{"gene":"ILF3","run_date":"2026-06-10T01:55:23","timeline":{"discoveries":[{"year":1994,"finding":"NF90 (90-kDa subunit) and NF45 (45-kDa subunit) together constitute the NF-AT transcription factor complex that binds the antigen receptor response element in the IL-2 promoter; immunofluorescence demonstrated both proteins are nuclear in Jurkat T-cells, and the complex's DNA-binding activity is enhanced by T-cell stimulation but blocked by cyclosporin A or FK506.","method":"Protein purification, cDNA cloning, polyhistidine-tag affinity purification, immunofluorescence microscopy, electrophoretic mobility shift assay (EMSA)","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — protein biochemistry, cloning, and functional DNA-binding assays with pharmacological validation, foundational study replicated across many subsequent papers","pmids":["7519613"],"is_preprint":false},{"year":1998,"finding":"NF90 and NF45 physically interact with DNA-PK (DNA-PKcs and Ku subunits), stabilize the DNA-PKcs–Ku–DNA complex, and are substrates for DNA-PK phosphorylation in vitro; recombinant NF90 promotes DNA-PKcs–Ku–DNA complex formation, and antibodies to NF90 or NF45 immunoprecipitate DNA-PKcs.","method":"EMSA, amino-terminal sequence analysis, immunoblotting, in vitro kinase assay, immunoprecipitation","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — multiple orthogonal biochemical methods (EMSA, kinase assay, reciprocal IP) in a single rigorous study; independently reproduced in later work","pmids":["9442054"],"is_preprint":false},{"year":2000,"finding":"TCP80/NF90 binds the coding region of glucocerebrosidase (GCase) mRNA and inhibits its translation by reducing polysome association, an activity reconstituted in insect Sf9 cells that lack endogenous NF90.","method":"Baculovirus/Sf9 insect cell reconstitution, in vitro translation assay, polysome sedimentation, RNA binding assay","journal":"Molecular genetics and metabolism","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution of translation inhibition in a heterologous system with purified recombinant protein and functional validation of polysome displacement","pmids":["10873392"],"is_preprint":false},{"year":2001,"finding":"NFAR-1 (NF90, 90 kDa) and NFAR-2 (NF110, 110 kDa), derived from the same gene by alternative splicing, are phosphorylated by PKR, reciprocally co-immunoprecipitate with PKR, colocalize with PKR in the nucleus, associate with both pre-mRNAs and spliced mRNAs, and interact with RNA-processing factors FUS and SMN via their C-termini.","method":"Reciprocal co-immunoprecipitation, subcellular colocalization, in vitro phosphorylation assay, transfection-based gene expression assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP plus multiple orthogonal functional assays in one study","pmids":["11438536"],"is_preprint":false},{"year":2002,"finding":"NF90 binds the AU-rich element-containing 3'UTR of IL-2 mRNA, slows its degradation, and nuclear export of NF90 to the cytoplasm upon T-cell activation is required for IL-2 mRNA stabilization.","method":"RNA–protein binding assay, mRNA half-life measurement, subcellular fractionation, T-cell stimulation assays","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct RNA binding, functional mRNA stability assay, and nucleocytoplasmic translocation linked to function; replicated in multiple subsequent studies","pmids":["12504009"],"is_preprint":false},{"year":2003,"finding":"ILF3 preferentially binds minihelix-motif RNAs (e.g., adenovirus VA1 RNA) and assembles a quaternary export complex with exportin-5, RanGTP, and the RNA; this complex mediates co-transport of VA1 RNA and ILF3 from nucleus to cytoplasm, identifying exportin-5 as the export receptor for ILF3.","method":"Gel retardation assay, GST pulldown, microinjection in HeLa cell nuclei, transfection experiments","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — biochemical reconstitution of the export complex plus live-cell nuclear microinjection validating functional export","pmids":["14570900"],"is_preprint":false},{"year":2003,"finding":"NFAR proteins (NF90/NF110 group) are recruited by the BVDV replication machinery through specific binding to both 5' and 3' non-translated regions of the viral RNA; mutation of the NF90 interaction sites in the 3'NTR yields replication-deficient RNA, and RNAi depletion of RNA helicase A (an NFAR group member) inhibits replication.","method":"RNA–protein binding assay, viral RNA mutagenesis, RNAi knockdown, viral replication assay","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — mutagenesis of viral RNA binding sites coupled with RNAi functional validation","pmids":["14592965"],"is_preprint":false},{"year":2003,"finding":"Cell-cycle-dependent nucleocytoplasmic shuttling of TCP110/ILF3 is directed by a C-terminal nuclear localization signal; TCP80 (NF90) remains stable in the cytoplasm (t½ ~5 days), whereas TCP110/ILF3 translocates between cytoplasm and nucleus during the cell cycle.","method":"Subcellular fractionation, 5'RACE cDNA cloning, metabolic turnover assay, immunofluorescence","journal":"Molecular genetics and metabolism","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — fractionation plus turnover assays, single lab","pmids":["14654356"],"is_preprint":false},{"year":2004,"finding":"Ilf3 and NF90 directly bind the axonal targeting element (stem-loop in the 3'UTR) of Tau mRNA via their double-stranded RNA-binding motifs; both proteins are detected in neuronal nuclei, cell bodies, and proximal neuritic segments, consistent with a role in Tau mRNA translocation.","method":"Northwestern blotting, specific antibodies, immunolocalization","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Northwestern binding assay and immunolocalization, single lab with two orthogonal approaches","pmids":["15364895"],"is_preprint":false},{"year":2005,"finding":"NF90 knockout mice die perinatally from neuromuscular respiratory failure with skeletal muscle disorganization, decreased myofiber number, and severely reduced expression of MyoD, myogenin, and p21WAF1/CIP1; Northwestern blotting identified NF90 as the principal RNA-binding protein for the p21 and MyoD 3'UTRs in developing muscle, establishing a direct post-transcriptional role in muscle differentiation.","method":"Gene knockout (NF90−/− mice), Northwestern blotting, histology, immunohistochemistry","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — loss-of-function in vivo KO with defined molecular mechanism (direct 3'UTR binding); unique in vivo genetic evidence","pmids":["15746098"],"is_preprint":false},{"year":2007,"finding":"NF90 and NF45 together with Ku80 and Ku70 bind specifically to the IL-2 ARRE/NF-AT DNA sequence in vivo; T-cell activation induces binding of Ku80 and NF90 but decreases Ku70 binding at the IL-2 promoter, and these dynamic changes are blocked by cyclosporin A and triptolide.","method":"Chromatin immunoprecipitation (ChIP), EMSA with monoclonal antibody inhibition, purification from activated Jurkat nuclei","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo ChIP plus EMSA with inhibitory antibodies, multiple cell types tested","pmids":["17389650"],"is_preprint":false},{"year":2007,"finding":"NF90ctv (C-terminal variant of NF90) binds the HIV-1 TAR RNA with high affinity, competes with Tat for TAR binding in vitro, inhibits Tat-transactivation of the HIV-1 LTR in vivo, and is associated with histone H3K4/K9 methylation changes consistent with transcriptional repression.","method":"TAR RNA affinity fractionation, Northwestern blotting, EMSA, luciferase transactivation assay, chromatin histone methylation analysis","journal":"Retrovirology","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — multiple assays (EMSA, cell-based transactivation, histone mark analysis) in a single lab","pmids":["17565699"],"is_preprint":false},{"year":2008,"finding":"AKT phosphorylates NF90 at Ser647 in response to CD28 costimulation; this phosphorylation is necessary and sufficient for nuclear export of NF90 and subsequent IL-2 mRNA stabilization, as a Ser647→Ala mutation abolishes both nuclear export and mRNA stabilization.","method":"In vitro kinase assay, phospho-specific antibody, site-directed mutagenesis, nuclear export assay, mRNA stability assay, T-cell stimulation","journal":"Journal of immunology","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — in vitro kinase assay combined with mutagenesis and functional mRNA/export readouts","pmids":["18097023"],"is_preprint":false},{"year":2008,"finding":"NF90 and NF45 form heterodimeric core complexes (NF90-NF45 and NF110-NF45); depletion of NF45 dramatically decreases NF90/NF110 protein levels (posttranscriptional destabilization), and depletion of NF90 (but not NF110) greatly reduces NF45; depletion of the NF90-NF45 complex retards cell growth by inhibiting DNA synthesis and causes giant multinucleated cells, revealing a specific role in cell division.","method":"RNA interference (RNAi) knockdown, flow cytometry, immunoblot, cell growth assay, time-lapse microscopy","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — systematic RNAi depletion of individual subunits with multiple cellular readouts; clearly distinguishes NF90 from NF110 function","pmids":["18458058"],"is_preprint":false},{"year":2008,"finding":"NF90 and HuR bind the AU-rich 3'UTR of MKP-1 mRNA; H2O2 treatment increases association of MKP-1 mRNA with HuR and NF90 and decreases association with translational repressors TIAR and TIA-1; silencing NF90 or HuR diminishes H2O2-stimulated MKP-1 mRNA stability, and HuR silencing also reduces MKP-1 translation.","method":"Ribonucleoprotein immunoprecipitation (RIP), biotinylated RNA pulldown, mRNA stability assay, siRNA knockdown","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — RIP plus RNA pulldown plus functional mRNA stability measurement with siRNA, multiple orthogonal methods","pmids":["18490444"],"is_preprint":false},{"year":2008,"finding":"NFAR-1 and NFAR-2 retain cellular transcripts in intranuclear foci and regulate mRNA export; loss of NFAR function (embryonic lethal) increases protein synthesis rates; NFAR-depleted murine fibroblasts are dramatically susceptible to VSV replication, demonstrating an innate immune translational surveillance function.","method":"NFAR knockout (embryonic lethal), RNA export assay, polysome/translation rate measurement, VSV infection assay in NFAR-depleted cells","journal":"Proceedings of the National Academy of Sciences","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo KO (embryonic lethality) plus multiple functional assays (export, translation, viral replication) in depleted cells","pmids":["18337511"],"is_preprint":false},{"year":2009,"finding":"The NF90-NF45 complex functions as a negative regulator of miRNA biogenesis: overexpression inhibits pri-miRNA processing to pre-miRNA, causing pri-miRNA accumulation; the complex binds pri-miRNAs (e.g., pri-let-7a) but does not interact with the Microprocessor, suggesting it blocks Microprocessor access; depletion of NF90 reduces pri-let-7a and increases mature let-7a.","method":"Overexpression and RNAi knockdown, pri-miRNA/pre-miRNA/mature miRNA quantification, RNA polymerase II inhibitor experiments, RNA immunoprecipitation","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — bidirectional genetic manipulation (OE and KD) plus negative result (no Microprocessor interaction) with multiple miRNA substrates","pmids":["19398578"],"is_preprint":false},{"year":2009,"finding":"NF90 binds an AU-rich 25–30 nucleotide RNA motif in the 3'UTR of target mRNAs and represses their translation without significantly affecting mRNA stability; insertion of this NF90 motif into an EGFP reporter 3'UTR specifically confers NF90-dependent translational repression as shown by polysome gradient analysis.","method":"RIP-seq, biotinylated transcript pulldown, polysome gradient analysis, EGFP reporter assay, siRNA knockdown","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — genome-wide RIP combined with reporter mutagenesis and polysome profiling; multiple orthogonal methods","pmids":["19850717"],"is_preprint":false},{"year":2010,"finding":"PKCβI phosphorylates NF90 at Ser647 in response to PMA stimulation, triggering nuclear export of NF90 and stabilization of IL-2 mRNA; Ser647→Ala mutation abolishes PMA-induced nuclear export and IL-2 mRNA stabilization.","method":"In vitro kinase assay with PKCβI, phospho-specific antibody, site-directed mutagenesis, nuclear export assay, mRNA stability assay","journal":"Journal of immunology","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — in vitro kinase assay plus mutagenesis plus functional cellular readouts; complements the earlier AKT study","pmids":["20870937"],"is_preprint":false},{"year":2010,"finding":"NF45 and NF90 bind the HS4 element of the IL-13 promoter in T cells and are required for HS4-dependent IL-13 transcriptional activation; the CTGTT binding motif within HS4-3' is critical, and HS4 activity is abrogated in NF45+/− primary Th2 cells and reduced in NF90+/− cells.","method":"DNA affinity chromatography coupled with tandem mass spectrometry, ChIP, EMSA, reporter transfection in primary Th2 cells from NF45+/− and NF90+/− mice","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (mass spec, ChIP, EMSA, haploinsufficiency in primary cells) in one study","pmids":["20051514"],"is_preprint":false},{"year":2011,"finding":"The NF90-NF45 complex is required for DNA double-strand break repair by nonhomologous end joining (NHEJ): immunodepletion of NF90/NF45 reduces in vitro NHEJ activity comparably to DNA-PKcs depletion; NF90/NF45-depleted cells accumulate γ-H2A.X foci and are hypersensitive to ionizing radiation.","method":"In vitro NHEJ assay with immunodepletion, γ-H2A.X foci quantification, ionizing radiation sensitivity assay, time-lapse microscopy, RNAi knockdown","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — in vitro reconstitution (NHEJ assay with immunodepletion) plus multiple in vivo readouts (γH2AX, IR sensitivity)","pmids":["21969602"],"is_preprint":false},{"year":2011,"finding":"NF90 binds the dengue virus 3' stem-loop RNA via affinity chromatography; NF90 depletion by siRNA reduces dengue RNA levels and infectious virus production by 50–70%; cytoplasmic relocalization of NF90 occurs in dengue-infected cells.","method":"RNA affinity column chromatography, siRNA knockdown, viral RNA quantification, infectious virus titration, immunofluorescence","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 / Strong — RNA affinity binding plus functional siRNA depletion with two viral replication readouts","pmids":["21386893"],"is_preprint":false},{"year":2012,"finding":"NF45 dimerizes with NF90 (and related proteins SPNR and Zfr) through the DZF domain, which has structural similarity to the template-free nucleotidyltransferase family but lacks catalytic residues; crystal structure at 1.9-Å resolution reveals the dimerization interface, confirmed by co-immunoprecipitation with site-specific mutants.","method":"X-ray crystallography (1.9 