14614,"output_tokens":5023,"usd":0.099322,"stage2_stop_reason":"end_turn"},"total_usd":0.200563,"stage1_batch_id":"msgbatch_01DUPACZi7AAWwuZiUjseQF4","stage2_batch_id":"msgbatch_018yt7chE3f48PEFtDwtj6AE","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2002,\n      \"finding\": \"FIH-1 is an asparaginyl hydroxylase (Fe(II)-dependent, O2-dependent dioxygenase) that hydroxylates an asparagine residue within the C-terminal transactivation domain (C-TAD) of HIF-1α, blocking its association with transcriptional coactivators and suppressing HIF transcriptional activity under normoxia.\",\n      \"method\": \"In vitro hydroxylase assay, Fe(II)-dependence tests, molecular O2 substrate assays\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct in vitro enzymatic assay establishing catalytic mechanism, independently corroborated by multiple subsequent studies\",\n      \"pmids\": [\"12080085\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"FIH-1 physically binds to HIF-1α and inhibits its transactivation function; FIH-1 also binds to VHL, and VHL recruits histone deacetylases to further repress HIF-1α transactivation, providing a dual mechanism for O2-dependent regulation of HIF-1α activity.\",\n      \"method\": \"Co-immunoprecipitation, mammalian two-hybrid, transactivation reporter assays\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP and functional reporter assays, replicated across subsequent studies\",\n      \"pmids\": [\"11641274\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"FIH-1 undergoes uncoupled O2-activation in the absence of its HIF-1α substrate, resulting in auto-hydroxylation of Trp296 (forming an Fe(III)-O-Trp296 chromophore) and oxidation of Met275; the enzyme active site contains high-spin Fe(III) in both outcomes, with distorted octahedral geometry as shown by XAS and EPR. Co(II)-substituted FIH-1 exists in a mixture of 5- and 6-coordinate forms, indicating the resting enzyme can readily undergo uncoupled O2-activation by loss of an H2O ligand.\",\n      \"method\": \"UV-Vis spectroscopy, EPR, XAS, in vitro uncoupling assay, Co(II) substitution\",\n      \"journal\": \"Journal of inorganic biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — multiple spectroscopic and biochemical methods in a single study establishing mechanistic intermediates\",\n      \"pmids\": [\"18805587\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Crystal structures of human FIH-1 in complex with quinol inhibitors (Clioquinol and 8-Hydroxyquinoline) show that these compounds bind the FIH-1 active site by coordinating the Fe(II) ion, thereby blocking binding of the co-substrate 2-oxoglutarate (2OG) and inhibiting hydroxylase activity.\",\n      \"method\": \"X-ray crystallography (crystal structures of FIH-1:inhibitor complexes)\",\n      \"journal\": \"Molecules and cells\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structures with two inhibitors provide direct structural mechanism for inhibition\",\n      \"pmids\": [\"20396966\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Kinetic isotope effect (KIE) studies with deuterated peptide substrate demonstrate that FIH performs canonical hydrogen atom transfer (HAT) on the asparagine substrate (Dkcat = 10 ± 1), that this step is partially rate-limiting on kcat, and that the deuterated substrate causes uncoupled O2-activation allowing spectroscopic observation of the ferryl intermediate. The close energy barrier between αKG decarboxylation and HAT is identified as the mechanistic basis for FIH's role as an O2-sensing enzyme.\",\n      \"method\": \"Steady-state kinetics with deuterated substrate, KIE measurements, spectroscopic detection of ferryl intermediate\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — mechanistic probe with KIE, ferryl intermediate detection, and internal competition analysis in a single rigorous study\",\n      \"pmids\": [\"34714626\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"FIH-1 transcription in renal cell carcinoma is repressed by the homeodomain protein CDP/Cut, which binds a defined site on the FIH-1 promoter in a PKCζ-dependent manner (with Ser987 phosphorylation of CDP as the regulatory phosphosite); this repression reduces FIH-1 levels and permits increased HIF-1 activity and downstream target gene (VEGF, GLUT-1) expression.\",\n      \"method\": \"Chromatin immunoprecipitation, luciferase promoter assay, PKCζ inhibition, site-directed mutagenesis (CDP S987)\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP, promoter reporter, and mutagenesis in a single study with multiple orthogonal methods\",\n      \"pmids\": [\"17682059\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"FIH-1 hydroxylates the ankyrin repeat domain (ARD)-containing protein ASPP2 at asparagine 986 (N986); this hydroxylation promotes binding of ASPP2 to the cell polarity regulator Par-3. FIH-1 depletion or hydroxylase inhibition (DMOG) causes ASPP2 to relocate from cell-cell contacts to the cytosol without affecting ASPP2 stability, p53 interaction, apoptosis, or proliferation.