{"gene":"NUCLEOLIN","run_date":"2026-06-10T05:19:52","timeline":{"discoveries":[{"year":1984,"finding":"Nucleolin (protein C23) localizes to the fibrillar shell surrounding the fibrillar center and to the fibrillar center of the nucleolus during interphase, and remains associated with NOR-containing structures (rDNA) during mitosis, as determined by silver staining and immunolocalization in Novikoff hepatoma cells.","method":"Silver staining, immunofluorescence, immunoelectron microscopy","journal":"Chromosoma","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct localization by multiple orthogonal imaging methods, replicated across labs and consistent with subsequent studies","pmids":["6206987"],"is_preprint":false},{"year":1983,"finding":"During mitosis, nucleolin (C23) remains associated with disappearing nucleoli through prophase, then reappears in prenucleolar bodies and subsequently the nucleolus during telophase, while B23 follows a distinct pattern; C23 co-localizes with silver-staining NOR proteins throughout mitosis.","method":"Dual immunofluorescence with anti-B23 and anti-C23 antibodies in PtK2 cells","journal":"Experimental cell research","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct cell-cycle-stage-resolved localization, consistent with multiple independent studies","pmids":["6345184"],"is_preprint":false},{"year":1986,"finding":"Nucleolin (C23) directly associates with rapidly labeled pre-rRNA (18S and 28S sequences) in preribosomal RNP particles, as demonstrated by UV cross-linking and immunoprecipitation from Novikoff hepatoma nucleoli.","method":"UV cross-linking, immunoprecipitation with anti-C23 antibody, dot-blot hybridization with rDNA probes, sucrose gradient fractionation","journal":"Biochemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — UV cross-linking and immunoprecipitation with multiple orthogonal methods in a single study","pmids":["3790520"],"is_preprint":false},{"year":1987,"finding":"Nucleolin is a natural preferential substrate for a nucleolar casein kinase II (type NII). The kinase phosphorylates nucleolin in vitro at serine residues in two highly acidic tryptic fragments (A: residues 21–49; C: residues 180–221) near the amino terminus, at canonical CKII consensus sites.","method":"Co-purification, in vitro kinase assay, peptide mapping, Km/Vmax analyses","journal":"Biochemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstituted kinase assay with peptide-level mapping, replicated by multiple independent groups","pmids":["3427111"],"is_preprint":false},{"year":1988,"finding":"Phosphopeptide patterns from nucleolin phosphorylated by purified casein kinase II (CKII) in vitro are identical to those from nucleolin isolated from tumor cells grown with [32P]-phosphate, establishing nucleolin as a physiological CKII substrate with ~2 mol phosphate per mol nucleolin.","method":"In vitro phosphorylation, partial tryptic digest, comparison of in vitro vs. in vivo phosphopeptide patterns","journal":"Biochemical and biophysical research communications","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct in vitro reconstitution compared to in vivo labeling, consistent with prior work","pmids":["3190709"],"is_preprint":false},{"year":1988,"finding":"Microinjection of anti-C23 antibody into Chironomus salivary gland nuclei caused a 2–3.5-fold stimulation of 32P incorporation into 38S pre-rRNA, selectively for pre-rRNA without affecting extranucleolar RNA, indicating nucleolin acts as a negative regulator of rDNA transcription.","method":"Microinjection of anti-C23 antibody into nuclei, 32P labeling, RNA extraction and electrophoresis","journal":"Experimental cell research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct functional perturbation in living cells with specific controls (preimmune serum, absorbed antibody), single lab","pmids":["3169130"],"is_preprint":false},{"year":1990,"finding":"Nucleolin is phosphorylated on threonines at TPXK motifs in the amino-terminal domain by p34cdc2 (Cdc2 kinase) specifically during mitosis, distinct from the interphase CKII-mediated serine phosphorylation; the same sites used in vivo during mitosis were phosphorylated by M-phase H1 kinase in vitro.","method":"In vitro kinase assay with starfish M-phase H1 kinase, in vivo 32P labeling, phosphoamino acid analysis, peptide mapping","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro kinase assay corroborated by in vivo labeling; sites matched consensus and were mapped at the peptide level","pmids":["2192260"],"is_preprint":false},{"year":1991,"finding":"Yeast NSR1 (nucleolin ortholog) specifically binds nuclear localization sequences (NLS), contains two RNA recognition motifs, and is required for normal cell growth; disruption of NSR1 causes severe growth defect, establishing functional conservation with mammalian nucleolin.","method":"Affinity purification on NLS peptide column, gene disruption, immunofluorescence, antibody-based cloning","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — biochemical purification plus genetic disruption, replicated and extended by multiple groups","pmids":["1706724"],"is_preprint":false},{"year":1992,"finding":"Yeast NSR1 is required for pre-rRNA processing: nsr1 deletion blocks initial 35S pre-rRNA processing, nearly eliminates 20S pre-rRNA, reduces 18S rRNA, and disrupts the free 40S ribosomal subunit pool, demonstrating a conserved role of the nucleolin ortholog in ribosome biogenesis.","method":"Pulse-labeling of rRNA, Northern blot, sucrose gradient fractionation of ribosomal subunits, antibiotic sensitivity assay","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods in genetic KO, independently replicated","pmids":["1508189"],"is_preprint":false},{"year":1992,"finding":"Yeast NSR1 is required for pre-rRNA processing and ribosome biogenesis: nsr1 deletion strains accumulate 35S pre-rRNA, show slow processing of 35S, impaired 18S rRNA methylation, and a reduced 40S:60S ratio; cold shock greatly exacerbates the processing defect.","method":"Pulse-labeling, pulse-chase rRNA analysis, Northern blot, NSR1 cold-shock induction assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic deletion with multiple rRNA processing readouts, replicated independently","pmids":["1644811","1644812"],"is_preprint":false},{"year":1993,"finding":"Nucleolar localization of NSR1 requires a bipartite NLS and is mediated redundantly by either the N-terminal acidic/serine-rich domain or both RNA recognition motifs (RRMs); point mutations in RNP consensus octamers cause nuclear mislocalization, and the RGG domain is necessary for nucleolar accumulation when one RNP octamer is mutated.","method":"Deletion analysis with beta-galactosidase fusion reporter, immunofluorescence, point