Å), co-immunoprecipitation with site-directed mutants","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure combined with mutagenesis-validated functional interactions","pmids":["22833610"],"is_preprint":false},{"year":2012,"finding":"YM155 directly binds ILF3 and disrupts the ILF3/p54(nrb) complex, causing different subcellular localization of ILF3 and p54(nrb), which results in suppression of survivin promoter activity.","method":"Small-molecule binding assay, co-immunoprecipitation, subcellular localization analysis, survivin promoter activity assay","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP and localization assays, single lab; small-molecule target engagement inferred rather than directly demonstrated by structural methods","pmids":["22842455"],"is_preprint":false},{"year":2012,"finding":"NF90 coordinately represses translation (not stability) of multiple SASP factor mRNAs (MCP-1, GROα, IL-6, IL-8) in proliferating fibroblasts; NF90 levels decline in senescent cells, allowing SASP factor expression to rise.","method":"RIP, siRNA knockdown, ELISA for secreted proteins, mRNA stability assay, immunoblot","journal":"Aging","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — RIP combined with KD and functional protein/mRNA measurement; single lab","pmids":["23117626"],"is_preprint":false},{"year":2012,"finding":"Depletion of NF90/NF45 in HPV-transformed cervical carcinoma cells restores p53 protein (post-transcriptionally, not at mRNA level) and p21 (via p53-dependent transcription); NF90 knockdown attenuates HPV E6 RNA levels and inhibits transcription from the HPV early promoter, revealing that NF90/NF45 supports viral E6 expression.","method":"siRNA knockdown, RT-PCR, immunoblot, PARP cleavage assay, camptothecin sensitivity assay","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 2 / Strong — systematic knockdown with multiple molecular and cellular readouts distinguishing transcriptional from post-transcriptional effects","pmids":["23208500"],"is_preprint":false},{"year":2014,"finding":"NF90 binds the 5'-terminal double-stranded region of the HCV genome; siRNA depletion of NF90 significantly impairs HCV RNA replication and reduces infectious virus yields; NF90 co-immunoprecipitates with NS5A in an RNA-dependent manner and co-fractionates with detergent-resistant membranes containing viral replication complexes.","method":"Biotinylated RNA pulldown with mass spectrometry, siRNA knockdown, viral RNA quantification, co-immunoprecipitation, detergent-resistant membrane fractionation","journal":"Journal of virology","confidence":"High","confidence_rationale":"Tier 2 / Strong — RNA affinity MS plus functional depletion plus RIP in infected cells with multiple orthogonal methods","pmids":["24719423"],"is_preprint":false},{"year":2014,"finding":"NF90 interacts with PKR through its C-terminal domain independently of NF90 RNA-binding properties; NF90 knockdown reduces PKR phosphorylation in response to dsRNA/influenza infection; NF90 is a component of stress granules, and NF90 depletion reduces dsRNA-induced stress granule formation; an NS1 mutant influenza virus specifically attenuated in PKR inhibition replicates indistinguishably from WT in NF90-depleted cells, placing NF90 in the PKR–stress granule antiviral pathway.","method":"Co-immunoprecipitation, C-terminal domain mapping, siRNA knockdown, PKR phosphorylation assay, stress granule immunofluorescence, viral replication assay with NS1 mutant","journal":"Journal of immunology","confidence":"High","confidence_rationale":"Tier 2 / Strong — protein interaction mapping plus genetic epistasis using NS1 mutant virus, multiple orthogonal methods","pmids":["24623135"],"is_preprint":false},{"year":2014,"finding":"NF90 binds the 3'UTR of cyclin E1 mRNA in vitro and in vivo (RIP), stabilizing it; NF90 knockdown decreases cyclin E1 mRNA half-life, delays G1/S transition, and inhibits HCC tumor growth in xenografts; ectopic NF90 expression rescues cyclin E1 mRNA stability.","method":"RIP, 3'UTR binding assay, mRNA half-life measurement (actinomycin D chase), siRNA knockdown, xenograft tumor assay, flow cytometry","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 2 / Strong — RIP plus functional stability rescue plus in vivo xenograft with multiple readouts","pmids":["25399696"],"is_preprint":false},{"year":2015,"finding":"The NF45/NF90 heterodimer associates with pre-60S ribosomal particles via NF90's double-stranded RNA-binding domains (identified by tandem affinity purification); depletion of NF45 and NF90 causes a 60S biogenesis defect, altered nucleolar morphology (spherical nucleoli), and a p53/p21 stress response suppressible by RPL11 depletion.","method":"Tandem affinity purification, density gradient sedimentation, RNAi knockdown, nucleolar morphology imaging, immunoblot, rRNA processing analysis","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — affinity purification identification plus functional depletion with specific ribosome biogenesis readout and domain mapping","pmids":["26240280"],"is_preprint":false},{"year":2015,"finding":"Cold-inducible RBM3 inhibits PERK phosphorylation through cooperation with NF90; NF90 is a novel interacting protein of PERK (identified by affinity purification coupled with mass spectrometry, confirmed by co-IP and proximity ligation assay), and this NF90–PERK interaction is RNA-dependent and required for RBM3-mediated inhibition of the PERK–eIF2α–CHOP ER stress pathway.","method":"Affinity purification with mass spectrometry, co-immunoprecipitation, proximity ligation assay, RBM3 knockout mouse organotypic cultures, siRNA knockdown, PERK phosphorylation assay","journal":"FASEB journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — mass spectrometry identification confirmed by two orthogonal binding assays (Co-IP + PLA) plus functional pathway measurement in KO model","pmids":["26472337"],"is_preprint":false},{"year":2011,"finding":"Alternatively spliced exon 3 of Ilf3/NF90 encodes a 13-amino-acid N-terminal motif that acts as a nucleolar localization signal; four arginines are essential for nucleolar targeting and three histidines stabilize proteins in the nucleolus; the long isoforms (L-Ilf3 and L-NF90) localize to the granular component of the nucleolus and exchange rapidly between nucleoli (FRAP); posttranslational modifications abrogate nucleolar targeting of L-Ilf3.","method":"Subcellular fractionation, confocal microscopy, FRAP, deletion and substitution mutagenesis, recombinant protein expression","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 / Strong — domain mutagenesis plus live-cell FRAP with multiple isoform comparisons establishing mechanistic basis for nucleolar targeting","pmids":["21811582"],"is_preprint":false},{"year":2015,"finding":"The NF45-NF90 and NF45-NF110 complexes act as direct transcriptional coactivators of the c-fos gene in a defined in vitro transcription system; their coactivator activity requires both the upstream enhancer and core promoter regions but not their dsRNA-binding activities; they cooperate with PC4 and Mediator and interact with activators and general transcription machinery; ChIP shows dynamic occupancy on the c-fos gene.","method":"In vitro transcription reconstitution, ChIP, RNAi knockdown, domain mutagenesis (dsRBM-inactive mutant), protein–protein interaction assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro transcription reconstitution plus in vivo ChIP and KD validation; direct transcriptional coactivator function established biochemically","pmids":["26381409"],"is_preprint":false},{"year":2016,"finding":"NF90/NF110 occupies ~9,081 genomic sites in K562 cells, predominantly at active promoters and strong enhancers co-localizing with POLR2A, MYC, and YY1; NF90/NF110 knockdown activates proliferative transcription factors (EGR1, MYC) and suppresses erythroid differentiation (KLF1), establishing NF90/NF110 as a chromatin-associated transcriptional regulator.","method":"ChIP-seq, RNA-seq after shRNA knockdown, comparison with 150 ENCODE ChIP-seq datasets","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome-wide ChIP-seq combined with functional shRNA knockdown and differential gene expression analysis","pmids":["29590119"],"is_preprint":false},{"year":2016,"finding":"NF90-NF45 complex binds pri-miR-7-1 in vitro and inhibits its processing to mature miR-7; NF90/NF45 depletion elevates mature miR-7, reducing EGFR levels and AKT phosphorylation in HCC cells.","method":"miRNA microarray, qRT-PCR, in vitro RNA binding assay, overexpression and siRNA knockdown, immunoblot","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vitro binding plus bidirectional genetic manipulation with pathway-level readout; consistent with earlier NF90-NF45/miRNA biogenesis findings","pmids":["27519414"],"is_preprint":false},{"year":2016,"finding":"NF90 is a novel interacting protein of IAV nonstructural protein NS1 (interaction dependent on NS1 RNA-binding properties); NS1 simultaneously associates with NF90 and PKR but restricts NF90–PKR interaction; NF90 coexpression antagonizes NS1-mediated inhibition of PKR phosphorylation and stress granule formation.","method":"Co-immunoprecipitation, domain mapping, PKR phosphorylation assay, stress granule immunofluorescence, siRNA knockdown","journal":"FEBS letters","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — Co-IP plus functional PKR/SG assays, single lab","pmids":["27423063"],"is_preprint":false},{"year":2016,"finding":"Formation of the NF90–NF45 heterodimer substantially improves NF90's RNA-binding capacity, affinity for both single- and double-stranded RNA, and alters its binding mode; NF45 acts as a conformational scaffold that enables cooperative interplay between NF90's RNA-binding motifs.","method":"Biophysical binding assays (ITC/SPR), biochemical RNA-binding assays with purified recombinant proteins, thermodynamic stability measurements","journal":"The Biochemical journal","confidence":"High","confidence_rationale":"Tier 1 / Moderate — quantitative biophysical characterization of purified proteins, single lab but rigorous in vitro biochemistry","pmids":["28062840"],"is_preprint":false},{"year":2017,"finding":"NF90/NF110 promote circRNA biogenesis in the nucleus by associating with intronic RNA pairs flanking circRNA-forming exons; NF90/NF110 also interact with mature circRNAs in the cytoplasm; upon viral infection, NF90/NF110 translocate to the cytoplasm, reducing circRNA production and becoming available to bind viral mRNAs as part of antiviral response.","method":"Genome-wide siRNA screen, circRNA expression reporter, RIP, subcellular fractionation, viral infection assay, fluorescence imaging","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome-wide functional screen plus mechanistic RIP and localization assays linking nuclear and cytoplasmic functions","pmids":["28625552"],"is_preprint":false},{"year":2017,"finding":"NF90-NF45 acts as a selective RNA chaperone: NF90 alone has RNA annealing and strand displacement activity via a 'matchmaking' plus charge-compensation mechanism; NF45 binding enhances matchmaking efficiency and substantially increases RNA chaperone activity; this activity stimulates the first step of HCV RNA replication in vitro and stabilizes regulatory structures in VEGF mRNA.","method":"In vitro RNA annealing and strand displacement assays, HCV RNA replication in vitro assay, RNA structure probing, biochemical binding assays with purified recombinant NF90 and NF90-NF45","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Strong — mechanistic in vitro biochemical reconstitution of RNA chaperone activity with defined molecular mechanism","pmids":["29040738"],"is_preprint":false},{"year":2018,"finding":"NF90/NF110 controls DICER expression by inhibiting processing of miR-3173 embedded in DICER pre-mRNA; miR-3173 in turn targets NF90, establishing a feedback amplification loop; NF90 overexpression in a nude mouse model reduces ovarian cancer proliferation, tumor size, and metastasis.","method":"miRNA processing assay, NF90 overexpression/knockdown, xenograft mouse model, miR-3173 reporter and functional assays","journal":"Cell research","confidence":"High","confidence_rationale":"Tier 2 / Strong — mechanistic feedback loop established by miRNA processing assay plus in vivo tumor model","pmids":["29563539"],"is_preprint":false},{"year":2018,"finding":"NF90 (ILF3) selectively suppresses translation of wild-type BAFF mRNA (but not the disease-associated BAFF-var mRNA lacking the NF90-binding 3'UTR region) by recruiting miR-15a to the BAFF-WT 3'UTR; this reveals a paradigm in which a UTR polymorphism prevents NF90-mediated miRNA recruitment and raises BAFF protein levels.","method":"RNA immunoprecipitation, reporter assays, miRNA pulldown/recruitment assay, translation measurement","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — RIP plus reporter assay demonstrating miRNA recruitment mechanism; single lab","pmids":["30272251"],"is_preprint":false},{"year":2019,"finding":"ILF3 is a substrate of the E3 ligase SPOP: EGF-MEK-ERK signaling phosphorylates ILF3, which prevents SPOP-mediated poly-ubiquitination and proteasomal degradation of ILF3; stabilized ILF3 then directly binds and stabilizes SGOC pathway gene mRNAs, promoting serine biosynthesis and colorectal cancer growth.","method":"Co-immunoprecipitation, ubiquitination assay, phosphorylation assay, mRNA stability assay, RIP, patient-derived xenograft tumor model","journal":"Cell research","confidence":"High","confidence_rationale":"Tier 2 / Strong — ubiquitination assay plus phosphorylation mapping plus functional in vivo PDX model with multiple orthogonal methods","pmids":["31772275"],"is_preprint":false},{"year":2019,"finding":"SRSF3 regulates alternative splicing of ILF3 by binding RNA sequence motifs to control exon 18 inclusion/exclusion or alternative 3' splice site selection, generating ILF3 isoforms 1 and 2 (pro-proliferative) versus isoforms 5 and 7 (anti-proliferative); isoform-7 induces apoptosis.","method":"Alternative splicing reporter assay, minigene splicing assay, siRNA knockdown of SRSF3, cancer cell proliferation and transformation assays","journal":"RNA","confidence":"High","confidence_rationale":"Tier 2 / Strong — minigene splicing assay with SRSF3 binding characterization plus isoform-specific functional assays","pmids":["30796096"],"is_preprint":false},{"year":2019,"finding":"ILF3 binds the SINEUP lncRNA AS Uchl1 through its RNA-binding motif 2 (RBM2) interacting with the embedded inverted SINE B2 (invSINEB2) element; ILF3 also binds a free Alu monomer sequence and transcriptome-wide SINE sequences; the invSINEB2 element moderately influences AS Uchl1 nuclear localization in an ILF3-dependent manner.","method":"RNA-interacting domainome technology, RNA pulldown, CLIP/eCLIP bioinformatic analysis, domain mutagenesis (RBM2), subcellular localization assay","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — RNA-interacting domainome technology plus domain mutagenesis confirming RBM2 specificity; single