\",\n      \"method\": \"Mass spectrometry substrate identification, Co-IP, in vitro hydroxylation assay, immunofluorescence localization, DMOG pharmacological inhibition\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — in vitro hydroxylation, MS identification of hydroxylation site, Co-IP, and localization in one study\",\n      \"pmids\": [\"23606740\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Gankyrin binds to and sequesters FIH-1, reducing FIH-1/HIF-1α interaction, thereby increasing HIF-1 transcriptional activity and VEGF production, and promoting vascular tumor formation in transgenic mouse livers.\",\n      \"method\": \"Co-immunoprecipitation, transgenic mouse model, cell-based HIF activity assays\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus in vivo transgenic model, single lab\",\n      \"pmids\": [\"23376718\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"FIH-1 positively regulates keratinocyte migration by binding to LRRK1 (a regulator of EGFR endosomal trafficking), preventing EGFR/LRRK1 complex formation, thereby enhancing EGFR signaling and downstream ERK1/2 activation. FIH-1 null mice show delayed wound healing.\",\n      \"method\": \"In vitro scratch wound assays, Co-IP (FIH-1/LRRK1 interaction), FIH-1 knockout mice, EGFR/MAPK pathway analysis\",\n      \"journal\": \"The American journal of pathology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, KO mouse phenotype, pathway analysis; single lab\",\n      \"pmids\": [\"25455687\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"FIH-1 negatively regulates glycogen metabolism in corneal epithelial cells through the AKT/GSK-3β/glycogen synthase pathway in a hydroxylase-independent manner (enzyme-dead FIH-1 fails to reduce glycogen), and independently of HIF-1α.\",\n      \"method\": \"Retroviral transduction of enzyme-dead FIH-1 mutant, antagomir knockdown, glycogen assay, AKT/GSK-3β western blot, HIF-1α reporter (negative result for HIF-1α dependence)\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — enzyme-dead mutant plus pathway rescue experiments; single lab\",\n      \"pmids\": [\"22532441\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"FIH-1 silencing in human vascular endothelial cells (HUVECs) leads to increased Notch2 activity, enhanced Hey-1 expression, and selective repression of survivin, resulting in apoptosis and growth arrest. FIH-1 is proposed to repress Notch2 activity, controlling endothelial cell survival.\",\n      \"method\": \"siRNA knockdown, western blot (Notch2, Hey-1, survivin), apoptosis assays\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — knockdown with defined pathway readout (Notch2→Hey-1→survivin) but single lab, no direct FIH-1/Notch2 binding demonstrated\",\n      \"pmids\": [\"25837583\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"FIH-1 directly interacts with the pro-apoptotic protein Bax and functions as a cytosolic retention factor for Bax, preventing its translocation to mitochondria in response to apoptotic stimuli; FIH-1 overexpression suppresses Bax-mediated apoptosis, while FIH-1 deficiency accelerates it.\",\n      \"method\": \"Co-immunoprecipitation, GST pull-down, apoptosis assays, subcellular fractionation\",\n      \"journal\": \"Molecular and cellular biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP, GST pull-down, and functional apoptosis assays; single lab\",\n      \"pmids\": [\"21069436\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"KANK3 is a substrate for HIF1AN asparaginyl hydroxylation; in vitro hydroxylation assay confirmed hydroxylation, and mass spectrometry identified three hydroxylated asparagine residues within the ankyrin repeat domain of KANK3. KANK3 tumor-suppressive effects on cell migration/invasion were not observed under hypoxic conditions, consistent with oxygen-dependent HIF1AN activity.\",\n      \"method\": \"In vitro hydroxylation assay, mass spectrometry, cell migration/invasion assays under normoxia vs. hypoxia\",\n      \"journal\": \"Cell biology international\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro assay and MS identification of hydroxylation sites; single lab, no mutagenesis validation of individual sites\",\n      \"pmids\": [\"29047187\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"FIH-1 inhibits HIF-1-mediated transcription of GLUT1 and VEGF-A even under hypoxic conditions in glioblastoma cells, and FIH-1 is more potent than PTEN in inhibiting HIF function in this context.