mutagenesis","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — systematic deletion and point mutation analysis with reporter fusions, single lab","pmids":["8245119"],"is_preprint":false},{"year":1994,"finding":"Nucleolin functions as a transcriptional repressor of the acute-phase response gene alpha-1 acid glycoprotein (AGP); nucleolin was purified from mouse hepatoma cells as the B-motif-binding factor and biochemical studies confirmed its repressor activity.","method":"Affinity purification, amino acid sequence analysis, transcription assay, DNA-binding assay","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — protein purification combined with functional transcription assay, single lab","pmids":["8065340"],"is_preprint":false},{"year":1996,"finding":"Nucleolin (C23) interacts with nucleolar protein B23 (nucleophosmin); co-immunoprecipitation from HeLa nuclear extract with either anti-C23 or anti-B23 monoclonal antibodies confirmed the interaction, which requires residues 540–628 of C23 (the nucleolar localization region) and residues 194–239 of B23.","method":"Two-hybrid system, co-immunoprecipitation, deletion mutant binding studies","journal":"European journal of biochemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — orthogonal methods (two-hybrid + reciprocal co-IP + deletion mapping), replicated","pmids":["8620867"],"is_preprint":false},{"year":1999,"finding":"B23 (nucleophosmin) and nucleolin (C23) interact in vivo during interphase and cytokinesis but not during prometaphase/metaphase; the interaction persists even after actinomycin D-induced translocation to the nucleoplasm, and mitotic phosphorylation of B23 alone does not explain loss of interaction during mitosis.","method":"Chemical cross-linking with DSP, co-immunoprecipitation, in vitro Cdc2 kinase phosphorylation of GST-B23 followed by co-IP","journal":"Cancer letters","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — cross-linking co-IP with cell-cycle synchronization and in vitro kinase experiment, single lab","pmids":["10503877"],"is_preprint":false},{"year":2000,"finding":"Cell-surface nucleolin is transported to the plasma membrane via an active, non-conventional pathway independent of the ER-Golgi complex; surface nucleolin clusters in an actin cytoskeleton-dependent manner upon antibody cross-linking and can be internalized, mediating intracellular import of ligands.","method":"Confocal and electron microscopy, antibody cross-linking experiments, inhibitor studies (low temperature, serum-free medium, glycoprotein transport inhibitors), actin disruption","journal":"Experimental cell research","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal localization and functional inhibition methods in a single study, replicated by other groups","pmids":["11112338"],"is_preprint":false},{"year":2001,"finding":"Nucleolin RBD1 and RBD2 together (RBD12), but neither alone, specifically recognize a stem-loop NRE (nucleolin recognition element) RNA structure; in the complex, the hairpin loop adopts a well-defined conformation distinct from the free RNA, with both loop E motif and hairpin loop interacting specifically with the protein.","method":"NMR spectroscopy of free and protein-bound RNA, titration of RBD1, RBD2, and RBD12 with sNRE","journal":"Journal of molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — NMR structure determination with functional binding titrations, single lab but rigorous","pmids":["11397095"],"is_preprint":false},{"year":2002,"finding":"Following cell stress (ionizing radiation, camptothecin, heat shock), nucleolin relocalizes from the nucleolus to the nucleoplasm in a p53-dependent manner; the p53 C-terminal regulatory domain is required for nucleolin-p53 complex formation and nucleolin mobilization, which is independent of p53 transactivation; nucleolin and p53 interact directly in vitro.","method":"Immunofluorescence, co-immunoprecipitation, in vitro binding assay with deletion mutants, p53-null and mutant cell lines","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct in vitro interaction confirmed, in vivo relocalization with genetic requirement for p53 C-terminal domain, multiple orthogonal methods","pmids":["12138209"],"is_preprint":false},{"year":2002,"finding":"Activation of nucleolin RNA-binding activity after genotoxic stress (UV or ionizing radiation) is mediated by the stress-activated protein kinase p38; nucleolin was identified as a genotoxic stress-responsive RNA-binding protein that binds stress-responsive mRNAs.","method":"Purification, RNA binding assays, p38 inhibitor treatment, identification of 40 mRNA ligands","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biochemical purification with RNA binding assays and kinase inhibitor studies, single lab","pmids":["12000845"],"is_preprint":false},{"year":2004,"finding":"Nucleolin interacts with the telomerase reverse transcriptase subunit hTERT through its RNA-binding domain 4 and RGG domain, with the interaction also involving hTERC (telomerase RNA); this nucleolin-hTERT interaction is critical for nucleolar localization of hTERT.","method":"Co-immunoprecipitation, domain deletion mapping, immunofluorescence colocalization","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — domain-mapped co-IP with functional localization readout, single lab","pmids":["15371412"],"is_preprint":false},{"year":2005,"finding":"Nucleolin functions as a macrophage cell-surface receptor for early apoptotic cells bearing polylactosaminyl CD43; anti-nucleolin antibody or deletion of the antibody-binding region abolishes binding to early apoptotic cells; nucleolin-transfected HEK293 cells acquire ability to bind early apoptotic cells.","method":"Antibody blocking, nucleolin transfection of HEK293, deletion mutant analysis, competitive inhibition with oligosaccharides","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — gain-of-function transfection plus deletion analysis and competitive inhibition, single lab","pmids":["16135517"],"is_preprint":false},{"year":2005,"finding":"Extranuclear nucleolin undergoes complex N- and O-glycosylations; N-glycosylation sites were mapped to N317 and N492 within RNA-binding domains 1 and 3, respectively, suggesting glycosylation may regulate RNA-binding function.","method":"SDS-PAGE, mass spectrometry, tunicamycin treatment, lectin binding, exoglycosidase digestion, MALDI-TOF, monosaccharide composition analysis","journal":"Biochemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — multiple orthogonal biochemical methods including MS-based glycan characterization with site identification, single lab","pmids":["15823039"],"is_preprint":false},{"year":2006,"finding":"Nucleolin is required for RNA polymerase I transcription of chromatin (but not naked DNA) templates in vitro, and is specifically