lab","pmids":["31570000"],"is_preprint":false},{"year":2019,"finding":"NF90 stabilizes PARP1 mRNA through binding to its 3'UTR (identified by RIP-seq and confirmed by 3'UTR assay); NF90 depletion decreases PARP1 mRNA and protein levels and sensitizes cells to PARP inhibitor Olaparib.","method":"RIP-seq, mRNA stability assay with 3'UTR reporter, siRNA knockdown, drug sensitivity assay","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — RIP-seq plus 3'UTR binding validation plus functional pharmacological consequence; single lab","pmids":["28487110"],"is_preprint":false},{"year":2020,"finding":"ILF3 is an essential host factor for efficient translation of IFNB1 and a subset of interferon-stimulated genes under conditions where cap-dependent translation is compromised; polysome profiling combined with next-generation sequencing showed ILF3 is required to establish both dsRNA-induced transcriptional and translational antiviral programs.","method":"Polysome profiling with next-generation sequencing, siRNA knockdown, IFNB1 reporter and protein assay, dsRNA stimulation","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 2 / Strong — polysome profiling plus RNA-seq provides genome-wide translational evidence; multiple functional readouts","pmids":["31701124"],"is_preprint":false},{"year":2020,"finding":"NF90 modulates the processing of a subset (~38) of highly stable, intronic pri-miRNAs: NF90 associates with the stem region of these pri-miRNAs in a manner largely exclusive of Microprocessor; loss of NF90 increases mature miRNA from 22 bound targets; mutations that destabilize pri-miRNA structure reduce NF90 binding as shown by EMSA.","method":"CLIP-seq/RIP-seq, miRNA microarray, EMSA with RNA structure mutants, NF90 knockdown","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome-wide CLIP combined with EMSA structure-function analysis and functional depletion experiments","pmids":["32427329"],"is_preprint":false},{"year":2020,"finding":"ILF3 stabilizes NEAT1 lncRNA through direct interaction; ATF3 transcriptionally represses ILF3 (ChIP assay shows ATF3 binding to ILF3 promoter); the ATF3/ILF3/NEAT1 axis regulates macrophage M2 polarization.","method":"RIP assay, RNA pulldown, RNA stability assay, ChIP assay, luciferase reporter, siRNA knockdown","journal":"Molecular medicine","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — direct binding (RIP/pulldown) plus transcriptional regulation (ChIP) plus functional polarization assay; single lab","pmids":["38395749"],"is_preprint":false},{"year":2021,"finding":"ILF3 is required for mTORC1-dependent amino acid sensing: ILF3 tethers the GATOR complexes to lysosomes; artificially targeting GATOR2 component WDR24 to lysosomes bypasses the requirement for ILF3; ILF3's role is evolutionarily conserved in human cells, mouse cells, and C. elegans.","method":"Genome-wide CRISPR/Cas9 screen (FACS-based pS6 assay), lysosome-targeting rescue experiment, epistasis analysis with GATOR complexes, cross-species conservation studies","journal":"Nature cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome-wide CRISPR screen plus genetic epistasis (lysosome-targeting rescue) plus evolutionary conservation; published in top journal with rigorous controls","pmids":["37037994"],"is_preprint":false},{"year":2021,"finding":"Tim-3 promotes ubiquitination and proteasomal degradation of NF90 via recruitment of E3 ubiquitin ligase TRIM47, which targets the zinc finger domain of NF90 at Lys297 via K48-linked ubiquitination; Tim-3 inactivation enhances NF90-dependent stress granule formation (increased PKR phosphorylation, eIF2α phosphorylation, G3BP1, and TIA-1) and protects mice from VSV challenge.","method":"Co-immunoprecipitation, ubiquitination assay (K48-linked), site-specific mutagenesis (Lys297), Tim-3 genetic inactivation, VSV infection mouse model, stress granule immunofluorescence","journal":"eLife","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — E3 ligase identification, site-specific ubiquitination mapping (K48/Lys297), domain-mapping, and in vivo mouse validation","pmids":["34110282"],"is_preprint":false},{"year":2021,"finding":"Macrophage ILF3 increases NF-κB activity by hastening p105 mRNA decay and represses anti-inflammatory Nrf2 signaling by facilitating ILF3/eIF4A1 complex-mediated enhancement of Keap1 translational efficiency, promoting abdominal aortic aneurysm progression.","method":"Multi-omics analysis, macrophage-specific transgenic and KO mice, mRNA stability assay (p105), translation efficiency assay (Keap1), co-immunoprecipitation (ILF3/eIF4A1), multiplex immunohistochemistry","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — mechanistic mRNA stability/translation assays with identified complex partner (eIF4A1) plus conditional KO/Tg in vivo validation","pmids":["39179537"],"is_preprint":false},{"year":2021,"finding":"ILF3 regulates ILF3 mRNA stability: ILF3 directly binds ILF3-AS1 and increases its transcript stability; HNRNPA2B1 recognizes m6A sites on ILF3 mRNA and enhances its stability, which in turn promotes AKT3 expression.","method":"m6A site mapping, RIP, mRNA stability assay, siRNA knockdown, immunohistochemistry","journal":"Journal of hematology & oncology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — RIP and stability assay, single lab; m6A reader mechanism inferred","pmids":["33794982"],"is_preprint":false},{"year":2021,"finding":"ILF3 (NF90/NF110) suppresses dendritic cell maturation and innate immune responses through a nuclear localization sequence-dependent mechanism, not through its dsRNA-binding domains; the DZF domain of NF110 isoform is additionally required; ILF3 regulates genes associated with cholesterol homeostasis alongside DC maturation genes.","method":"SiRNA depletion and overexpression in human monocyte-derived DCs, NLS and DZF domain mutagenesis, flow cytometry for DC maturation markers, RNA-seq","journal":"Journal of immunology","confidence":"High","confidence_rationale":"Tier 2 / Strong — domain mutagenesis (NLS, DZF) plus primary human cell functional assays plus RNA-seq; two isoforms compared","pmids":["34031149"],"is_preprint":false},{"year":2021,"finding":"circACTA2 competes with CDK4 mRNA for binding to ILF3 (both shown by oligo pulldown and RIP); Ang II increases circACTA2 expression, facilitating its association with ILF3, thereby reducing ILF3-CDK4 mRNA interaction, decreasing CDK4 mRNA stability and protein levels, and promoting VSMC senescence.","method":"Oligo pulldown, RNA immunoprecipitation (RIP), mRNA stability assay, siRNA knockdown of circACTA2, cellular senescence assay (SA β-gal, p21/CDK4 levels)","journal":"Aging","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — RNA competition mechanism supported by RIP and pulldown, single lab","pmids":["33885378"],"is_preprint":false},{"year":2023,"finding":"circSLC38A1 interacts with ILF3 (RNA pulldown, mass spectrometry, RIP) and stabilizes ILF3 protein by modulating its ubiquitination; the circSLC38A1–ILF3 complex drives transcription of TGF-β2, promoting bladder cancer invasion and metastasis.","method":"RNA pulldown with mass spectrometry, RIP, ubiquitination assay, CUT&Tag-seq, RNA-seq, in vivo mouse metastasis model","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — MS-identified interaction confirmed by RIP and ubiquitination assay plus transcriptional readout; single lab","pmids":["36697384"],"is_preprint":false},{"year":2023,"finding":"PRMT1 mediates arginine methylation of ILF3, stabilizing the ILF3 protein; stabilized ILF3 in turn binds and stabilizes IL-8 mRNA, promoting M2 polarization of macrophages and prostate cancer progression.","method":"Co-immunoprecipitation, GST pulldown, mass spectrometry, RNA pulldown, RIP, in vivo mouse model","journal":"Carcinogenesis","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — multiple binding assays (IP, GST, RIP) identifying arginine methylation as PTM stabilizing ILF3; single lab","pmids":["36394342"],"is_preprint":false},{"year":2024,"finding":"Lefamulin directly binds the Ala99 site of ILF3 protein and interferes with GCN5/CBP-mediated acetylation of Lys100, disrupting ILF3-mediated transcription of MRPL12 and impairing mitochondrial biogenesis/function; this mechanism underlies lefamulin's ability to overcome sorafenib resistance in HCC.","method":"Drug–protein binding assay (direct binding to Ala99), site-directed mutagenesis (Lys100), acetylation assay (GCN5/CBP), ChIP/CUT&Tag for MRPL12 transcription, CDX and hydrodynamic mouse models","journal":"Advanced science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct binding site identified and acetylation mechanism mapped with mutagenesis; in vivo validation; single lab","pmids":["38874478"],"is_preprint":false},{"year":2020,"finding":"USP11 interacts with NF90 and promotes its deubiquitination, thereby stabilizing NF90 protein levels in HCC cells; the effect of USP11 on HCC cell proliferation and metastasis is dependent on NF90.","method":"Mass spectrometry, co-immunoprecipitation, ubiquitination assay, siRNA knockdown epistasis, in vivo mouse model","journal":"American journal of cancer research","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — Co-IP plus ubiquitination assay plus functional epistasis; single lab","pmids":["32509388"],"is_preprint":false},{"year":2024,"finding":"Hyperglycemia/AGEs increase vascular smooth muscle cell NF90 activity; NF90 then stabilizes FBXW7 mRNA, and increased FBXW7 promotes ubiquitination and degradation of AGE receptor 1 (AGER1), causing accumulation of AGEs and atherosclerotic calcification; VSMC-specific NF90/110 knockout in male mice decreases AGEs-induced calcification.","method":"VSMC-specific NF90/110 KO mice, mRNA stability assay (FBXW7), ubiquitination assay (AGER1), multi-omics, immunohistochemistry","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional KO mouse model plus mechanistic mRNA stability and ubiquitination assays; published in top journal","pmids":["38862515"],"is_preprint":false}],"current_model":"ILF3/NF90 is a multifunctional dsRNA-binding protein that operates as: (1) a post-transcriptional regulator—binding AU-rich and structured 3'UTR/5'UTR elements to stabilize or translationally repress target mRNAs (IL-2, MKP-1, cyclin E1, PARP1, VEGF, SGOC genes, CDK4, MRPL12, p21, MyoD); (2) a negative regulator of miRNA biogenesis—competing with the Microprocessor for highly stable intronic pri-miRNAs as an NF90–NF45 complex; (3) a transcriptional coactivator/regulator—occupying active promoters and enhancers (IL-2, IL-13, c-fos, IEGs, BMP2, STAT1) in complex with NF45, Ku proteins, and DNA-PK; (4) an RNA chaperone—facilitating RNA annealing and strand displacement as an NF90–NF45 heterodimer; (5) a nucleo-cytoplasmic shuttle protein whose export is triggered by AKT- or PKCβI-mediated phosphorylation at Ser647; (6) a participant in DNA double-strand break repair via NHEJ through its interaction with DNA-PK; (7) a ribosome biogenesis factor required for 60S subunit maturation via its dsRNA-binding domains; (8) an mTORC1 amino acid sensing mediator that tethers GATOR complexes to lysosomes; and (9) a viral restriction/host factor whose antiviral activity involves PKR activation, stress granule formation, and competition with viral RNA elements, while its stability is regulated by SPOP-mediated ubiquitination (controlled by EGF-ERK phosphorylation), USP11-mediated deubiquitination, TRIM47-mediated K48-ubiquitination at Lys297 (promoted by Tim-3), PRMT1-mediated arginine methylation, and GCN5/CBP-mediated acetylation at Lys100."},"narrative":{"mechanistic_narrative":"ILF3 (NF90/NF110) is a multifunctional double-stranded RNA-binding protein that, in obligate heterodimer with NF45, governs gene expression at the transcriptional, post-transcriptional, and translational levels [PMID:18458058, PMID:22833610, PMID:28062840]. NF45 dimerizes with NF90 through the DZF domain — structurally related to nucleotidyltransferases but catalytically inactive — and this association is reciprocally stabilizing and acts as a conformational scaffold that markedly enhances NF90's affinity for single- and double-stranded RNA [PMID:22833610, PMID:18458058, PMID:28062840]. As a post-transcriptional regulator, NF90 binds AU-rich and structured 3'UTR elements to control the fate of target mRNAs, stabilizing transcripts such as IL-2, MKP-1, cyclin E1, and PARP1 while translationally repressing others through a defined AU-rich motif without altering stability [PMID:12504009, PMID:18490444, PMID:25399696, PMID:19850717]. The complex also acts as an RNA chaperone with annealing and strand-displacement activity [PMID:29040738] and as a negative regulator of miRNA biogenesis, binding the stems of highly stable intronic pri-miRNAs in a manner largely exclusive of the Microprocessor [PMID:19398578, PMID:32427329]. In the nucleus ILF3 functions as a chromatin-associated transcriptional regulator, occupying active promoters and enhancers (IL-2, IL-13, c-fos) in complexes with NF45, Ku proteins, and DNA-PK, and through the latter interaction contributes to non-homologous end-joining repair of DNA double-strand breaks [PMID:17389650, PMID:20051514, PMID:26381409, PMID:29590119, PMID:9442054, PMID:21969602]. ILF3 additionally serves as a 60S ribosome biogenesis factor via its dsRNA-binding domains [PMID:26240280] and as a mediator of mTORC1 amino-acid sensing that tethers GATOR complexes to lysosomes [PMID:37037994]. Its nucleo-cytoplasmic distribution is controlled by phosphorylation at Ser647 by AKT or PKCβI, which triggers export and downstream mRNA stabilization [PMID:18097023, PMID:20870937], and isoform-specific nucleolar targeting is conferred by an alternatively spliced N-terminal signal [PMID:21811582]. In antiviral defense, ILF3 binds viral RNA elements, activates PKR, and promotes stress-granule formation [PMID:24623135, PMID:31701124], while its abundance is set by competing ubiquitination and stabilizing modifications — SPOP-mediated degradation antagonized by EGF-ERK phosphorylation, TRIM47-mediated K48 ubiquitination at Lys297, USP11-mediated deubiquitination, and PRMT1-mediated arginine methylation [PMID:31772275, PMID:34110282, PMID:32509388, PMID:36394342].","teleology":[{"year":1994,"claim":"Established ILF3 as a DNA-binding transcription factor subunit by identifying NF90 and NF45 as the components binding the IL-2 promoter antigen receptor response element, defining its founding role in T-cell gene activation.","evidence":"Protein purification, cDNA cloning, immunofluorescence, and EMSA with pharmacological (cyclosporin A/FK506) validation in Jurkat T cells","pmids":["7519613"],"confidence":"High","gaps":["Direct versus bridged DNA contact not resolved","Mechanism coupling T-cell stimulation to DNA-binding activity unknown"]},{"year":1998,"claim":"Linked NF90/NF45 to DNA double-strand break machinery by showing physical interaction with and phosphorylation by DNA-PK, foreshadowing a DNA repair role.","evidence":"EMSA, in vitro kinase assay, and reciprocal immunoprecipitation with DNA-PKcs/Ku","pmids":["9442054"],"confidence":"High","gaps":["Functional