\",\n      \"method\": \"Retroviral FIH-1 expression, siRNA knockdown, quantitative RT-PCR for HIF target genes (GLUT1, VEGF-A) under hypoxia\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — gain- and loss-of-function with defined transcriptional readouts; single lab\",\n      \"pmids\": [\"24465898\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"In VHL-defective CCRCC cells expressing both HIF-1α and HIF-2α, FIH-1 knockdown increases HIF target gene expression and apoptosis. In 786-O cells expressing only HIF-2α, FIH-1 knockdown has no effect; re-introduction of HIF-1α restores sensitivity to FIH-1 knockdown. This establishes FIH-1's substrate preference for HIF-1α over HIF-2α in modulating both target gene expression and cell viability.\",\n      \"method\": \"siRNA knockdown, retroviral gene expression, qRT-PCR, Annexin V/propidium iodide apoptosis assay\",\n      \"journal\": \"British journal of cancer\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic complementation (HIF-1α rescue) plus matched isogenic cell lines; single lab\",\n      \"pmids\": [\"21386837\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"FIH-1 interacts with Plectin1 and STAT1 (novel binding partners identified by BioID proteomics), as well as known partners ASPP2 and HDAC1, to positively regulate ΔNp63α expression in keratinocytes. ASPP2 knockdown upregulates ΔNp63α; FIH-1 regulates GADD45α (a negative regulator of ΔNp63α) via HDAC1 interaction.\",\n      \"method\": \"BioID proteomics, Co-IP, siRNA knockdown, FIH-1 conditional transgenic and knockout mouse\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — BioID proximity labeling, Co-IP, and in vivo mouse model; single lab\",\n      \"pmids\": [\"31914679\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"ANKDD1A directly interacts with FIH1 and upregulates FIH1 activity, leading to decreased HIF1α transcriptional activity and reduced HIF1α protein half-life. This decreases glucose uptake, lactate production, and autophagy while inducing apoptosis in glioblastoma cells under hypoxia.\",\n      \"method\": \"Co-immunoprecipitation (ANKDD1A-FIH1 interaction), HIF1α stability assay, glucose/lactate measurement, autophagy and apoptosis assays\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — Co-IP plus functional readouts; single lab\",\n      \"pmids\": [\"30082910\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"The N-terminal disordered region of Mint3 (amino acids 78–88 as core binding site) binds FIH-1 with a large enthalpy and entropy change consistent with disorder-to-order transition upon binding; thermodynamic mapping by ITC with truncated Mint3 constructs identified the core binding site and distinguished affinity-contributing from enthalpy-only-contributing regions.\",\n      \"method\": \"Circular dichroism, NMR, hydrogen/deuterium exchange-MS, isothermal titration calorimetry (ITC) with truncation series\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — multiple orthogonal biophysical methods (CD, NMR, HDX-MS, ITC) with systematic truncation analysis in one rigorous study\",\n      \"pmids\": [\"34655613\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"FIH-1 knockout in intestinal epithelial cells (FIH-1 silencing in HIEC-6 cells) activates the PI3K-AKT signaling pathway and cell cycle redistribution into S-phase, conferring radioprotection by mitigating radiation-induced cell damage.\",\n      \"method\": \"shRNA knockdown, colony formation assay, cell cycle analysis, apoptosis assay, RNA-Seq, western blot\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — RNA-Seq plus western blot pathway analysis with functional radiosensitivity assays; single lab\",\n      \"pmids\": [\"40762438\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ASB9 (E3 ubiquitin ligase) directly interacts with HIF1AN and promotes its ubiquitination and degradation; ASB9 overexpression inhibits human SSC line proliferation and increases apoptosis, effects reversed by HIF1AN re-expression.\",\n      \"method\": \"Co-immunoprecipitation, western blot (HIF1AN protein level), functional proliferation/apoptosis assays with rescue\",\n      \"journal\": \"Biological research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — Co-IP demonstrating direct interaction, functional rescue; single lab\",\n      \"pmids\": [\"36683111\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"FIH-1 knockout in colon epithelial cells (Vil1-Cre/FIH+f/+f mice) attenuates chronic colitis severity and reduces infiltrating macrophages and immune-response gene expression, but does not alter colorectal tumor occurrence in an AOM/DSS model.