associated by ChIP with rRNA genes transcribed by RNAP I but not RNAP II or III; siRNA knockdown of nucleolin specifically inhibits RNAP I transcription.","method":"In vitro chromatin transcription assay, ChIP, immunofluorescence, siRNA knockdown","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution on chromatin templates, ChIP, and RNAi knockdown with multiple orthogonal readouts","pmids":["17130237"],"is_preprint":false},{"year":2007,"finding":"Nucleolin depletion by RNAi results in disorganized nucleoli at interphase, prolonged cell cycle with misaligned chromosomes, syntelic kinetochore-microtubule attachments with reduced centromere stretching, and defects in spindle organization; during mitosis, CDC2-phosphorylated nucleolin associates with spindle poles from prometaphase to anaphase.","method":"RNAi depletion, high-resolution microscopy, antibody specific for CDC2-phosphorylated nucleolin, kinetochore-microtubule attachment analysis","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 2 / Moderate — clean RNAi depletion with specific mitotic phenotypes and phospho-specific localization, single lab with multiple orthogonal readouts","pmids":["17535846"],"is_preprint":false},{"year":2007,"finding":"Endostatin specifically binds cell-surface nucleolin with high affinity; blockage or knockdown of nucleolin abolishes endostatin's antiendothelial activity in vitro and antiangiogenic/antitumor activity in vivo; endostatin is internalized into cell nuclei via nucleolin; endostatin inhibits mitosis-related phosphorylation of nucleolin in the nucleus.","method":"Binding assay, neutralizing antibody, siRNA knockdown, colocalization, internalization assay, in vivo tumor model","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods including in vivo validation and mechanistic phosphorylation studies, single lab","pmids":["17615292"],"is_preprint":false},{"year":1999,"finding":"Urokinase (uPA) induces formation of a signaling complex on the cell surface containing uPAR, nucleolin, and casein kinase 2; nucleolin and CK2 were isolated by affinity chromatography and co-localized with uPAR; uPA activates CK2 in this complex leading to nucleolin phosphorylation; blocking nucleolin or CK2 inhibits uPA-induced cell proliferation.","method":"Affinity chromatography, nano-electrospray MS, immunoblotting, laser scanning and immunoelectron microscopy, co-immunoprecipitation, in vitro kinase assay, cell proliferation assay","journal":"Current biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — affinity isolation, MS identification, co-IP, in vitro kinase assay, and functional inhibition studies with multiple orthogonal methods","pmids":["10607589"],"is_preprint":false},{"year":2011,"finding":"Nucleolin interacts with the miRNA microprocessor components DGCR8 and Drosha in the nucleus; nucleolin directly and specifically binds the primary miR-15a/16 transcript; nucleolin is required for primary-to-precursor miRNA processing of miR-15a/16 in vitro; nuclear localization of nucleolin is critical for this function.","method":"Overexpression and knockdown studies, in vitro processing assay with cell extracts, co-immunoprecipitation, direct RNA binding assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro processing assay with rescue by nucleolin addition, plus direct binding and co-IP, single lab","pmids":["22049078"],"is_preprint":false},{"year":2012,"finding":"Nucleolin depletion increases H3K9me2 (heterochromatin mark) and decreases H4K12Ac and H3K4me3 (euchromatin marks) at rRNA genes; ChIP-seq shows nucleolin enrichment at rDNA coding and promoter regions, preferentially at unmethylated genes; nucleolin depletion causes RNAP I accumulation at the start of transcription units and decreased UBF along the gene; nucleolin antagonizes binding of TTF-1 at the T0 terminator, thereby inhibiting TIP5 and HDAC1 recruitment and repressive heterochromatin formation.","method":"ChIP-seq, ChIP, siRNA knockdown, histone mark analysis, RNAP I and UBF distribution analysis","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome-scale ChIP-seq with functional siRNA knockdown and multiple chromatin mark readouts, single lab with rigorous controls","pmids":["22859736"],"is_preprint":false},{"year":2012,"finding":"Nucleolin suppresses p53 mRNA translation using both 5'- and 3'-UTRs; nucleolin binds the same 5'-3'-UTR interaction region critical for RPL26 recruitment; nucleolin oligomerizes (dimerizes) through its RNA-binding domain; RPL26 disrupts NCL dimerization; NCL's RNA-binding domain mediates both dimerization and translational repression, and is the domain that interacts with RPL26.","method":"Translation assays, RNA binding assays, co-immunoprecipitation, domain deletion and point mutation analysis, dimerization assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — domain-level dissection with multiple orthogonal methods and functional rescue experiments, single lab","pmids":["22433872"],"is_preprint":false},{"year":2012,"finding":"Nucleolin binds LINE-1 ORF2 IRES and functions as an IRES trans-acting factor (ITAF) for ORF2 translation; NCL knockdown specifically reduces ORF2 IRES activity, L1 and L1-assisted Alu retrotransposition without altering L1 RNA or ORF1 protein abundance.","method":"RNA affinity chromatography, RNAi knockdown, IRES activity assay, retrotransposition assay","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — RNA affinity chromatography plus functional IRES/retrotransposition assays, single lab","pmids":["23161687"],"is_preprint":false},{"year":2011,"finding":"HuR promotes nucleolin translation via the nucleolin 3'UTR without affecting mRNA levels; miR-494 inhibits nucleolin expression by enhancing NCL mRNA association with Argonaute-containing complexes and promoting NCL mRNA transport to processing bodies (PBs); HuR and miR-494 functionally compete to control nucleolin abundance.","method":"MS2-tagged 3'UTR pull-down, immunoprecipitation, HuR silencing, miR-494 transfection, P-body tracking, ribosome loading assay","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods in a single study, single lab","pmids":["21859890"],"is_preprint":false},{"year":2015,"finding":"Cell-surface nucleolin mediates EV71 binding and infection; EV71 interacts directly with nucleolin via the VP1 capsid protein; knockdown of nucleolin reduces EV71 binding, infection, and production; expression of human nucleolin on mouse cells confers EV71 infection.","method":"Glycoproteomics, immunoprecipitation with EV71 particles, anti-nucleolin antibody blocking, siRNA knockdown, gain-of-function expression in mouse cells","journal":"Journal of virology","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods including gain-of-function and knockdown with multiple viral readouts, single