consequence of NF90 phosphorylation by DNA-PK not defined here","Direct repair contribution established only later"]},{"year":2003,"claim":"Revealed how ILF3 is exported from the nucleus by reconstituting an exportin-5/RanGTP/minihelix-RNA quaternary complex, identifying the export receptor and coupling ILF3 trafficking to structured RNA.","evidence":"Gel retardation, GST pulldown, and HeLa nuclear microinjection of VA1 RNA","pmids":["14570900"],"confidence":"High","gaps":["Whether endogenous cellular RNAs use the same export route unclear","Relationship to phosphorylation-triggered export not addressed"]},{"year":2003,"claim":"Demonstrated that ILF3 is hijacked as a host factor by RNA viruses, binding pestivirus 5' and 3' non-translated regions required for replication.","evidence":"RNA-protein binding, viral RNA mutagenesis, and RNAi in BVDV replication assays","pmids":["14592965"],"confidence":"High","gaps":["Whether NF90 promotes or restricts replication context-dependent","Direct enzymatic contribution to replication undefined"]},{"year":2002,"claim":"Defined ILF3 as a post-transcriptional mRNA regulator by showing AU-rich 3'UTR binding stabilizes IL-2 mRNA and that nuclear export is required for this stabilization, connecting localization to function.","evidence":"RNA-protein binding, mRNA half-life, and subcellular fractionation in activated T cells","pmids":["12504009"],"confidence":"High","gaps":["Signal triggering export not yet identified at this stage","Stabilization mechanism (decay factor competition) not resolved"]},{"year":2005,"claim":"Provided in vivo genetic proof of a developmental post-transcriptional role: NF90 knockout mice die perinatally with muscle defects traced to direct binding of MyoD and p21 3'UTRs.","evidence":"NF90-/- mouse knockout, Northwestern blotting, and histology","pmids":["15746098"],"confidence":"High","gaps":["Whether 3'UTR binding stabilizes or translationally controls these mRNAs in vivo not dissected","Tissue-specific requirement versus global essentiality unclear"]},{"year":2008,"claim":"Identified the kinases and phosphosite (Ser647) controlling ILF3 export, showing AKT (CD28) and later PKCβI (PMA) phosphorylation is necessary and sufficient for export and IL-2 mRNA stabilization.","evidence":"In vitro kinase assays, phospho-specific antibodies, Ser647Ala mutagenesis, and export/stability readouts","pmids":["18097023","20870937"],"confidence":"High","gaps":["Phosphatase reversing Ser647 unknown","How phosphorylation alters export receptor engagement not shown"]},{"year":2008,"claim":"Established the NF90-NF45 heterodimer as the functional unit and demonstrated its requirement for cell division, with reciprocal subunit stabilization and multinucleation upon depletion.","evidence":"RNAi depletion of individual subunits, flow cytometry, and time-lapse microscopy","pmids":["18458058"],"confidence":"High","gaps":["Molecular target underlying division defect not identified here","NF90 versus NF110 functional divergence only partly resolved"]},{"year":2009,"claim":"Defined two complementary post-transcriptional mechanisms: translational repression via a discrete AU-rich 3'UTR motif, and negative control of miRNA biogenesis by pri-miRNA binding that excludes the Microprocessor.","evidence":"RIP-seq, polysome profiling, EGFP reporter mutagenesis, and pri/pre/mature miRNA quantification","pmids":["19850717","19398578"],"confidence":"High","gaps":["How NF90 discriminates repression from stabilization targets unclear","Selectivity for specific pri-miRNAs not yet mapped to structure"]},{"year":2011,"claim":"Provided direct functional evidence that NF90-NF45 is required for non-homologous end joining, extending the early DNA-PK interaction into a defined DNA repair role.","evidence":"In vitro NHEJ assay with immunodepletion, γH2AX foci, and ionizing radiation sensitivity","pmids":["21969602"],"confidence":"High","gaps":["Step in NHEJ at which NF90/NF45 acts undefined","Whether RNA binding is required for repair function not tested"]},{"year":2012,"claim":"Provided the structural basis for heterodimer assembly, showing NF45 dimerizes with NF90 through a catalytically dead nucleotidyltransferase-like DZF domain.","evidence":"1.9-Å X-ray crystallography and Co-IP with interface mutants","pmids":["22833610"],"confidence":"High","gaps":["Structure of RNA-bound holocomplex not determined","How dimerization is coupled to RNA-binding enhancement not structurally shown"]},{"year":2015,"claim":"Identified ILF3 as a ribosome biogenesis factor required for 60S maturation, acting through its dsRNA-binding domains on pre-60S particles.","evidence":"Tandem affinity purification, density gradient sedimentation, RNAi, and rRNA processing/nucleolar morphology analysis","pmids":["26240280"],"confidence":"High","gaps":["Specific rRNA or assembly intermediate substrate not defined","Relationship to nucleolar isoform targeting not integrated"]},{"year":2015,"claim":"Established ILF3 as a direct transcriptional coactivator independent of RNA binding, reconstituting c-fos activation in vitro with PC4 and Mediator.","evidence":"In vitro transcription reconstitution, ChIP, and dsRBM-inactive mutant analysis","pmids":["26381409"],"confidence":"High","gaps":["Generality across activator-dependent genes not established","Contacts with specific general transcription factors not mapped"]},{"year":2016,"claim":"Mapped the genome-wide chromatin footprint of NF90/NF110 at active promoters and enhancers and linked occupancy to control of proliferation and differentiation programs.","evidence":"ChIP-seq with ENCODE comparison and RNA-seq after shRNA knockdown in K562","pmids":["29590119"],"confidence":"High","gaps":["Direct versus indirect regulation of individual loci not resolved","Mechanism of chromatin recruitment unknown"]},{"year":2017,"claim":"Defined the biochemical basis of NF90-NF45 RNA chaperone activity (annealing/strand displacement) and connected nuclear circRNA biogenesis to cytoplasmic antiviral redistribution.","evidence":"In vitro annealing/strand-displacement assays and genome-wide siRNA screen with RIP and fractionation","pmids":["29040738","28625552"],"confidence":"High","gaps":["In vivo substrates of chaperone activity beyond test cases unclear","Trigger for infection-induced cytoplasmic translocation not molecularly defined"]},{"year":2020,"claim":"Resolved the antiviral pathway placement of ILF3, showing it activates PKR, promotes stress-granule formation, and is required for translation of IFNB1 and a subset of interferon-stimulated genes.","evidence":"Co-IP/domain mapping, PKR phosphorylation and stress-granule assays, and polysome profiling with NGS","pmids":["24623135","31701124"],"confidence":"High","gaps":["How ILF3 selects ISG transcripts for translation unclear","Balance between restriction and pro-viral host-factor roles not unified"]},{"year":2020,"claim":"Refined the miRNA biogenesis mechanism, defining the subset of highly stable intronic pri-miRNAs whose stem structure is recognized by NF90 to the exclusion of the Microprocessor.","evidence":"CLIP-seq/RIP-seq, miRNA microarray, and EMSA with RNA structure mutants","pmids":["32427329"],"confidence":"High","gaps":["Determinants distinguishing NF90-regulated from unregulated pri-miRNAs incomplete","Whether NF45 is required for stem recognition not addressed"]},{"year":2021,"claim":"Identified ILF3 as a conserved mediator of mTORC1 amino-acid sensing that tethers GATOR complexes to lysosomes, extending ILF3 beyond nucleic-acid biology into nutrient signaling.","evidence":"Genome-wide CRISPR screen, lysosome-targeting rescue epistasis, and cross-species conservation","pmids":["37037994"],"confidence":"High","gaps":["Whether RNA binding contributes to GATOR tethering unknown","Direct ILF3-GATOR contacts not structurally defined"]},{"year":2021,"claim":"Defined the post-translational control of ILF3 abundance, showing EGF-ERK phosphorylation blocks SPOP-mediated degradation and that TRIM47 (downstream of Tim-3) ubiquitinates Lys297 to drive degradation, with stabilized ILF3 controlling metabolic and antiviral outputs.","evidence":"Ubiquitination assays, site-specific (Lys297) mutagenesis, in vivo PDX and VSV mouse models","pmids":["31772275","34110282"],"confidence":"High","gaps":["Crosstalk between competing ubiquitination/deubiquitination/methylation events not integrated","Whether these PTMs alter localization or only abundance unclear"]},{"year":null,"claim":"How ILF3's many functions — transcriptional coactivation, mRNA stability/translation control, miRNA biogenesis, ribosome assembly, DNA repair, and mTORC1 sensing — are partitioned across isoforms, localizations, and PTM states within a single cell remains unresolved.","evidence":"","pmids":[],"confidence":"High","gaps":["No unifying model linking NF90 versus NF110 isoforms to specific functions","Structure of RNA- or DNA-bound functional complexes not determined","Rules governing which targets are stabilized versus repressed unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[4,17,36,38,46]},{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[0,10,19,32]},{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[32,33]},{"term_id":"GO:0045182","term_label":"translation regulator activity","supporting_discovery_ids":[2,17,24]},{"term_id":"GO:0140098","term_label":"catalytic activity, acting on RNA","supporting_discovery_ids":[38,16,46]},{"term_id":"GO:0060089","term_label":"molecular transducer activity","supporting_discovery_ids":[48]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[0,3,7]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[4,27]},{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[29,31]},{"term_id":"GO:0005654","term_label":"nucleoplasm","supporting_discovery_ids":[15,37]},{"term_id":"GO:0005764","term_label":"lysosome","supporting_discovery_ids":[48]}],"pathway":[{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[16,17,37,38,46]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[0,10,19,32,33]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[27,45,49]},{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[1,20]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[13,28]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[48]},{"term_id":"R-HSA-1852241","term_label":"Organelle biogenesis and maintenance","supporting_discovery_ids":[29]}],"complexes":["NF90-NF45 heterodimer","NF110-NF45 heterodimer","DNA-PK (DNA-PKcs–Ku) complex"],"partners":["ILF2","PRKDC","XRCC5","XRCC6","EIF2AK2","EIF4A1","PERK","NS5A"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q12906","full_name":"Interleukin enhancer-binding factor 3","aliases":["Double-stranded RNA-binding protein 76","DRBP76","M-phase phosphoprotein 4","MPP4","Nuclear factor associated with dsRNA","NFAR","Nuclear factor of activated T-cells 90 kDa","NF-AT-90","Translational control protein 80","TCP80"],"length_aa":894,"mass_kda":95.3,"function":"RNA-binding protein that plays an essential role in the biogenesis of circular RNAs (circRNAs) which are produced by back-splicing circularization of pre-mRNAs. Within the nucleus, promotes circRNAs processing by stabilizing the regulatory elements residing in the flanking introns of the circularized exons. Plays thereby a role in the back-splicing of a subset of circRNAs (PubMed:28625552). As a consequence, participates in a wide range of transcriptional and post-transcriptional processes. Binds to poly-U elements and AU-rich elements (AREs) in the 3'-UTR of target mRNAs (PubMed:14731398). Upon viral infection, ILF3 accumulates in the cytoplasm and participates in the innate antiviral response (PubMed:21123651, PubMed:34110282). Mechanistically, ILF3 becomes phosphorylated and activated by the double-stranded RNA-activated protein kinase/PKR which releases ILF3 from cellular mature circRNAs. In turn, unbound ILF3 molecules are able to interact with and thus inhibit viral mRNAs (PubMed:21123651, PubMed:28625552) (Microbial infection) Plays a positive role in HIV-1 virus production by binding to and thereby stabilizing HIV-1 RNA, together with ILF3","subcellular_location":"Nucleus, nucleolus; Cytoplasm; Nucleus","url":"https://www.uniprot.org/uniprotkb/Q12906/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/ILF3","classification":"Common Essential","n_dependent_lines":1178,"n_total_lines":1208,"dependency_fraction":0.9751655629139073},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000129351","cell_line_id":"CID001039","localizations":[{"compartment":"nucleolus_gc","grade":3},{"compartment":"nucleoplasm","grade":3}],"interactors":[{"gene":"ILF2","stoichiometry":10.0},{"gene":"DDX21","stoichiometry":4.0},{"gene":"HNRNPA2B1","stoichiometry":4.0},{"gene":"RPL39P5;RPL39","stoichiometry":4.0},{"gene":"RPS21","stoichiometry":4.0},{"gene":"RPL7A","stoichiometry":4.0},{"gene":"RPS25","stoichiometry":4.0},{"gene":"RPS28","stoichiometry":4.0},{"gene":"RPS16","stoichiometry":4.0},{"gene":"RPL27","stoichiometry":4.0}],"url":"https://opencell.sf.czbiohub.org/target/CID001039","total_profiled":1310},"omim":[{"mim_id":"615176","title":"NPTN INTRONIC TRANSCRIPT 1, NONCODING; 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the DNase I hypersensitive site II in HLA-DR alpha gene was identified as NF90.","date":"1999","source":"Biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/10079079","citation_count":21,"is_preprint":false},{"pmid":"35188449","id":"PMC_35188449","title":"LncRNA LINC00649 promotes the growth and metastasis of triple-negative breast cancer by maintaining the stability of HIF-1α through the NF90/NF45 complex.","date":"2022","source":"Cell cycle (Georgetown, Tex.)","url":"https://pubmed.ncbi.nlm.nih.gov/35188449","citation_count":20,"is_preprint":false},{"pmid":"14654356","id":"PMC_14654356","title":"Cell cycle dependent intracellular distribution of two spliced isoforms of TCP/ILF3 proteins.","date":"2003","source":"Molecular genetics and metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/14654356","citation_count":19,"is_preprint":false},{"pmid":"27423063","id":"PMC_27423063","title":"NF90 is a novel influenza A virus NS1-interacting protein that antagonizes the inhibitory role of NS1 on PKR phosphorylation.","date":"2016","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/27423063","citation_count":19,"is_preprint":false},{"pmid":"34343541","id":"PMC_34343541","title":"ILF3 is responsible for hyperlipidemia-induced arteriosclerotic calcification by mediating BMP2 and STAT1 transcription.","date":"2021","source":"Journal of molecular and cellular cardiology","url":"https://pubmed.ncbi.nlm.nih.gov/34343541","citation_count":18,"is_preprint":false},{"pmid":"31612313","id":"PMC_31612313","title":"LncRNA LINC00470 promotes proliferation through association with NF45/NF90 complex in hepatocellular carcinoma.","date":"2019","source":"Human cell","url":"https://pubmed.ncbi.nlm.nih.gov/31612313","citation_count":18,"is_preprint":false},{"pmid":"38874478","id":"PMC_38874478","title":"Lefamulin Overcomes Acquired Drug Resistance via Regulating Mitochondrial Homeostasis by Targeting ILF3 in Hepatocellular Carcinoma.","date":"2024","source":"Advanced science (Weinheim, Baden-Wurttemberg, Germany)","url":"https://pubmed.ncbi.nlm.nih.gov/38874478","citation_count":17,"is_preprint":false},{"pmid":"30588088","id":"PMC_30588088","title":"The positive feedback loop between ILF3 and lncRNA ILF3-AS1 promotes melanoma proliferation, migration, and invasion.","date":"2018","source":"Cancer management and research","url":"https://pubmed.ncbi.nlm.nih.gov/30588088","citation_count":17,"is_preprint":false},{"pmid":"21811582","id":"PMC_21811582","title":"L-Ilf3 and L-NF90 traffic to the nucleolus granular component: alternatively-spliced exon 3 encodes a nucleolar localization motif.","date":"2011","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/21811582","citation_count":17,"is_preprint":false},{"pmid":"31022259","id":"PMC_31022259","title":"Inducible expression of immediate early genes is regulated through dynamic chromatin association by NF45/ILF2 and NF90/NF110/ILF3.","date":"2019","source":"PloS 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disease","url":"https://pubmed.ncbi.nlm.nih.gov/40140647","citation_count":14,"is_preprint":false},{"pmid":"38395749","id":"PMC_38395749","title":"Role of ATF3 triggering M2 macrophage polarization to protect against the inflammatory injury of sepsis through ILF3/NEAT1 axis.","date":"2024","source":"Molecular medicine (Cambridge, Mass.)","url":"https://pubmed.ncbi.nlm.nih.gov/38395749","citation_count":14,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":50772,"output_tokens":14597,"usd":0.185636,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":27014,"output_tokens":6234,"usd":0.14546,"stage2_stop_reason":"end_turn"},"total_usd":0.331096,"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\": 1994,\n      \"finding\": \"NF90 (90-kDa subunit) and NF45 (45-kDa subunit) together constitute the NF-AT transcription factor complex that binds the antigen receptor response element in the IL-2 promoter; immunofluorescence demonstrated both proteins are nuclear in Jurkat T-cells, and the complex's DNA-binding activity is enhanced by T-cell stimulation but blocked by cyclosporin A or FK506.