\",\n      \"method\": \"Conditional knockout mouse (Vil1-Cre), AOM/DSS colitis-cancer model, RNA-seq of colon tissue, histology, immunohistochemistry\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo conditional KO with transcriptomics and histology; single lab\",\n      \"pmids\": [\"35121641\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"FIH-1 does not interact with IKKα/β or IκBα (negative result by immunoprecipitation), and FIH-1 overexpression does not affect IKKα/β or p65 protein levels, indicating FIH-1 does not regulate the classical NF-κB pathway.\",\n      \"method\": \"Immunoprecipitation, western blot, FIH-1 overexpression\",\n      \"journal\": \"The Journal of toxicological sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — negative result from Co-IP and overexpression; single lab, single method per assay\",\n      \"pmids\": [\"40024754\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"FIH-1 (HIF1AN) physically interacts with Mindbomb (Mib) E3 ubiquitin ligase in zebrafish and cultured cells (co-localization confirmed); depletion of fih-1 in zebrafish embryos induces ectopic VEGF-A expression and ectopic intersegmental vessel sprouting, while fih-1 overexpression attenuates ISV formation in a VEGF-A-rescuable manner, establishing FIH-1 as an anti-angiogenic regulator of VEGF-A signaling.\",\n      \"method\": \"Yeast two-hybrid (Mib interaction), co-localization in cultured cells, zebrafish morpholino knockdown, fih-1 mRNA overexpression, ISV imaging\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — yeast two-hybrid plus in vivo zebrafish gain- and loss-of-function with rescue; single lab\",\n      \"pmids\": [\"25347788\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"PML (promyelocytic leukemia protein) directly binds HIF1AN and promotes its ubiquitination-mediated degradation; PML knockdown or HIF1AN upregulation suppresses osteogenic differentiation of BMSCs; HIF1α directly binds the SOD3 promoter; PML overexpression promotes osteogenesis via HIF1AN/HIF1α/SOD3 axis and PI3K/AKT pathway.\",\n      \"method\": \"Co-immunoprecipitation, immunofluorescence, ChIP, dual-luciferase reporter, western blot, ALP/Alizarin red staining\",\n      \"journal\": \"International journal of stem cells\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and ChIP with functional differentiation readouts; single lab\",\n      \"pmids\": [\"40059075\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"HIF1AN (FIH-1) is an Fe(II)- and O2-dependent asparaginyl hydroxylase that hydroxylates Asn803 of HIF-1α's C-terminal transactivation domain, blocking CBP/p300 co-activator recruitment and suppressing HIF-1 transcriptional activity under normoxia; it also hydroxylates ankyrin repeat domain (ARD)-containing substrates (including ASPP2, KANK3, and Notch) to modulate their protein–protein interactions, and performs HIF-1α-independent functions including regulation of AKT/GSK-3β-dependent glycogen metabolism, cytosolic retention of Bax to suppress apoptosis, and EGFR/ERK-mediated keratinocyte migration via LRRK1 binding, with its own expression controlled transcriptionally by the CDP/Cut–PKCζ axis and its protein stability regulated by ASB9 and PML-mediated ubiquitination.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"HIF1AN (FIH-1) is an Fe(II)- and O2-dependent asparaginyl hydroxylase that functions as the principal oxygen sensor restraining HIF transcriptional activity under normoxia by hydroxylating an asparagine in the C-terminal transactivation domain of HIF-1\\u03b1, blocking coactivator recruitment and binding both HIF-1\\u03b1 and VHL to achieve dual O2-dependent repression [#0, #1]. Its catalytic cycle proceeds through canonical hydrogen-atom transfer on the asparagine via a ferryl intermediate, and the near-equal energy barrier between \\u03b1KG decarboxylation and HAT is the mechanistic basis for its O2-sensing role; in the absence of substrate the enzyme undergoes uncoupled O2-activation with auto-hydroxylation of Trp296, and quinol inhibitors block activity by coordinating the active-site Fe and excluding 2-oxoglutarate [#2, #3, #4]. FIH-1 shows substrate preference for HIF-1\\u03b1 over HIF-2\\u03b1 and suppresses HIF target genes such as GLUT1 and VEGF-A even under hypoxia [#13, #14]. Beyond HIF, FIH-1 hydroxylates ankyrin-repeat-domain substrates including ASPP2 at N986\\u2014promoting ASPP2/Par-3 binding and junctional localization\\u2014and KANK3, coupling its enzymatic activity to protein-protein interactions [#6, #12]. It also performs HIF-1\\u03b1-independent functions: hydroxylase-independent suppression of glycogen metabolism via the AKT/GSK-3\\u03b2 axis, cytosolic retention of pro-apoptotic Bax to suppress apoptosis, and regulation of keratinocyte migration and \\u0394Np63\\u03b1 expression through binding partners including LRRK1, Plectin1, STAT1, ASPP2 and HDAC1 [#9, #11, #8, #15]. FIH-1 abundance is set transcriptionally by CDP/Cut\\u2013PKC\\u03b6 repression and post-translationally by ASB9- and PML-mediated ubiquitination and degradation, and its activity is modulated by interacting proteins that either sequester it (Gankyrin) or stimulate it (ANKDD1A) to tune HIF output [#5, #19, #23, #7, #16].