lab","pmids":["25673703"],"is_preprint":false},{"year":2016,"finding":"Nucleolin depletion reduces axonal levels of importin β1 mRNA and protein; subcellular sequestration of nucleolin or importin β1 enhances axonal growth and causes a subcellular shift in protein synthesis; nucleolin associates with importin β1 mRNA in axons and with kinesins for anterograde transport.","method":"RNAi/siRNA knockdown, subcellular fractionation, in situ hybridization, cell growth assays in neurons and fibroblasts","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple cell types with knockdown and localization experiments, single lab","pmids":["27477284"],"is_preprint":false},{"year":2017,"finding":"NCL phosphorylation at six CK2 consensus sites in the N-terminus is required for activation of PARN deadenylase activity upon oncogenic stimuli and UV stress; NCL directly interacts with PARN; under non-stress conditions NCL forms complexes with p53 and HuR; phosphorylation state of NCL determines specificity of its protein-protein and protein-RNA interactions governing mRNA deadenylation.","method":"In vitro deadenylase assay, co-immunoprecipitation, phospho-mutant (NCL-6/S*A) analysis, RNA binding assay","journal":"RNA biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro PARN assay with phosphomutant and co-IP, single lab","pmids":["29168431"],"is_preprint":false},{"year":2018,"finding":"LINE1 RNA recruits nucleolin and Kap1/Trim28 to repress Dux (master activator of the 2-cell program) in ESCs; LINE1 RNA also mediates binding of nucleolin and Kap1 to rDNA, promoting rRNA synthesis; in pre-implantation embryos, LINE1 RNA is required for Dux silencing, rRNA synthesis, and exit from the 2-cell stage.","method":"RNA immunoprecipitation, ChIP, siRNA/shRNA knockdown, RNA FISH, embryo functional assays","journal":"Cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal IP, ChIP, and functional embryo assays with multiple orthogonal methods","pmids":["29937225"],"is_preprint":false},{"year":2019,"finding":"Binding of cellular nucleolin to the HCV core RNA G-quadruplex (G4) structure stabilizes the G4 and suppresses HCV replication; NCL co-localizes with HCV particles; HCV infection upregulates NCL; silencing NCL greatly enhances viral RNA replication.","method":"Direct binding assay (in vitro and in-cell), colocalization, NCL knockdown with viral replication assay, G4-mutant virus comparison","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct binding plus functional knockdown assay with G4-mutant virus control, single lab","pmids":["30462330"],"is_preprint":false},{"year":2018,"finding":"Nucleolin mediates RHDV internalization via clathrin-dependent endocytosis; NCL interacts specifically with RHDV capsid protein VP60 through NCL N-terminal residues 285–318 and VP60's DVN motif (472Asp-Val-Asn474); NCL also interacts with clathrin light chain A C-terminus; blocking the NCL-VP60 interaction with a DVN peptide markedly reduces virus internalization.","method":"Biochemical inhibitors, RNAi, co-immunoprecipitation, domain deletion analysis, competitive peptide inhibition, in vivo animal experiments","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 2 / Strong — domain-mapped co-IP, functional blocking with specific peptide, and in vivo animal validation","pmids":["30339712"],"is_preprint":false},{"year":2019,"finding":"Shear stress-activated AMPK phosphorylates nucleolin at serine 328 (S328), which sequesters nucleolin in the nucleus and inhibits processing of miR-93 and miR-484 that target KLF2 and eNOS mRNAs; this identifies an AMPK-NCL-miR-93/miR-484 axis in endothelial mechanotransduction.","method":"In silico analysis, phosphorylation site mapping (S328), AMPK inhibitor and activator studies, anti-miR rescue experiments, posttranslational modification analysis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — phospho-site identified with functional rescue experiments, single lab","pmids":["31182601"],"is_preprint":false},{"year":2020,"finding":"Nucleolin strongly and preferentially binds G-quadruplex structures with long central loops (5–9 nucleotides) over short-loop variants; photo-cross-linking identified a 15-amino acid fragment in helix α2 of RBD2 as a loop-contact site; the RGG domain participates in terminal guanine quartet recognition; thus NCL recognizes G4 via concerted action of RBD2 (loop) and RGG (quartet) domains.","method":"Binding assays, photo-cross-linking with BrU-modified G4 sequences, quantitative proteomics (LC-MS/MS), G4 ligand competition assays","journal":"Biochemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — photo-cross-linking with MS-based site mapping plus systematic loop-length and ligand competition experiments, single lab","pmids":["32191439"],"is_preprint":false},{"year":2022,"finding":"A cysteine tRNA-derived fragment (5'-tRFCys) directly binds nucleolin and promotes its oligomerization into a higher-order ribonucleoprotein complex that stabilizes bound metabolic mRNAs (Mthfd1l and Pafah1b1) against exonucleolytic degradation, driving breast cancer metastasis.","method":"Small RNA profiling, direct binding assay, oligomerization assay, RNA stability assay, functional metastasis assays","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct binding established, oligomerization and mRNA stability mechanistically linked, functional metastasis readout, single lab with multiple orthogonal methods","pmids":["35654044"],"is_preprint":false},{"year":2014,"finding":"Nucleolin directly activates NFκB signaling; nucleolin overexpression increases NFκB phosphorylation and upregulates DNMT1; NFκB inactivation diminishes DNMT1 promoter activity, establishing a nucleolin-NFκB-DNMT1 axis in leukemia.","method":"Forced expression and RNAi knockdown, NFκB reporter assay, DNMT1 promoter activity assay, Western blot, in vitro and in vivo proliferation/tumorigenesis assays","journal":"Oncotarget","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional assays with promoter reporter and genetic knockdown/overexpression, single lab","pmids":["25015109"],"is_preprint":false},{"year":2017,"finding":"Nucleolin identifies nucleolin as binding partner of topoisomerase II-alpha (TopIIA); this interaction is mapped to RNA-binding domain 3 of nucleolin and is essential for blocking DNA damage and apoptosis; nucleolin silencing decreases TopIIA decatenation activity and enhances formation of TopIIA-DNA cleavable complexes in the presence of etoposide.","method":"Co-immunoprecipitation, domain deletion mapping, TopIIA decatenation assay, cleavable complex assay, siRNA knockdown with DNA damage assays","journal":"Leukemia","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — domain-mapped co-IP with enzymatic activity assay, single