\",\n      \"method\": \"Protein purification, cDNA cloning, polyhistidine-tag affinity purification, immunofluorescence microscopy, electrophoretic mobility shift assay (EMSA)\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — protein biochemistry, cloning, and functional DNA-binding assays with pharmacological validation, foundational study replicated across many subsequent papers\",\n      \"pmids\": [\"7519613\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"NF90 and NF45 physically interact with DNA-PK (DNA-PKcs and Ku subunits), stabilize the DNA-PKcs–Ku–DNA complex, and are substrates for DNA-PK phosphorylation in vitro; recombinant NF90 promotes DNA-PKcs–Ku–DNA complex formation, and antibodies to NF90 or NF45 immunoprecipitate DNA-PKcs.\",\n      \"method\": \"EMSA, amino-terminal sequence analysis, immunoblotting, in vitro kinase assay, immunoprecipitation\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — multiple orthogonal biochemical methods (EMSA, kinase assay, reciprocal IP) in a single rigorous study; independently reproduced in later work\",\n      \"pmids\": [\"9442054\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"TCP80/NF90 binds the coding region of glucocerebrosidase (GCase) mRNA and inhibits its translation by reducing polysome association, an activity reconstituted in insect Sf9 cells that lack endogenous NF90.\",\n      \"method\": \"Baculovirus/Sf9 insect cell reconstitution, in vitro translation assay, polysome sedimentation, RNA binding assay\",\n      \"journal\": \"Molecular genetics and metabolism\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution of translation inhibition in a heterologous system with purified recombinant protein and functional validation of polysome displacement\",\n      \"pmids\": [\"10873392\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"NFAR-1 (NF90, 90 kDa) and NFAR-2 (NF110, 110 kDa), derived from the same gene by alternative splicing, are phosphorylated by PKR, reciprocally co-immunoprecipitate with PKR, colocalize with PKR in the nucleus, associate with both pre-mRNAs and spliced mRNAs, and interact with RNA-processing factors FUS and SMN via their C-termini.\",\n      \"method\": \"Reciprocal co-immunoprecipitation, subcellular colocalization, in vitro phosphorylation assay, transfection-based gene expression assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP plus multiple orthogonal functional assays in one study\",\n      \"pmids\": [\"11438536\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"NF90 binds the AU-rich element-containing 3'UTR of IL-2 mRNA, slows its degradation, and nuclear export of NF90 to the cytoplasm upon T-cell activation is required for IL-2 mRNA stabilization.\",\n      \"method\": \"RNA–protein binding assay, mRNA half-life measurement, subcellular fractionation, T-cell stimulation assays\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct RNA binding, functional mRNA stability assay, and nucleocytoplasmic translocation linked to function; replicated in multiple subsequent studies\",\n      \"pmids\": [\"12504009\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"ILF3 preferentially binds minihelix-motif RNAs (e.g., adenovirus VA1 RNA) and assembles a quaternary export complex with exportin-5, RanGTP, and the RNA; this complex mediates co-transport of VA1 RNA and ILF3 from nucleus to cytoplasm, identifying exportin-5 as the export receptor for ILF3.\",\n      \"method\": \"Gel retardation assay, GST pulldown, microinjection in HeLa cell nuclei, transfection experiments\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — biochemical reconstitution of the export complex plus live-cell nuclear microinjection validating functional export\",\n      \"pmids\": [\"14570900\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"NFAR proteins (NF90/NF110 group) are recruited by the BVDV replication machinery through specific binding to both 5' and 3' non-translated regions of the viral RNA; mutation of the NF90 interaction sites in the 3'NTR yields replication-deficient RNA, and RNAi depletion of RNA helicase A (an NFAR group member) inhibits replication.\",\n      \"method\": \"RNA–protein binding assay, viral RNA mutagenesis, RNAi knockdown, viral replication assay\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — mutagenesis of viral RNA binding sites coupled with RNAi functional validation\",\n      \"pmids\": [\"14592965\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Cell-cycle-dependent nucleocytoplasmic shuttling of TCP110/ILF3 is directed by a C-terminal nuclear localization signal; TCP80 (NF90) remains stable in the cytoplasm (t½ ~5 days), whereas TCP110/ILF3 translocates between cytoplasm and nucleus during the cell cycle.\",\n      \"method\": \"Subcellular fractionation, 5'RACE cDNA cloning, metabolic turnover assay, immunofluorescence\",\n      \"journal\": \"Molecular genetics and metabolism\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — fractionation plus turnover assays, single lab\",\n      \"pmids\": [\"14654356\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Ilf3 and NF90 directly bind the axonal targeting element (stem-loop in the 3'UTR) of Tau mRNA via their double-stranded RNA-binding motifs; both proteins are detected in neuronal nuclei, cell bodies, and proximal neuritic segments, consistent with a role in Tau mRNA translocation.\",\n      \"method\": \"Northwestern blotting, specific antibodies, immunolocalization\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Northwestern binding assay and immunolocalization, single lab with two orthogonal approaches\",\n      \"pmids\": [\"15364895\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"NF90 knockout mice die perinatally from neuromuscular respiratory failure with skeletal muscle disorganization, decreased myofiber number, and severely reduced expression of MyoD, myogenin, and p21WAF1/CIP1; Northwestern blotting identified NF90 as the principal RNA-binding protein for the p21 and MyoD 3'UTRs in developing muscle, establishing a direct post-transcriptional role in muscle differentiation.\",\n      \"method\": \"Gene knockout (NF90−/− mice), Northwestern blotting, histology, immunohistochemistry\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — loss-of-function in vivo KO with defined molecular mechanism (direct 3'UTR binding); unique in vivo genetic evidence\",\n      \"pmids\": [\"15746098\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"NF90 and NF45 together with Ku80 and Ku70 bind specifically to the IL-2 ARRE/NF-AT DNA sequence in vivo; T-cell activation induces binding of Ku80 and NF90 but decreases Ku70 binding at the IL-2 promoter, and these dynamic changes are blocked by cyclosporin A and triptolide.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP), EMSA with monoclonal antibody inhibition, purification from activated Jurkat nuclei\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo ChIP plus EMSA with inhibitory antibodies, multiple cell types tested\",\n      \"pmids\": [\"17389650\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"NF90ctv (C-terminal variant of NF90) binds the HIV-1 TAR RNA with high affinity, competes with Tat for TAR binding in vitro, inhibits Tat-transactivation of the HIV-1 LTR in vivo, and is associated with histone H3K4/K9 methylation changes consistent with transcriptional repression.\",\n      \"method\": \"TAR RNA affinity fractionation, Northwestern blotting, EMSA, luciferase transactivation assay, chromatin histone methylation analysis\",\n      \"journal\": \"Retrovirology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — multiple assays (EMSA, cell-based transactivation, histone mark analysis) in a single lab\",\n      \"pmids\": [\"17565699\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"AKT phosphorylates NF90 at Ser647 in response to CD28 costimulation; this phosphorylation is necessary and sufficient for nuclear export of NF90 and subsequent IL-2 mRNA stabilization, as a Ser647→Ala mutation abolishes both nuclear export and mRNA stabilization.\",\n      \"method\": \"In vitro kinase assay, phospho-specific antibody, site-directed mutagenesis, nuclear export assay, mRNA stability assay, T-cell stimulation\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — in vitro kinase assay combined with mutagenesis and functional mRNA/export readouts\",\n      \"pmids\": [\"18097023\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"NF90 and NF45 form heterodimeric core complexes (NF90-NF45 and NF110-NF45); depletion of NF45 dramatically decreases NF90/NF110 protein levels (posttranscriptional destabilization), and depletion of NF90 (but not NF110) greatly reduces NF45; depletion of the NF90-NF45 complex retards cell growth by inhibiting DNA synthesis and causes giant multinucleated cells, revealing a specific role in cell division.\",\n      \"method\": \"RNA interference (RNAi) knockdown, flow cytometry, immunoblot, cell growth assay, time-lapse microscopy\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — systematic RNAi depletion of individual subunits with multiple cellular readouts; clearly distinguishes NF90 from NF110 function\",\n      \"pmids\": [\"18458058\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"NF90 and HuR bind the AU-rich 3'UTR of MKP-1 mRNA; H2O2 treatment increases association of MKP-1 mRNA with HuR and NF90 and decreases association with translational repressors TIAR and TIA-1; silencing NF90 or HuR diminishes H2O2-stimulated MKP-1 mRNA stability, and HuR silencing also reduces MKP-1 translation.\",\n      \"method\": \"Ribonucleoprotein immunoprecipitation (RIP), biotinylated RNA pulldown, mRNA stability assay, siRNA knockdown\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — RIP plus RNA pulldown plus functional mRNA stability measurement with siRNA, multiple orthogonal methods\",\n      \"pmids\": [\"18490444\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"NFAR-1 and NFAR-2 retain cellular transcripts in intranuclear foci and regulate mRNA export; loss of NFAR function (embryonic lethal) increases protein synthesis rates; NFAR-depleted murine fibroblasts are dramatically susceptible to VSV replication, demonstrating an innate immune translational surveillance function.\",\n      \"method\": \"NFAR knockout (embryonic lethal), RNA export assay, polysome/translation rate measurement, VSV infection assay in NFAR-depleted cells\",\n      \"journal\": \"Proceedings of the National Academy of Sciences\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo KO (embryonic lethality) plus multiple functional assays (export, translation, viral replication) in depleted cells\",\n      \"pmids\": [\"18337511\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"The NF90-NF45 complex functions as a negative regulator of miRNA biogenesis: overexpression inhibits pri-miRNA processing to pre-miRNA, causing pri-miRNA accumulation; the complex binds pri-miRNAs (e.g., pri-let-7a) but does not interact with the Microprocessor, suggesting it blocks Microprocessor access; depletion of NF90 reduces pri-let-7a and increases mature let-7a.\",\n      \"method\": \"Overexpression and RNAi knockdown, pri-miRNA/pre-miRNA/mature miRNA quantification, RNA polymerase II inhibitor experiments, RNA immunoprecipitation\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — bidirectional genetic manipulation (OE and KD) plus negative result (no Microprocessor interaction) with multiple miRNA substrates\",\n      \"pmids\": [\"19398578\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"NF90 binds an AU-rich 25–30 nucleotide RNA motif in the 3'UTR of target mRNAs and represses their translation without significantly affecting mRNA stability; insertion of this NF90 motif into an EGFP reporter 3'UTR specifically confers NF90-dependent translational repression as shown by polysome gradient analysis.\",\n      \"method\": \"RIP-seq, biotinylated transcript pulldown, polysome gradient analysis, EGFP reporter assay, siRNA knockdown\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — genome-wide RIP combined with reporter mutagenesis and polysome profiling; multiple orthogonal methods\",\n      \"pmids\": [\"19850717\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"PKCβI phosphorylates NF90 at Ser647 in response to PMA stimulation, triggering nuclear export of NF90 and stabilization of IL-2 mRNA; Ser647→Ala mutation abolishes PMA-induced nuclear export and IL-2 mRNA stabilization.\",\n      \"method\": \"In vitro kinase assay with PKCβI, phospho-specific antibody, site-directed mutagenesis, nuclear export assay, mRNA stability assay\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — in vitro kinase assay plus mutagenesis plus functional cellular readouts; complements the earlier AKT study\",\n      \"pmids\": [\"20870937\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"NF45 and NF90 bind the HS4 element of the IL-13 promoter in T cells and are required for HS4-dependent IL-13 transcriptional activation; the CTGTT binding motif within HS4-3' is critical, and HS4 activity is abrogated in NF45+/− primary Th2 cells and reduced in NF90+/− cells.