\",\n  \"teleology\": [\n    {\n      \"year\": 2001,\n      \"claim\": \"Established that FIH-1 is a physical inhibitor of HIF-1\\u03b1 transactivation, defining a new oxygen-regulatory node distinct from the VHL/proteolysis axis.\",\n      \"evidence\": \"Co-IP, mammalian two-hybrid, and transactivation reporter assays showing FIH-1 binds HIF-1\\u03b1 and VHL\",\n      \"pmids\": [\"11641274\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define the enzymatic basis of inhibition\", \"Mechanism of VHL-mediated HDAC recruitment not resolved\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Identified the molecular mechanism: FIH-1 is an Fe(II)/O2-dependent asparaginyl hydroxylase that modifies the HIF-1\\u03b1 C-TAD asparagine to block coactivator binding, explaining O2-dependent HIF suppression.\",\n      \"evidence\": \"In vitro hydroxylase assays with Fe(II)- and molecular O2-dependence tests\",\n      \"pmids\": [\"12080085\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Catalytic intermediates and rate-limiting steps not characterized\", \"Non-HIF substrates not yet known\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Showed FIH-1 expression itself is controlled, with CDP/Cut repressing the FIH-1 promoter in a PKC\\u03b6-dependent manner to derepress HIF activity in renal carcinoma.\",\n      \"evidence\": \"ChIP, luciferase promoter assays, PKC\\u03b6 inhibition, and CDP S987 mutagenesis\",\n      \"pmids\": [\"17682059\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Generality of CDP/Cut control beyond RCC unknown\", \"Upstream signals activating PKC\\u03b6 not defined\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Defined the active-site chemistry of substrate-free FIH-1, revealing uncoupled O2-activation, Trp296 auto-hydroxylation, and high-spin Fe(III) geometry.\",\n      \"evidence\": \"UV-Vis, EPR, XAS, Co(II) substitution, and in vitro uncoupling assays\",\n      \"pmids\": [\"18805587\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological consequence of auto-hydroxylation unclear\", \"Did not capture the productive ferryl intermediate\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Provided structural mechanism of inhibition, showing quinol compounds coordinate the active-site Fe and exclude 2OG, and revealed FIH-1 acts as a cytosolic Bax retention factor independent of its HIF role.\",\n      \"evidence\": \"X-ray crystallography of FIH-1:inhibitor complexes; Co-IP, GST pull-down, fractionation, and apoptosis assays for Bax\",\n      \"pmids\": [\"20396966\", \"21069436\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether Bax retention requires hydroxylase activity not established\", \"Structural basis of Bax interaction unknown\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Demonstrated FIH-1 substrate preference for HIF-1\\u03b1 over HIF-2\\u03b1 in modulating target genes and cell viability, using isogenic CCRCC lines and HIF-1\\u03b1 complementation.\",\n      \"evidence\": \"siRNA knockdown, retroviral HIF-1\\u03b1 re-introduction, qRT-PCR, and apoptosis assays\",\n      \"pmids\": [\"21386837\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of isoform discrimination not addressed\", \"Single lab\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Revealed a hydroxylase-independent, HIF-1\\u03b1-independent function in glycogen metabolism via the AKT/GSK-3\\u03b2 pathway, broadening FIH-1 biology beyond catalysis.\",\n      \"evidence\": \"Enzyme-dead FIH-1 mutant, antagomir knockdown, glycogen assays, and pathway western blots\",\n      \"pmids\": [\"22532441\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular target linking FIH-1 to AKT not identified\", \"Generality beyond corneal epithelium unknown\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Extended FIH-1 substrate range to ARD proteins, showing ASPP2 N986 hydroxylation controls ASPP2/Par-3 binding and junctional localization, and identifying Gankyrin as a sequestering regulator that boosts HIF activity in vivo.\",\n      \"evidence\": \"MS site identification, in vitro hydroxylation, Co-IP, immunofluorescence (ASPP2); Co-IP and transgenic mouse (Gankyrin)\",\n      \"pmids\": [\"23606740\", \"23376718\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional consequence of ASPP2 mislocalization on tissue physiology unclear\", \"Whether Gankyrin affects FIH-1 catalysis directly not shown\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Expanded FIH-1 into angiogenesis, endothelial survival, keratinocyte migration and continued HIF suppression under hypoxia, linking it to Notch2, LRRK1/EGFR, and Mindbomb.