lab","pmids":["28690315"],"is_preprint":false},{"year":2024,"finding":"Nucleolin is lactylated predominantly at lysine K477 by the acyltransferase P300 in response to glycolytic hyperactivity; K477 lactylation promotes efficient translation of MADD mRNA by preventing alternative splicing that generates a premature termination codon; the resulting MADD protein activates ERK signaling and drives iCCA tumor growth.","method":"Proteomics, mass spectrometry, macromolecule interaction studies, P300 acyltransferase assay, alternative splicing analysis, xenograft tumor model, clinical sample validation","journal":"Journal of hepatology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — site-specific PTM identified by MS with P300 as writer enzyme, mechanistic link to splicing and ERK signaling validated in vivo, single lab with multiple methods","pmids":["38679071"],"is_preprint":false},{"year":2021,"finding":"Nucleolin acts as a cell-surface receptor for C1QTNF4; interaction is mediated by the second C1q-like domain of C1QTNF4 and the C-terminus of nucleolin; upon cell binding, C1QTNF4 is actively internalized in monocytes; nucleolin may act as a docking molecule with context-dependent coreceptors.","method":"Mass spectrometric analysis of cell-surface proteins, co-immunoprecipitation, domain deletion analysis, imaging flow cytometry internalization assay","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — domain-mapped MS identification plus co-IP and internalization imaging, single lab","pmids":["33676896"],"is_preprint":false},{"year":2025,"finding":"Crystal structure (2.6 Å) of nucleolin bound to the MYC promoter G-quadruplex (MycG4) reveals: MycG4 adopts a folded parallel three-tetrad structure with two K+ ions; RBD1, RBD2, and Linker12 interact with the 6-nt central loop and 5' flanking region; RBD3 and RBD4 bind the 1-nt loops (confirmed by NMR); all four RBDs are required for high-affinity MycG4 binding; CUT&Tag sequencing confirmed nucleolin binding to MycG4 in cells, suggesting G4s are primary cellular substrates of nucleolin.","method":"X-ray crystallography (2.6 Å), NMR spectroscopy, CUT&Tag sequencing, binding affinity measurements","journal":"Science","confidence":"High","confidence_rationale":"Tier 1 / Strong — atomic-resolution crystal structure plus NMR validation plus in-cell chromatin binding, multiple orthogonal methods in a single rigorous study","pmids":["40245140"],"is_preprint":false},{"year":2024,"finding":"The E3 ubiquitin ligase TRIM21 mediates K48-linked polyubiquitination and proteasomal degradation of nucleolin; circRNA circ0006646 protects nucleolin from TRIM21-mediated ubiquitination by preventing the TRIM21-NCL interaction.","method":"Multi-omics proteomics, co-immunoprecipitation, ubiquitination assay, proteasome inhibitor treatment, in vivo xenograft and lentivirus models","journal":"Advanced science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ubiquitination assay with TRIM21 as E3 ligase identified, K48-linkage specified, but findings come from a circRNA-focused study; single lab","pmids":["38357830"],"is_preprint":false},{"year":2011,"finding":"Cell-surface nucleolin on alveolar macrophages directly binds LPS via affinity chromatography; LPS co-localizes with C23 on cell surface and in cytoplasm; siRNA knockdown of C23 reduces LPS internalization, LPS-induced NF-κB DNA binding, and TNF-α and IL-6 protein expression.","method":"LPS affinity chromatography, immunofluorescence colocalization, siRNA knockdown, NF-κB DNA binding assay, cytokine measurement","journal":"Cell biology international","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct binding demonstrated with functional siRNA knockdown, single lab","pmids":["21309751"],"is_preprint":false},{"year":2019,"finding":"Nucleolin on the plasma membrane of cardiomyocytes binds mitochondrial DNA (mtDNA) with ~10-fold greater affinity than nuclear DNA; blocking nucleolin with midkine reduces IL-1β/TNF-α expression and inhibits cellular uptake of CpG-DNA; nucleolin inhibition with AS1411 reduces IL-6 release; nucleolin facilitates CpG-DNA internalization independently of classical endocytosis.","method":"Microscale thermophoresis (direct binding), immunofluorescence, nucleolin inhibitor (midkine and AS1411) studies, CpG-DNA uptake assay, cytokine measurement","journal":"British journal of pharmacology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct binding measured with MST plus functional inhibition studies, single lab","pmids":["31412132"],"is_preprint":false},{"year":2023,"finding":"G-quadruplex structures in MALAT1 lncRNA (in the 3' region) are specifically bound by nucleolin and nucleophosmin; G4-disrupting G-to-A mutations in MALAT1 abolish localization of both NCL and NPM to nuclear speckles; truncated NCL (ΔNCL) binds all three MALAT1 rG4s with high affinity in vitro.","method":"rG4 domain-specific RNA pull-down, mass spectrometry, RNA immunoprecipitation, imaging, G-to-A mutagenesis, in vitro biophysical binding studies","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — RNA pull-down with MS identification, mutagenesis functional readout, in vitro binding; single lab","pmids":["37558241"],"is_preprint":false}],"current_model":"Nucleolin (NCL/C23) is a multidomain nucleolar phosphoprotein whose tripartite structure (N-terminal acidic/serine domain, four RNA-binding domains RBD1–4, and C-terminal RGG domain) enables it to: (1) bind pre-rRNA and chromatin at rDNA loci to facilitate RNA polymerase I transcriptional elongation and maintain a euchromatic rDNA state by antagonizing TTF-1/TIP5/HDAC1-mediated repression; (2) participate in pre-rRNA processing and ribosome assembly (conserved from yeast NSR1 to mammals); (3) undergo cell-cycle-regulated phosphorylation—by CKII at serine residues during interphase (promoting rDNA transcription) and by Cdc2/CDK1 at threonine TPXK motifs during mitosis (linked to nucleolar chromatin reorganization); (4) recognize G-quadruplex structures (including the MYC promoter G4, revealed by a 2.6-Å crystal structure) through concerted action of RBD1–4 and the RGG domain; (5) suppress p53 mRNA translation by binding 5'–3'-UTR interaction regions and forming homodimers that are disrupted by RPL26 after DNA damage; (6) interact with LINE1 RNA as a nuclear scaffold to repress Dux and promote rRNA synthesis in ESCs; (7) translocate to the cell surface via a non-conventional, ER-Golgi-independent, actin-dependent pathway where it acts as a receptor/co-receptor for diverse ligands (endostatin, EV71, RHDV VP60, C1QTNF4, LPS, apoptotic cells) mediating internalization; and (8) be post-translationally regulated by CKII-mediated phosphorylation (controlling PARN deadenylase activation and mRNA stability), p38-mediated activation of RNA-binding after genotoxic stress, AMPK-mediated S328 phosphorylation (sequestering NCL in the nucleus to inhibit miRNA processing), P300-catalyzed K477 lactylation (redirecting RNA splicing to upregulate MADD/ERK), and TRIM21-mediated K48-ubiquitination leading to proteasomal degradation."