\",\n      \"method\": \"DNA affinity chromatography coupled with tandem mass spectrometry, ChIP, EMSA, reporter transfection in primary Th2 cells from NF45+/− and NF90+/− mice\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (mass spec, ChIP, EMSA, haploinsufficiency in primary cells) in one study\",\n      \"pmids\": [\"20051514\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"The NF90-NF45 complex is required for DNA double-strand break repair by nonhomologous end joining (NHEJ): immunodepletion of NF90/NF45 reduces in vitro NHEJ activity comparably to DNA-PKcs depletion; NF90/NF45-depleted cells accumulate γ-H2A.X foci and are hypersensitive to ionizing radiation.\",\n      \"method\": \"In vitro NHEJ assay with immunodepletion, γ-H2A.X foci quantification, ionizing radiation sensitivity assay, time-lapse microscopy, RNAi knockdown\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — in vitro reconstitution (NHEJ assay with immunodepletion) plus multiple in vivo readouts (γH2AX, IR sensitivity)\",\n      \"pmids\": [\"21969602\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"NF90 binds the dengue virus 3' stem-loop RNA via affinity chromatography; NF90 depletion by siRNA reduces dengue RNA levels and infectious virus production by 50–70%; cytoplasmic relocalization of NF90 occurs in dengue-infected cells.\",\n      \"method\": \"RNA affinity column chromatography, siRNA knockdown, viral RNA quantification, infectious virus titration, immunofluorescence\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — RNA affinity binding plus functional siRNA depletion with two viral replication readouts\",\n      \"pmids\": [\"21386893\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"NF45 dimerizes with NF90 (and related proteins SPNR and Zfr) through the DZF domain, which has structural similarity to the template-free nucleotidyltransferase family but lacks catalytic residues; crystal structure at 1.9-Å resolution reveals the dimerization interface, confirmed by co-immunoprecipitation with site-specific mutants.\",\n      \"method\": \"X-ray crystallography (1.9 Å), co-immunoprecipitation with site-directed mutants\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure combined with mutagenesis-validated functional interactions\",\n      \"pmids\": [\"22833610\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"YM155 directly binds ILF3 and disrupts the ILF3/p54(nrb) complex, causing different subcellular localization of ILF3 and p54(nrb), which results in suppression of survivin promoter activity.\",\n      \"method\": \"Small-molecule binding assay, co-immunoprecipitation, subcellular localization analysis, survivin promoter activity assay\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP and localization assays, single lab; small-molecule target engagement inferred rather than directly demonstrated by structural methods\",\n      \"pmids\": [\"22842455\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"NF90 coordinately represses translation (not stability) of multiple SASP factor mRNAs (MCP-1, GROα, IL-6, IL-8) in proliferating fibroblasts; NF90 levels decline in senescent cells, allowing SASP factor expression to rise.\",\n      \"method\": \"RIP, siRNA knockdown, ELISA for secreted proteins, mRNA stability assay, immunoblot\",\n      \"journal\": \"Aging\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — RIP combined with KD and functional protein/mRNA measurement; single lab\",\n      \"pmids\": [\"23117626\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Depletion of NF90/NF45 in HPV-transformed cervical carcinoma cells restores p53 protein (post-transcriptionally, not at mRNA level) and p21 (via p53-dependent transcription); NF90 knockdown attenuates HPV E6 RNA levels and inhibits transcription from the HPV early promoter, revealing that NF90/NF45 supports viral E6 expression.\",\n      \"method\": \"siRNA knockdown, RT-PCR, immunoblot, PARP cleavage assay, camptothecin sensitivity assay\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — systematic knockdown with multiple molecular and cellular readouts distinguishing transcriptional from post-transcriptional effects\",\n      \"pmids\": [\"23208500\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"NF90 binds the 5'-terminal double-stranded region of the HCV genome; siRNA depletion of NF90 significantly impairs HCV RNA replication and reduces infectious virus yields; NF90 co-immunoprecipitates with NS5A in an RNA-dependent manner and co-fractionates with detergent-resistant membranes containing viral replication complexes.\",\n      \"method\": \"Biotinylated RNA pulldown with mass spectrometry, siRNA knockdown, viral RNA quantification, co-immunoprecipitation, detergent-resistant membrane fractionation\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — RNA affinity MS plus functional depletion plus RIP in infected cells with multiple orthogonal methods\",\n      \"pmids\": [\"24719423\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"NF90 interacts with PKR through its C-terminal domain independently of NF90 RNA-binding properties; NF90 knockdown reduces PKR phosphorylation in response to dsRNA/influenza infection; NF90 is a component of stress granules, and NF90 depletion reduces dsRNA-induced stress granule formation; an NS1 mutant influenza virus specifically attenuated in PKR inhibition replicates indistinguishably from WT in NF90-depleted cells, placing NF90 in the PKR–stress granule antiviral pathway.\",\n      \"method\": \"Co-immunoprecipitation, C-terminal domain mapping, siRNA knockdown, PKR phosphorylation assay, stress granule immunofluorescence, viral replication assay with NS1 mutant\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — protein interaction mapping plus genetic epistasis using NS1 mutant virus, multiple orthogonal methods\",\n      \"pmids\": [\"24623135\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"NF90 binds the 3'UTR of cyclin E1 mRNA in vitro and in vivo (RIP), stabilizing it; NF90 knockdown decreases cyclin E1 mRNA half-life, delays G1/S transition, and inhibits HCC tumor growth in xenografts; ectopic NF90 expression rescues cyclin E1 mRNA stability.\",\n      \"method\": \"RIP, 3'UTR binding assay, mRNA half-life measurement (actinomycin D chase), siRNA knockdown, xenograft tumor assay, flow cytometry\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — RIP plus functional stability rescue plus in vivo xenograft with multiple readouts\",\n      \"pmids\": [\"25399696\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"The NF45/NF90 heterodimer associates with pre-60S ribosomal particles via NF90's double-stranded RNA-binding domains (identified by tandem affinity purification); depletion of NF45 and NF90 causes a 60S biogenesis defect, altered nucleolar morphology (spherical nucleoli), and a p53/p21 stress response suppressible by RPL11 depletion.\",\n      \"method\": \"Tandem affinity purification, density gradient sedimentation, RNAi knockdown, nucleolar morphology imaging, immunoblot, rRNA processing analysis\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — affinity purification identification plus functional depletion with specific ribosome biogenesis readout and domain mapping\",\n      \"pmids\": [\"26240280\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Cold-inducible RBM3 inhibits PERK phosphorylation through cooperation with NF90; NF90 is a novel interacting protein of PERK (identified by affinity purification coupled with mass spectrometry, confirmed by co-IP and proximity ligation assay), and this NF90–PERK interaction is RNA-dependent and required for RBM3-mediated inhibition of the PERK–eIF2α–CHOP ER stress pathway.\",\n      \"method\": \"Affinity purification with mass spectrometry, co-immunoprecipitation, proximity ligation assay, RBM3 knockout mouse organotypic cultures, siRNA knockdown, PERK phosphorylation assay\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — mass spectrometry identification confirmed by two orthogonal binding assays (Co-IP + PLA) plus functional pathway measurement in KO model\",\n      \"pmids\": [\"26472337\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Alternatively spliced exon 3 of Ilf3/NF90 encodes a 13-amino-acid N-terminal motif that acts as a nucleolar localization signal; four arginines are essential for nucleolar targeting and three histidines stabilize proteins in the nucleolus; the long isoforms (L-Ilf3 and L-NF90) localize to the granular component of the nucleolus and exchange rapidly between nucleoli (FRAP); posttranslational modifications abrogate nucleolar targeting of L-Ilf3.\",\n      \"method\": \"Subcellular fractionation, confocal microscopy, FRAP, deletion and substitution mutagenesis, recombinant protein expression\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — domain mutagenesis plus live-cell FRAP with multiple isoform comparisons establishing mechanistic basis for nucleolar targeting\",\n      \"pmids\": [\"21811582\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"The NF45-NF90 and NF45-NF110 complexes act as direct transcriptional coactivators of the c-fos gene in a defined in vitro transcription system; their coactivator activity requires both the upstream enhancer and core promoter regions but not their dsRNA-binding activities; they cooperate with PC4 and Mediator and interact with activators and general transcription machinery; ChIP shows dynamic occupancy on the c-fos gene.\",\n      \"method\": \"In vitro transcription reconstitution, ChIP, RNAi knockdown, domain mutagenesis (dsRBM-inactive mutant), protein–protein interaction assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro transcription reconstitution plus in vivo ChIP and KD validation; direct transcriptional coactivator function established biochemically\",\n      \"pmids\": [\"26381409\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"NF90/NF110 occupies ~9,081 genomic sites in K562 cells, predominantly at active promoters and strong enhancers co-localizing with POLR2A, MYC, and YY1; NF90/NF110 knockdown activates proliferative transcription factors (EGR1, MYC) and suppresses erythroid differentiation (KLF1), establishing NF90/NF110 as a chromatin-associated transcriptional regulator.\",\n      \"method\": \"ChIP-seq, RNA-seq after shRNA knockdown, comparison with 150 ENCODE ChIP-seq datasets\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome-wide ChIP-seq combined with functional shRNA knockdown and differential gene expression analysis\",\n      \"pmids\": [\"29590119\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"NF90-NF45 complex binds pri-miR-7-1 in vitro and inhibits its processing to mature miR-7; NF90/NF45 depletion elevates mature miR-7, reducing EGFR levels and AKT phosphorylation in HCC cells.\",\n      \"method\": \"miRNA microarray, qRT-PCR, in vitro RNA binding assay, overexpression and siRNA knockdown, immunoblot\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vitro binding plus bidirectional genetic manipulation with pathway-level readout; consistent with earlier NF90-NF45/miRNA biogenesis findings\",\n      \"pmids\": [\"27519414\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"NF90 is a novel interacting protein of IAV nonstructural protein NS1 (interaction dependent on NS1 RNA-binding properties); NS1 simultaneously associates with NF90 and PKR but restricts NF90–PKR interaction; NF90 coexpression antagonizes NS1-mediated inhibition of PKR phosphorylation and stress granule formation.\",\n      \"method\": \"Co-immunoprecipitation, domain mapping, PKR phosphorylation assay, stress granule immunofluorescence, siRNA knockdown\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — Co-IP plus functional PKR/SG assays, single lab\",\n      \"pmids\": [\"27423063\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Formation of the NF90–NF45 heterodimer substantially improves NF90's RNA-binding capacity, affinity for both single- and double-stranded RNA, and alters its binding mode; NF45 acts as a conformational scaffold that enables cooperative interplay between NF90's RNA-binding motifs.\",\n      \"method\": \"Biophysical binding assays (ITC/SPR), biochemical RNA-binding assays with purified recombinant proteins, thermodynamic stability measurements\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — quantitative biophysical characterization of purified proteins, single lab but rigorous in vitro biochemistry\",\n      \"pmids\": [\"28062840\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"NF90/NF110 promote circRNA biogenesis in the nucleus by associating with intronic RNA pairs flanking circRNA-forming exons; NF90/NF110 also interact with mature circRNAs in the cytoplasm; upon viral infection, NF90/NF110 translocate to the cytoplasm, reducing circRNA production and becoming available to bind viral mRNAs as part of antiviral response.\",\n      \"method\": \"Genome-wide siRNA screen, circRNA expression reporter, RIP, subcellular fractionation, viral infection assay, fluorescence imaging\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome-wide functional screen plus mechanistic RIP and localization assays linking nuclear and cytoplasmic functions\",\n      \"pmids\": [\"28625552\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"NF90-NF45 acts as a selective RNA chaperone: NF90 alone has RNA annealing and strand displacement activity via a 'matchmaking' plus charge-compensation mechanism; NF45 binding enhances matchmaking efficiency and substantially increases RNA chaperone activity; this activity stimulates the first step of HCV RNA replication in vitro and stabilizes regulatory structures in VEGF mRNA.\",\n      \"method\": \"In vitro RNA annealing and strand displacement assays, HCV RNA replication in vitro assay, RNA structure probing, biochemical binding assays with purified recombinant NF90 and NF90-NF45\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — mechanistic in vitro biochemical reconstitution of RNA chaperone activity with defined molecular mechanism\",\n      \"pmids\": [\"29040738\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"NF90/NF110 controls DICER expression by inhibiting processing of miR-3173 embedded in DICER pre-mRNA; miR-3173 in turn targets NF90, establishing a feedback amplification loop; NF90 overexpression in a nude mouse model reduces ovarian cancer proliferation, tumor size, and metastasis.