\",\n      \"evidence\": \"siRNA/Notch readouts (HUVEC); scratch assays, Co-IP, KO mice (LRRK1); zebrafish morpholino, Y2H, ISV imaging (Mib); retroviral/siRNA qRT-PCR in glioblastoma\",\n      \"pmids\": [\"25837583\", \"25455687\", \"25347788\", \"24465898\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct FIH-1/Notch2 binding not demonstrated\", \"Whether LRRK1 and Mib effects depend on hydroxylation unresolved\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Confirmed KANK3 as an ARD hydroxylation substrate with three hydroxylated asparagines, reinforcing FIH-1's role as a regulator of ARD-containing tumor suppressors under oxygen control.\",\n      \"evidence\": \"In vitro hydroxylation, mass spectrometry, and normoxia-vs-hypoxia migration/invasion assays\",\n      \"pmids\": [\"29047187\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Individual sites not validated by mutagenesis\", \"Functional impact of each hydroxylation site undefined\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Identified ANKDD1A as a direct activator that upregulates FIH-1 to lower HIF1\\u03b1 stability and activity, suppressing glycolysis and autophagy in glioblastoma.\",\n      \"evidence\": \"Co-IP, HIF1\\u03b1 stability assays, glucose/lactate measurement, autophagy/apoptosis assays\",\n      \"pmids\": [\"30082910\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which ANKDD1A enhances catalysis unknown\", \"Single lab\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Mapped an interactome (Plectin1, STAT1, ASPP2, HDAC1) through which FIH-1 positively regulates \\u0394Np63\\u03b1 in keratinocytes, integrating it into epidermal gene regulation.\",\n      \"evidence\": \"BioID proteomics, Co-IP, siRNA, and conditional transgenic/knockout mice\",\n      \"pmids\": [\"31914679\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether these interactions require hydroxylase activity unclear\", \"Direct hydroxylation of new partners not tested\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Resolved the catalytic basis of O2 sensing\\u2014partially rate-limiting HAT through a ferryl intermediate with a close barrier to \\u03b1KG decarboxylation\\u2014and thermodynamically characterized disordered-substrate (Mint3) binding via disorder-to-order transition.\",\n      \"evidence\": \"KIE with deuterated substrate and ferryl detection (kinetics); CD, NMR, HDX-MS, ITC with truncations (Mint3)\",\n      \"pmids\": [\"34714626\", \"34655613\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In-cell relevance of the kinetic barrier not tested\", \"Structure of the FIH-1/Mint3 complex not determined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Established post-translational control of FIH-1 abundance, with ASB9 directly binding and targeting HIF1AN for ubiquitin-mediated degradation in spermatogonial stem cells.\",\n      \"evidence\": \"Co-IP, HIF1AN protein-level western blots, and proliferation/apoptosis rescue assays\",\n      \"pmids\": [\"36683111\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Ubiquitination sites not mapped\", \"Generality beyond SSC lines unknown\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Refined the regulatory and signaling boundaries of FIH-1: PML drives its ubiquitin-mediated degradation to promote osteogenesis, while FIH-1 was shown not to regulate the classical NF-\\u03baB pathway.\",\n      \"evidence\": \"Co-IP, ChIP, luciferase, differentiation assays (PML); IP and overexpression negative results (NF-\\u03baB)\",\n      \"pmids\": [\"40059075\", \"40024754\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"PML and ASB9 relationship to each other unresolved\", \"Negative NF-\\u03baB result is single-lab\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How FIH-1 selects among its many catalytic (HIF-1\\u03b1, ASPP2, KANK3) and non-catalytic (Bax, LRRK1, AKT) functions in different tissues, and which depend on hydroxylase activity versus scaffolding, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified rule distinguishing hydroxylation-dependent from independent functions\", \"Tissue-specific substrate hierarchy undefined\", \"Structural models for most non-HIF substrates lacking\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016491\", \"supporting_discovery_ids\": [0, 2, 4, 6, 12]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 6, 12]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [1, 11, 13]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [6, 11]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [0, 1, 13, 14]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [1, 13, 15]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [11, 10]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"HIF1A\", \"VHL\", \"ASPP2\", \"KANK3\", \"LRRK1\", \"BAX\", \"HDAC1\", \"ANKDD1A\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}