},"narrative":{"mechanistic_narrative":"Nucleolin (NCL/C23) is a multidomain nucleolar phosphoprotein that couples ribosomal DNA transcription, pre-rRNA processing, and ribosome assembly to a broad set of nucleic-acid recognition activities through its tandem RNA-binding domains (RBD1–4) and C-terminal RGG domain [PMID:3790520, PMID:11397095, PMID:40245140]. It localizes to the fibrillar regions of the nucleolus during interphase and stays associated with NOR/rDNA structures throughout mitosis [PMID:6206987, PMID:6345184], where it binds pre-rRNA in preribosomal RNP particles [PMID:3790520]; the conserved yeast ortholog NSR1 is required for 35S pre-rRNA processing and 40S subunit formation, establishing this ribosome-biogenesis role across eukaryotes [PMID:1706724, PMID:1508189, PMID:1644811, PMID:1644812]. At rDNA, nucleolin is selectively required for RNA polymerase I transcription of chromatin templates and maintains a euchromatic, transcriptionally active state by antagonizing TTF-1 binding and the recruitment of TIP5/HDAC1-mediated repressive heterochromatin [PMID:17130237, PMID:22859736]. Its activity is governed by cell-cycle-coupled phosphorylation—by casein kinase II on N-terminal acidic-domain serines during interphase and by Cdc2/CDK1 on TPXK threonine motifs during mitosis, the latter directing nucleolin to spindle poles and proper kinetochore-microtubule attachment [PMID:3427111, PMID:3190709, PMID:2192260, PMID:17535846]. Through concerted action of its RBD and RGG domains, nucleolin recognizes G-quadruplex structures, demonstrated at atomic resolution for the MYC promoter G4, where all four RBDs engage the folded quadruplex [PMID:32191439, PMID:40245140]. Beyond rRNA, nucleolin regulates gene expression post-transcriptionally: it represses p53 mRNA translation by binding a 5'–3'-UTR interaction region and homodimerizing through its RNA-binding domain, an interaction disrupted by RPL26 [PMID:22433872]; it participates in microprocessor-dependent miRNA maturation with Drosha/DGCR8 [PMID:22049078]; and it stabilizes bound mRNAs as part of higher-order ribonucleoprotein assemblies [PMID:35654044]. A second functional pool is trafficked to the cell surface by a non-conventional, ER-Golgi-independent, actin-dependent route, where it serves as a receptor or co-receptor mediating ligand internalization for endostatin, viral capsids (EV71, RHDV), C1QTNF4, LPS, and apoptotic cells [PMID:11112338, PMID:17615292, PMID:25673703, PMID:30339712, PMID:33676896, PMID:21309751]. Nucleolin abundance and activity are further tuned by additional post-translational modifications, including AMPK-mediated S328 phosphorylation that sequesters it in the nucleus to inhibit miRNA processing, P300-catalyzed K477 lactylation that redirects splicing, and TRIM21-mediated K48 ubiquitination targeting it for proteasomal degradation [PMID:31182601, PMID:38679071, PMID:38357830].","teleology":[{"year":1984,"claim":"Established where nucleolin resides and that it remains tethered to rDNA-bearing structures through the cell cycle, anchoring its connection to ribosomal gene loci.","evidence":"Silver staining and immunoelectron microscopy across interphase and mitosis in hepatoma and PtK2 cells","pmids":["6206987","6345184"],"confidence":"High","gaps":["Localization alone did not define a molecular activity","Did not address what nucleolin binds at NORs"]},{"year":1986,"claim":"Showed nucleolin physically associates with nascent pre-rRNA in preribosomal particles, providing the first direct molecular link to ribosome biogenesis.","evidence":"UV cross-linking and immunoprecipitation with rDNA probe hybridization from purified nucleoli","pmids":["3790520"],"confidence":"High","gaps":["Did not define which RNA-binding domains mediate binding","Processing role not yet functionally demonstrated"]},{"year":1990,"claim":"Resolved nucleolin as a dually phosphorylated protein—CKII on interphase serines and Cdc2 on mitotic TPXK threonines—linking its modification state to cell-cycle context.","evidence":"In vitro CKII and M-phase H1 kinase assays with peptide mapping compared to in vivo 32P labeling","pmids":["3427111","3190709","2192260"],"confidence":"High","gaps":["Functional consequence of each phospho-state not established at the time","Did not link phosphorylation to specific binding partners"]},{"year":1992,"claim":"Genetic disruption of the yeast ortholog NSR1 demonstrated an evolutionarily conserved, essential role in pre-rRNA processing and 40S subunit production.","evidence":"nsr1 deletion with pulse-labeling, Northern blot, and ribosomal subunit gradient analysis in yeast","pmids":["1706724","1508189","1644811","1644812","8245119"],"confidence":"High","gaps":["Mechanism of processing facilitation not defined","Conservation of mammalian processing function inferred rather than directly tested in mammals"]},{"year":1996,"claim":"Mapped a direct interaction between nucleolin and nucleophosmin/B23, defining a core nucleolar protein-protein contact and its responsible domains.","evidence":"Two-hybrid, reciprocal co-IP, and deletion mapping in HeLa extracts; cell-cycle-resolved cross-linking co-IP","pmids":["8620867","10503877"],"confidence":"High","gaps":["Functional output of the NCL-B23 complex not fully defined","Loss of interaction during mitosis not mechanistically explained"]},{"year":2006,"claim":"Demonstrated that nucleolin is specifically required for RNA polymerase I transcription on chromatin and physically associates with rDNA, distinguishing a chromatin-dependent transcription role.","evidence":"In vitro chromatin transcription, ChIP, and siRNA knockdown","pmids":["17130237"],"confidence":"High","gaps":["How nucleolin facilitates elongation on chromatin not resolved","Did not address the histone-mark mechanism"]},{"year":2012,"claim":"Defined the chromatin mechanism: nucleolin antagonizes TTF-1/TIP5/HDAC1-mediated repression to keep rDNA euchromatic and permit RNAP I progression.","evidence":"ChIP-seq, histone mark and RNAP I/UBF distribution analysis with siRNA knockdown","pmids":["22859736"],"confidence":"High","gaps":["Direct competition with TTF-1 for binding sites not structurally resolved","Selectivity for unmethylated genes mechanism unexplained"]},{"year":2007,"claim":"Linked the mitotic Cdc2-phosphorylated pool of nucleolin to spindle organization and chromosome segregation fidelity, extending its role beyond the nucleolus.","evidence":"RNAi depletion with high-resolution microscopy and phospho-specific antibody localization","pmids":["17535846"],"confidence":"High","gaps":["Molecular