\",\n      \"method\": \"miRNA processing assay, NF90 overexpression/knockdown, xenograft mouse model, miR-3173 reporter and functional assays\",\n      \"journal\": \"Cell research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — mechanistic feedback loop established by miRNA processing assay plus in vivo tumor model\",\n      \"pmids\": [\"29563539\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"NF90 (ILF3) selectively suppresses translation of wild-type BAFF mRNA (but not the disease-associated BAFF-var mRNA lacking the NF90-binding 3'UTR region) by recruiting miR-15a to the BAFF-WT 3'UTR; this reveals a paradigm in which a UTR polymorphism prevents NF90-mediated miRNA recruitment and raises BAFF protein levels.\",\n      \"method\": \"RNA immunoprecipitation, reporter assays, miRNA pulldown/recruitment assay, translation measurement\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — RIP plus reporter assay demonstrating miRNA recruitment mechanism; single lab\",\n      \"pmids\": [\"30272251\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"ILF3 is a substrate of the E3 ligase SPOP: EGF-MEK-ERK signaling phosphorylates ILF3, which prevents SPOP-mediated poly-ubiquitination and proteasomal degradation of ILF3; stabilized ILF3 then directly binds and stabilizes SGOC pathway gene mRNAs, promoting serine biosynthesis and colorectal cancer growth.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, phosphorylation assay, mRNA stability assay, RIP, patient-derived xenograft tumor model\",\n      \"journal\": \"Cell research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — ubiquitination assay plus phosphorylation mapping plus functional in vivo PDX model with multiple orthogonal methods\",\n      \"pmids\": [\"31772275\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"SRSF3 regulates alternative splicing of ILF3 by binding RNA sequence motifs to control exon 18 inclusion/exclusion or alternative 3' splice site selection, generating ILF3 isoforms 1 and 2 (pro-proliferative) versus isoforms 5 and 7 (anti-proliferative); isoform-7 induces apoptosis.\",\n      \"method\": \"Alternative splicing reporter assay, minigene splicing assay, siRNA knockdown of SRSF3, cancer cell proliferation and transformation assays\",\n      \"journal\": \"RNA\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — minigene splicing assay with SRSF3 binding characterization plus isoform-specific functional assays\",\n      \"pmids\": [\"30796096\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"ILF3 binds the SINEUP lncRNA AS Uchl1 through its RNA-binding motif 2 (RBM2) interacting with the embedded inverted SINE B2 (invSINEB2) element; ILF3 also binds a free Alu monomer sequence and transcriptome-wide SINE sequences; the invSINEB2 element moderately influences AS Uchl1 nuclear localization in an ILF3-dependent manner.\",\n      \"method\": \"RNA-interacting domainome technology, RNA pulldown, CLIP/eCLIP bioinformatic analysis, domain mutagenesis (RBM2), subcellular localization assay\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — RNA-interacting domainome technology plus domain mutagenesis confirming RBM2 specificity; single lab\",\n      \"pmids\": [\"31570000\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"NF90 stabilizes PARP1 mRNA through binding to its 3'UTR (identified by RIP-seq and confirmed by 3'UTR assay); NF90 depletion decreases PARP1 mRNA and protein levels and sensitizes cells to PARP inhibitor Olaparib.\",\n      \"method\": \"RIP-seq, mRNA stability assay with 3'UTR reporter, siRNA knockdown, drug sensitivity assay\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — RIP-seq plus 3'UTR binding validation plus functional pharmacological consequence; single lab\",\n      \"pmids\": [\"28487110\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"ILF3 is an essential host factor for efficient translation of IFNB1 and a subset of interferon-stimulated genes under conditions where cap-dependent translation is compromised; polysome profiling combined with next-generation sequencing showed ILF3 is required to establish both dsRNA-induced transcriptional and translational antiviral programs.\",\n      \"method\": \"Polysome profiling with next-generation sequencing, siRNA knockdown, IFNB1 reporter and protein assay, dsRNA stimulation\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — polysome profiling plus RNA-seq provides genome-wide translational evidence; multiple functional readouts\",\n      \"pmids\": [\"31701124\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"NF90 modulates the processing of a subset (~38) of highly stable, intronic pri-miRNAs: NF90 associates with the stem region of these pri-miRNAs in a manner largely exclusive of Microprocessor; loss of NF90 increases mature miRNA from 22 bound targets; mutations that destabilize pri-miRNA structure reduce NF90 binding as shown by EMSA.\",\n      \"method\": \"CLIP-seq/RIP-seq, miRNA microarray, EMSA with RNA structure mutants, NF90 knockdown\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome-wide CLIP combined with EMSA structure-function analysis and functional depletion experiments\",\n      \"pmids\": [\"32427329\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"ILF3 stabilizes NEAT1 lncRNA through direct interaction; ATF3 transcriptionally represses ILF3 (ChIP assay shows ATF3 binding to ILF3 promoter); the ATF3/ILF3/NEAT1 axis regulates macrophage M2 polarization.\",\n      \"method\": \"RIP assay, RNA pulldown, RNA stability assay, ChIP assay, luciferase reporter, siRNA knockdown\",\n      \"journal\": \"Molecular medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — direct binding (RIP/pulldown) plus transcriptional regulation (ChIP) plus functional polarization assay; single lab\",\n      \"pmids\": [\"38395749\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ILF3 is required for mTORC1-dependent amino acid sensing: ILF3 tethers the GATOR complexes to lysosomes; artificially targeting GATOR2 component WDR24 to lysosomes bypasses the requirement for ILF3; ILF3's role is evolutionarily conserved in human cells, mouse cells, and C. elegans.\",\n      \"method\": \"Genome-wide CRISPR/Cas9 screen (FACS-based pS6 assay), lysosome-targeting rescue experiment, epistasis analysis with GATOR complexes, cross-species conservation studies\",\n      \"journal\": \"Nature cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome-wide CRISPR screen plus genetic epistasis (lysosome-targeting rescue) plus evolutionary conservation; published in top journal with rigorous controls\",\n      \"pmids\": [\"37037994\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Tim-3 promotes ubiquitination and proteasomal degradation of NF90 via recruitment of E3 ubiquitin ligase TRIM47, which targets the zinc finger domain of NF90 at Lys297 via K48-linked ubiquitination; Tim-3 inactivation enhances NF90-dependent stress granule formation (increased PKR phosphorylation, eIF2α phosphorylation, G3BP1, and TIA-1) and protects mice from VSV challenge.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay (K48-linked), site-specific mutagenesis (Lys297), Tim-3 genetic inactivation, VSV infection mouse model, stress granule immunofluorescence\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — E3 ligase identification, site-specific ubiquitination mapping (K48/Lys297), domain-mapping, and in vivo mouse validation\",\n      \"pmids\": [\"34110282\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Macrophage ILF3 increases NF-κB activity by hastening p105 mRNA decay and represses anti-inflammatory Nrf2 signaling by facilitating ILF3/eIF4A1 complex-mediated enhancement of Keap1 translational efficiency, promoting abdominal aortic aneurysm progression.\",\n      \"method\": \"Multi-omics analysis, macrophage-specific transgenic and KO mice, mRNA stability assay (p105), translation efficiency assay (Keap1), co-immunoprecipitation (ILF3/eIF4A1), multiplex immunohistochemistry\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — mechanistic mRNA stability/translation assays with identified complex partner (eIF4A1) plus conditional KO/Tg in vivo validation\",\n      \"pmids\": [\"39179537\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ILF3 regulates ILF3 mRNA stability: ILF3 directly binds ILF3-AS1 and increases its transcript stability; HNRNPA2B1 recognizes m6A sites on ILF3 mRNA and enhances its stability, which in turn promotes AKT3 expression.\",\n      \"method\": \"m6A site mapping, RIP, mRNA stability assay, siRNA knockdown, immunohistochemistry\",\n      \"journal\": \"Journal of hematology & oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — RIP and stability assay, single lab; m6A reader mechanism inferred\",\n      \"pmids\": [\"33794982\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ILF3 (NF90/NF110) suppresses dendritic cell maturation and innate immune responses through a nuclear localization sequence-dependent mechanism, not through its dsRNA-binding domains; the DZF domain of NF110 isoform is additionally required; ILF3 regulates genes associated with cholesterol homeostasis alongside DC maturation genes.\",\n      \"method\": \"SiRNA depletion and overexpression in human monocyte-derived DCs, NLS and DZF domain mutagenesis, flow cytometry for DC maturation markers, RNA-seq\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — domain mutagenesis (NLS, DZF) plus primary human cell functional assays plus RNA-seq; two isoforms compared\",\n      \"pmids\": [\"34031149\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"circACTA2 competes with CDK4 mRNA for binding to ILF3 (both shown by oligo pulldown and RIP); Ang II increases circACTA2 expression, facilitating its association with ILF3, thereby reducing ILF3-CDK4 mRNA interaction, decreasing CDK4 mRNA stability and protein levels, and promoting VSMC senescence.\",\n      \"method\": \"Oligo pulldown, RNA immunoprecipitation (RIP), mRNA stability assay, siRNA knockdown of circACTA2, cellular senescence assay (SA β-gal, p21/CDK4 levels)\",\n      \"journal\": \"Aging\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — RNA competition mechanism supported by RIP and pulldown, single lab\",\n      \"pmids\": [\"33885378\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"circSLC38A1 interacts with ILF3 (RNA pulldown, mass spectrometry, RIP) and stabilizes ILF3 protein by modulating its ubiquitination; the circSLC38A1–ILF3 complex drives transcription of TGF-β2, promoting bladder cancer invasion and metastasis.\",\n      \"method\": \"RNA pulldown with mass spectrometry, RIP, ubiquitination assay, CUT&Tag-seq, RNA-seq, in vivo mouse metastasis model\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — MS-identified interaction confirmed by RIP and ubiquitination assay plus transcriptional readout; single lab\",\n      \"pmids\": [\"36697384\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"PRMT1 mediates arginine methylation of ILF3, stabilizing the ILF3 protein; stabilized ILF3 in turn binds and stabilizes IL-8 mRNA, promoting M2 polarization of macrophages and prostate cancer progression.\",\n      \"method\": \"Co-immunoprecipitation, GST pulldown, mass spectrometry, RNA pulldown, RIP, in vivo mouse model\",\n      \"journal\": \"Carcinogenesis\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — multiple binding assays (IP, GST, RIP) identifying arginine methylation as PTM stabilizing ILF3; single lab\",\n      \"pmids\": [\"36394342\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Lefamulin directly binds the Ala99 site of ILF3 protein and interferes with GCN5/CBP-mediated acetylation of Lys100, disrupting ILF3-mediated transcription of MRPL12 and impairing mitochondrial biogenesis/function; this mechanism underlies lefamulin's ability to overcome sorafenib resistance in HCC.\",\n      \"method\": \"Drug–protein binding assay (direct binding to Ala99), site-directed mutagenesis (Lys100), acetylation assay (GCN5/CBP), ChIP/CUT&Tag for MRPL12 transcription, CDX and hydrodynamic mouse models\",\n      \"journal\": \"Advanced science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding site identified and acetylation mechanism mapped with mutagenesis; in vivo validation; single lab\",\n      \"pmids\": [\"38874478\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"USP11 interacts with NF90 and promotes its deubiquitination, thereby stabilizing NF90 protein levels in HCC cells; the effect of USP11 on HCC cell proliferation and metastasis is dependent on NF90.\",\n      \"method\": \"Mass spectrometry, co-immunoprecipitation, ubiquitination assay, siRNA knockdown epistasis, in vivo mouse model\",\n      \"journal\": \"American journal of cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — Co-IP plus ubiquitination assay plus functional epistasis; single lab\",\n      \"pmids\": [\"32509388\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Hyperglycemia/AGEs increase vascular smooth muscle cell NF90 activity; NF90 then stabilizes FBXW7 mRNA, and increased FBXW7 promotes ubiquitination and degradation of AGE receptor 1 (AGER1), causing accumulation of AGEs and atherosclerotic calcification; VSMC-specific NF90/110 knockout in male mice decreases AGEs-induced calcification.\",\n      \"method\": \"VSMC-specific NF90/110 KO mice, mRNA stability assay (FBXW7), ubiquitination assay (AGER1), multi-omics, immunohistochemistry\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional KO mouse model plus mechanistic mRNA stability and ubiquitination assays; published in top journal\",\n      \"pmids\": [\"38862515\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ILF3/NF90 is a multifunctional dsRNA-binding protein that operates as: (1) a post-transcriptional regulator—binding AU-rich and structured 3'UTR/5'UTR elements to stabilize or translationally repress target mRNAs (IL-2, MKP-1, cyclin E1, PARP1, VEGF, SGOC genes, CDK4, MRPL12, p21, MyoD); (2) a negative regulator of miRNA biogenesis—competing with the Microprocessor for highly stable intronic pri-miRNAs as an NF90–NF45 complex; (3) a transcriptional coactivator/regulator—occupying active promoters and enhancers (IL-2, IL-13, c-fos, IEGs, BMP2, STAT1) in complex with NF45, Ku proteins, and DNA-PK; (4) an RNA chaperone—facilitating RNA annealing and strand displacement as an NF90–NF45 heterodimer; (5) a nucleo-cytoplasmic shuttle protein whose export is triggered by AKT- or PKCβI-mediated phosphorylation at Ser647; (6) a participant in DNA double-strand break repair via NHEJ through its interaction with DNA-PK; (7) a ribosome biogenesis factor required for 60S subunit maturation via its dsRNA-binding domains; (8) an mTORC1 amino acid sensing mediator that tethers GATOR complexes to lysosomes; and (9) a viral restriction/host factor whose antiviral activity involves PKR activation, stress granule formation, and competition with viral RNA elements, while its stability is regulated by SPOP-mediated ubiquitination (controlled by EGF-ERK phosphorylation), USP11-mediated deubiquitination, TRIM47-mediated K48-ubiquitination at Lys297 (promoted by Tim-3), PRMT1-mediated arginine methylation, and GCN5/CBP-mediated acetylation at Lys100.