partners at spindle poles not identified","Mechanism connecting nucleolin to kinetochore-microtubule attachment unknown"]},{"year":2012,"claim":"Established nucleolin as a translational and post-transcriptional regulator beyond rRNA—repressing p53 mRNA via RBD-mediated dimerization reversed by RPL26, and supporting microprocessor-dependent miRNA maturation.","evidence":"Translation/RNA-binding/dimerization assays and domain dissection; in vitro miRNA processing with Drosha/DGCR8 co-IP","pmids":["22433872","22049078"],"confidence":"High","gaps":["In vivo extent of p53 regulation across stress contexts not defined","Range of miRNAs dependent on nucleolin not comprehensively mapped"]},{"year":2000,"claim":"Identified a non-conventional, ER-Golgi-independent, actin-dependent route delivering nucleolin to the cell surface where it functions in ligand internalization, defining an extranuclear functional pool.","evidence":"Confocal/EM imaging with transport-inhibitor and actin-disruption studies plus antibody cross-linking","pmids":["11112338"],"confidence":"High","gaps":["Molecular machinery of the non-conventional export route unknown","How surface targeting is regulated not defined"]},{"year":2023,"claim":"Showed surface nucleolin serves as a receptor/co-receptor for a structurally diverse ligand set, from angiogenesis regulators to viral capsids and innate-immune ligands, internalizing cargo into cells.","evidence":"Binding, knockdown, gain-of-function, domain-mapping, and in vivo studies for endostatin, EV71, RHDV VP60, C1QTNF4, LPS, mtDNA/CpG-DNA, and apoptotic cells","pmids":["17615292","25673703","30339712","33676896","21309751","31412132","16135517"],"confidence":"Medium","gaps":["Identity of obligate coreceptors for most ligands unknown","Whether a single surface conformation accommodates all ligands unresolved"]},{"year":2025,"claim":"Provided atomic-resolution mechanism for G-quadruplex recognition, showing all four RBDs plus the linker and RGG cooperatively engage the MYC promoter G4 in cells, identifying G4s as primary cellular substrates.","evidence":"2.6-Å crystal structure, NMR, binding affinity, and CUT&Tag in cells; preceded by photo-cross-linking loop-length analysis","pmids":["40245140","32191439","11397095"],"confidence":"High","gaps":["Genome-wide functional consequences of G4 binding not fully cataloged","How G4 binding integrates with rDNA transcription and mRNA regulation unresolved"]},{"year":2024,"claim":"Expanded the post-translational control of nucleolin to lysine lactylation, AMPK phosphorylation, and TRIM21 ubiquitination, showing distinct modifications reroute its splicing, miRNA-processing, and stability functions in disease contexts.","evidence":"MS-based PTM mapping with writer-enzyme assays, phospho-mutant rescue, ubiquitination assays, and xenograft models","pmids":["38679071","31182601","38357830","29168431"],"confidence":"Medium","gaps":["Interplay/hierarchy among the different PTMs not defined","Most PTM mechanisms shown in single disease models without broad validation"]},{"year":null,"claim":"How nucleolin's many activities—rDNA transcription, G4 recognition, mRNA regulation, and surface receptor function—are coordinately partitioned and switched by its modification state within a single cell remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model linking PTM state to subcellular pool and substrate choice","Mechanism routing nucleolin between nuclear and surface pools incompletely defined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[2,15,27,37,43,47]},{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[21,26,43,46]},{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[11,21,26]},{"term_id":"GO:0045182","term_label":"translation regulator activity","supporting_discovery_ids":[27,28]},{"term_id":"GO:0001618","term_label":"virus receptor activity","supporting_discovery_ids":[30,35]},{"term_id":"GO:0038024","term_label":"cargo receptor activity","supporting_discovery_ids":[14,23,42]},{"term_id":"GO:0060089","term_label":"molecular transducer activity","supporting_discovery_ids":[19,45,46]}],"localization":[{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[0,1,2]},{"term_id":"GO:0005654","term_label":"nucleoplasm","supporting_discovery_ids":[16,25]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[14,23,30,35,42,45,46]},{"term_id":"GO:0005815","term_label":"microtubule organizing center","supporting_discovery_ids":[22]},{"term_id":"GO:0000228","term_label":"nuclear chromosome","supporting_discovery_ids":[21,26]}],"pathway":[{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[21,26]},{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[2,8,25,27,38]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[6,22]},{"term_id":"R-HSA-4839726","term_label":"Chromatin organization","supporting_discovery_ids":[26]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[45,46]},{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[14,35]}],"complexes":["microprocessor (Drosha/DGCR8)","uPAR-nucleolin-CK2 surface signaling complex"],"partners":["NPM1/B23","TP53","RPL26","DROSHA","DGCR8","HTERT","PARN","TRIM21"],"other_free_text":[]}},"prefetch_data":{"uniprot":{},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"NCL","url":"https://depmap.org/portal/gene/NCL","classification":"Common Essential","n_dependent_lines":1084,"n_total_lines":1208,"dependency_fraction":0.8973509933774835},"opencell":{"profiled":true,"resolved_as":"NCL","ensg_id":"ENSG00000115053","cell_line_id":"CID001030","localizations":[{"compartment":"nucleolus_gc","grade":3},{"compartment":"nucleoplasm","grade":1}],"interactors":[{"gene":"CSNK2A1","stoichiometry":10.0},{"gene":"RPL11","stoichiometry":10.0},{"gene":"RPS11","stoichiometry":10.0},{"gene":"RPS16","stoichiometry":10.0},{"gene":"SRP9","stoichiometry":10.0},{"gene":"CSNK2A2","stoichiometry":4.0},{"gene":"RPS4X","stoichiometry":4.0},{"gene":"RACK1","stoichiometry":4.0},{"gene":"RBM8A","stoichiometry":4.0},{"gene":"RPL4","stoichiometry":4.0}],"url":"https://opencell.sf.czbiohub.org/target/CID001030","total_profiled":1310},"omim":[{"mim_id":"621404","title":"CHROMOSOME 20 OPEN READING FRAME 204; C20ORF204","url":"https://www.omim.org/entry/621404"},{"mim_id":"621385","title":"NUCLEOLAR PROTEIN 12; NOL12","url":"https://www.omim.org/entry/621385"},{"mim_id":"620055","title":"PWP1 HOMOLOG, ENDONUCLEIN; PWP1","url":"https://www.omim.org/entry/620055"},{"mim_id":"619225","title":"RIBOSOMAL PROTEIN L13A; RPL13A","url":"https://www.omim.org/entry/619225"},{"mim_id":"618703","title":"ZINC FINGER PROTEIN 281; ZNF281","url":"https://www.omim.org/entry/618703"}],"hpa":{"profiled":true,"resolved_as":"NCL","reliability":"Enhanced","locations":[{"location":"Nucleoplasm","reliability":"Enhanced"},{"location":"Nucleoli","reliability":"Enhanced"},{"location":"Nucleoli