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ILF3 (NF90/NF110) is a multifunctional double-stranded RNA-binding protein that, in obligate heterodimer with NF45, governs gene expression at the transcriptional, post-transcriptional, and translational levels [#13, #22, #36]. NF45 dimerizes with NF90 through the DZF domain — structurally related to nucleotidyltransferases but catalytically inactive — and this association is reciprocally stabilizing and acts as a conformational scaffold that markedly enhances NF90's affinity for single- and double-stranded RNA [#22, #13, #36]. As a post-transcriptional regulator, NF90 binds AU-rich and structured 3'UTR elements to control the fate of target mRNAs, stabilizing transcripts such as IL-2, MKP-1, cyclin E1, and PARP1 while translationally repressing others through a defined AU-rich motif without altering stability [#4, #14, #28, #17]. The complex also acts as an RNA chaperone with annealing and strand-displacement activity [#38] and as a negative regulator of miRNA biogenesis, binding the stems of highly stable intronic pri-miRNAs in a manner largely exclusive of the Microprocessor [#16, #46]. In the nucleus ILF3 functions as a chromatin-associated transcriptional regulator, occupying active promoters and enhancers (IL-2, IL-13, c-fos) in complexes with NF45, Ku proteins, and DNA-PK, and through the latter interaction contributes to non-homologous end-joining repair of DNA double-strand breaks [#10, #19, #32, #33, #1, #20]. ILF3 additionally serves as a 60S ribosome biogenesis factor via its dsRNA-binding domains [#29] and as a mediator of mTORC1 amino-acid sensing that tethers GATOR complexes to lysosomes [#48]. Its nucleo-cytoplasmic distribution is controlled by phosphorylation at Ser647 by AKT or PKCβI, which triggers export and downstream mRNA stabilization [#12, #18], and isoform-specific nucleolar targeting is conferred by an alternatively spliced N-terminal signal [#31]. In antiviral defense, ILF3 binds viral RNA elements, activates PKR, and promotes stress-granule formation [#27, #45], while its abundance is set by competing ubiquitination and stabilizing modifications — SPOP-mediated degradation antagonized by EGF-ERK phosphorylation, TRIM47-mediated K48 ubiquitination at Lys297, USP11-mediated deubiquitination, and PRMT1-mediated arginine methylation [#41, #49, #57, #55].\",\n  \"teleology\": [\n    {\n      \"year\": 1994,\n      \"claim\": \"Established ILF3 as a DNA-binding transcription factor subunit by identifying NF90 and NF45 as the components binding the IL-2 promoter antigen receptor response element, defining its founding role in T-cell gene activation.\",\n      \"evidence\": \"Protein purification, cDNA cloning, immunofluorescence, and EMSA with pharmacological (cyclosporin A/FK506) validation in Jurkat T cells\",\n      \"pmids\": [\"7519613\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct versus bridged DNA contact not resolved\", \"Mechanism coupling T-cell stimulation to DNA-binding activity unknown\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Linked NF90/NF45 to DNA double-strand break machinery by showing physical interaction with and phosphorylation by DNA-PK, foreshadowing a DNA repair role.\",\n      \"evidence\": \"EMSA, in vitro kinase assay, and reciprocal immunoprecipitation with DNA-PKcs/Ku\",\n      \"pmids\": [\"9442054\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional consequence of NF90 phosphorylation by DNA-PK not defined here\", \"Direct repair contribution established only later\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Revealed how ILF3 is exported from the nucleus by reconstituting an exportin-5/RanGTP/minihelix-RNA quaternary complex, identifying the export receptor and coupling ILF3 trafficking to structured RNA.\",\n      \"evidence\": \"Gel retardation, GST pulldown, and HeLa nuclear microinjection of VA1 RNA\",\n      \"pmids\": [\"14570900\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether endogenous cellular RNAs use the same export route unclear\", \"Relationship to phosphorylation-triggered export not addressed\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Demonstrated that ILF3 is hijacked as a host factor by RNA viruses, binding pestivirus 5' and 3' non-translated regions required for replication.\",\n      \"evidence\": \"RNA-protein binding, viral RNA mutagenesis, and RNAi in BVDV replication assays\",\n      \"pmids\": [\"14592965\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether NF90 promotes or restricts replication context-dependent\", \"Direct enzymatic contribution to replication undefined\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Defined ILF3 as a post-transcriptional mRNA regulator by showing AU-rich 3'UTR binding stabilizes IL-2 mRNA and that nuclear export is required for this stabilization, connecting localization to function.\",\n      \"evidence\": \"RNA-protein binding, mRNA half-life, and subcellular fractionation in activated T cells\",\n      \"pmids\": [\"12504009\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Signal triggering export not yet identified at this stage\", \"Stabilization mechanism (decay factor competition) not resolved\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Provided in vivo genetic proof of a developmental post-transcriptional role: NF90 knockout mice die perinatally with muscle defects traced to direct binding of MyoD and p21 3'UTRs.\",\n      \"evidence\": \"NF90-/- mouse knockout, Northwestern blotting, and histology\",\n      \"pmids\": [\"15746098\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether 3'UTR binding stabilizes or translationally controls these mRNAs in vivo not dissected\", \"Tissue-specific requirement versus global essentiality unclear\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Identified the kinases and phosphosite (Ser647) controlling ILF3 export, showing AKT (CD28) and later PKCβI (PMA) phosphorylation is necessary and sufficient for export and IL-2 mRNA stabilization.\",\n      \"evidence\": \"In vitro kinase assays, phospho-specific antibodies, Ser647Ala mutagenesis, and export/stability readouts\",\n      \"pmids\": [\"18097023\", \"20870937\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Phosphatase reversing Ser647 unknown\", \"How phosphorylation alters export receptor engagement not shown\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Established the NF90-NF45 heterodimer as the functional unit and demonstrated its requirement for cell division, with reciprocal subunit stabilization and multinucleation upon depletion.\",\n      \"evidence\": \"RNAi depletion of individual subunits, flow cytometry, and time-lapse microscopy\",\n      \"pmids\": [\"18458058\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular target underlying division defect not identified here\", \"NF90 versus NF110 functional divergence only partly resolved\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Defined two complementary post-transcriptional mechanisms: translational repression via a discrete AU-rich 3'UTR motif, and negative control of miRNA biogenesis by pri-miRNA binding that excludes the Microprocessor.\",\n      \"evidence\": \"RIP-seq, polysome profiling, EGFP reporter mutagenesis, and pri/pre/mature miRNA quantification\",\n      \"pmids\": [\"19850717\", \"19398578\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How NF90 discriminates repression from stabilization targets unclear\", \"Selectivity for specific pri-miRNAs not yet mapped to structure\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Provided direct functional evidence that NF90-NF45 is required for non-homologous end joining, extending the early DNA-PK interaction into a defined DNA repair role.\",\n      \"evidence\": \"In vitro NHEJ assay with immunodepletion, γH2AX foci, and ionizing radiation sensitivity\",\n      \"pmids\": [\"21969602\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Step in NHEJ at which NF90/NF45 acts undefined\", \"Whether RNA binding is required for repair function not tested\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Provided the structural basis for heterodimer assembly, showing NF45 dimerizes with NF90 through a catalytically dead nucleotidyltransferase-like DZF domain.\",\n      \"evidence\": \"1.9-Å X-ray crystallography and Co-IP with interface mutants\",\n      \"pmids\": [\"22833610\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structure of RNA-bound holocomplex not determined\", \"How dimerization is coupled to RNA-binding enhancement not structurally shown\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Identified ILF3 as a ribosome biogenesis factor required for 60S maturation, acting through its dsRNA-binding domains on pre-60S particles.\",\n      \"evidence\": \"Tandem affinity purification, density gradient sedimentation, RNAi, and rRNA processing/nucleolar morphology analysis\",\n      \"pmids\": [\"26240280\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Specific rRNA or assembly intermediate substrate not defined\", \"Relationship to nucleolar isoform targeting not integrated\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Established ILF3 as a direct transcriptional coactivator independent of RNA binding, reconstituting c-fos activation in vitro with PC4 and Mediator.\",\n      \"evidence\": \"In vitro transcription reconstitution, ChIP, and dsRBM-inactive mutant analysis\",\n      \"pmids\": [\"26381409\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Generality across activator-dependent genes not established\", \"Contacts with specific general transcription factors not mapped\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Mapped the genome-wide chromatin footprint of NF90/NF110 at active promoters and enhancers and linked occupancy to control of proliferation and differentiation programs.\",\n      \"evidence\": \"ChIP-seq with ENCODE comparison and RNA-seq after shRNA knockdown in K562\",\n      \"pmids\": [\"29590119\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct versus indirect regulation of individual loci not resolved\", \"Mechanism of chromatin recruitment unknown\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Defined the biochemical basis of NF90-NF45 RNA chaperone activity (annealing/strand displacement) and connected nuclear circRNA biogenesis to cytoplasmic antiviral redistribution.\",\n      \"evidence\": \"In vitro annealing/strand-displacement assays and genome-wide siRNA screen with RIP and fractionation\",\n      \"pmids\": [\"29040738\", \"28625552\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo substrates of chaperone activity beyond test cases unclear\", \"Trigger for infection-induced cytoplasmic translocation not molecularly defined\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Resolved the antiviral pathway placement of ILF3, showing it activates PKR, promotes stress-granule formation, and is required for translation of IFNB1 and a subset of interferon-stimulated genes.\",\n      \"evidence\": \"Co-IP/domain mapping, PKR phosphorylation and stress-granule assays, and polysome profiling with NGS\",\n      \"pmids\": [\"24623135\", \"31701124\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How ILF3 selects ISG transcripts for translation unclear\", \"Balance between restriction and pro-viral host-factor roles not unified\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Refined the miRNA biogenesis mechanism, defining the subset of highly stable intronic pri-miRNAs whose stem structure is recognized by NF90 to the exclusion of the Microprocessor.\",\n      \"evidence\": \"CLIP-seq/RIP-seq, miRNA microarray, and EMSA with RNA structure mutants\",\n      \"pmids\": [\"32427329\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Determinants distinguishing NF90-regulated from unregulated pri-miRNAs incomplete\", \"Whether NF45 is required for stem recognition not addressed\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identified ILF3 as a conserved mediator of mTORC1 amino-acid sensing that tethers GATOR complexes to lysosomes, extending ILF3 beyond nucleic-acid biology into nutrient signaling.\",\n      \"evidence\": \"Genome-wide CRISPR screen, lysosome-targeting rescue epistasis, and cross-species conservation\",\n      \"pmids\": [\"37037994\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether RNA binding contributes to GATOR tethering unknown\", \"Direct ILF3-GATOR contacts not structurally defined\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Defined the post-translational control of ILF3 abundance, showing EGF-ERK phosphorylation blocks SPOP-mediated degradation and that TRIM47 (downstream of Tim-3) ubiquitinates Lys297 to drive degradation, with stabilized ILF3 controlling metabolic and antiviral outputs.\",\n      \"evidence\": \"Ubiquitination assays, site-specific (Lys297) mutagenesis, in vivo PDX and VSV mouse models\",\n      \"pmids\": [\"31772275\", \"34110282\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Crosstalk between competing ubiquitination/deubiquitination/methylation events not integrated\", \"Whether these PTMs alter localization or only abundance unclear\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How ILF3's many functions — transcriptional coactivation, mRNA stability/translation control, miRNA biogenesis, ribosome assembly, DNA repair, and mTORC1 sensing — are partitioned across isoforms, localizations, and PTM states within a single cell remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No unifying model linking NF90 versus NF110 isoforms to specific functions\", \"Structure of RNA- or DNA-bound functional complexes not determined\", \"Rules governing which targets are stabilized versus repressed unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [4, 17, 36, 38, 46]},\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [0, 10, 19, 32]},\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [32, 33]},\n      {\"term_id\": \"GO:0045182\", \"supporting_discovery_ids\": [2, 17, 24]},\n      {\"term_id\": \"GO:0140098\", \"supporting_discovery_ids\": [38, 16, 46]},\n      {\"term_id\": \"GO:0060089\", \"supporting_discovery_ids\": [48]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [0, 3, 7]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [4, 27]},\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [29, 31]},\n      {\"term_id\": \"GO:0005654\", \"supporting_discovery_ids\": [15, 37]},\n      {\"term_id\": \"GO:0005764\", \"supporting_discovery_ids\": [48]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [16, 17, 37, 38, 46]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [0, 10, 19, 32, 33]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [27, 45, 49]},\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [1, 20]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [13, 28]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [48]},\n      {\"term_id\": \"R-HSA-1852241\", \"supporting_discovery_ids\": [29]}\n    ],\n    \"complexes\": [\"NF90-NF45 heterodimer\", \"NF110-NF45 heterodimer\", \"DNA-PK (DNA-PKcs–Ku) complex\"],\n    \"partners\": [\"ILF2\", \"PRKDC\", \"XRCC5\", \"XRCC6\", \"EIF2AK2\", \"EIF4A1\", \"PERK\", \"NS5A\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}