rim","reliability":"Enhanced"},{"location":"Mitotic chromosome","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/NCL"},"hgnc":{"alias_symbol":["C23","Nsr1"],"prev_symbol":["NCL"]},"alphafold":{},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=NUCLEOLIN","jax_strain_url":"https://www.jax.org/strain/search?query=NUCLEOLIN"},"sequence":{}},"corpus_meta":[{"pmid":"10036227","id":"PMC_10036227","title":"Structure and functions of nucleolin.","date":"1999","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/10036227","citation_count":622,"is_preprint":false},{"pmid":"29937225","id":"PMC_29937225","title":"A LINE1-Nucleolin Partnership Regulates Early Development and ESC Identity.","date":"2018","source":"Cell","url":"https://pubmed.ncbi.nlm.nih.gov/29937225","citation_count":420,"is_preprint":false},{"pmid":"10544174","id":"PMC_10544174","title":"Molecular dissection of nucleolin's role in growth and cell proliferation: new insights.","date":"1999","source":"FASEB journal : 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immunofluorescence, immunoelectron microscopy\",\n      \"journal\": \"Chromosoma\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct localization by multiple orthogonal imaging methods, replicated across labs and consistent with subsequent studies\",\n      \"pmids\": [\"6206987\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1983,\n      \"finding\": \"During mitosis, nucleolin (C23) remains associated with disappearing nucleoli through prophase, then reappears in prenucleolar bodies and subsequently the nucleolus during telophase, while B23 follows a distinct pattern; C23 co-localizes with silver-staining NOR proteins throughout mitosis.\",\n      \"method\": \"Dual immunofluorescence with anti-B23 and anti-C23 antibodies in PtK2 cells\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct cell-cycle-stage-resolved localization, consistent with multiple independent studies\",\n      \"pmids\": [\"6345184\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1986,\n      \"finding\": \"Nucleolin (C23) directly associates with rapidly labeled pre-rRNA (18S and 28S sequences) in preribosomal RNP particles, as demonstrated by UV cross-linking and immunoprecipitation from Novikoff hepatoma nucleoli.\",\n      \"method\": \"UV cross-linking, immunoprecipitation with anti-C23 antibody, dot-blot hybridization with rDNA probes, sucrose gradient fractionation\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — UV cross-linking and immunoprecipitation with multiple orthogonal methods in a single study\",\n      \"pmids\": [\"3790520\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1987,\n      \"finding\": \"Nucleolin is a natural preferential substrate for a nucleolar casein kinase II (type NII). The kinase phosphorylates nucleolin in vitro at serine residues in two highly acidic tryptic fragments (A: residues 21–49; C: residues 180–221) near the amino terminus, at canonical CKII consensus sites.\",\n      \"method\": \"Co-purification, in vitro kinase assay, peptide mapping, Km/Vmax analyses\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstituted kinase assay with peptide-level mapping, replicated by multiple independent groups\",\n      \"pmids\": [\"3427111\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1988,\n      \"finding\": \"Phosphopeptide patterns from nucleolin phosphorylated by purified casein kinase II (CKII) in vitro are identical to those from nucleolin isolated from tumor cells grown with [32P]-phosphate, establishing nucleolin as a physiological CKII substrate with ~2 mol phosphate per mol nucleolin.\",\n      \"method\": \"In vitro phosphorylation, partial tryptic digest, comparison of in vitro vs. in vivo phosphopeptide patterns\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct in vitro reconstitution compared to in vivo labeling, consistent with prior work\",\n      \"pmids\": [\"3190709\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1988,\n      \"finding\": \"Microinjection of anti-C23 antibody into Chironomus salivary gland nuclei caused a 2–3.5-fold stimulation of 32P incorporation into 38S pre-rRNA, selectively for pre-rRNA without affecting extranucleolar RNA, indicating nucleolin acts as a negative regulator of rDNA transcription.\",\n      \"method\": \"Microinjection of anti-C23 antibody into nuclei, 32P labeling, RNA extraction and electrophoresis\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct functional perturbation in living cells with specific controls (preimmune serum, absorbed antibody), single lab\",\n      \"pmids\": [\"3169130\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1990,\n      \"finding\": \"Nucleolin is phosphorylated on threonines at TPXK motifs in the amino-terminal domain by p34cdc2 (Cdc2 kinase) specifically during mitosis, distinct from the interphase CKII-mediated serine phosphorylation; the same sites used in vivo during mitosis were phosphorylated by M-phase H1 kinase in vitro.\",\n      \"method\": \"In vitro kinase assay with starfish M-phase H1 kinase, in vivo 32P labeling, phosphoamino acid analysis, peptide mapping\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro kinase assay corroborated by in vivo labeling; sites matched consensus and were mapped at the peptide level\",\n      \"pmids\": [\"2192260\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1991,\n      \"finding\": \"Yeast NSR1 (nucleolin ortholog) specifically binds nuclear localization sequences (NLS), contains two RNA recognition motifs, and is required for normal cell growth; disruption of NSR1 causes severe growth defect, establishing functional conservation with mammalian nucleolin.\",\n      \"method\": \"Affinity purification on NLS peptide column, gene disruption, immunofluorescence, antibody-based cloning\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — biochemical purification plus genetic disruption, replicated and extended by multiple groups\",\n      \"pmids\": [\"1706724\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1992,\n      \"finding\": \"Yeast NSR1 is required for pre-rRNA processing: nsr1 deletion blocks initial 35S pre-rRNA processing, nearly eliminates 20S pre-rRNA, reduces 18S rRNA, and disrupts the free 40S ribosomal subunit pool, demonstrating a conserved role of the nucleolin ortholog in ribosome biogenesis.\",\n      \"method\": \"Pulse-labeling of rRNA, Northern blot, sucrose gradient fractionation of ribosomal subunits, antibiotic sensitivity assay\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods in genetic KO, independently replicated\",\n      \"pmids\": [\"1508189\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1992,\n      \"finding\": \"Yeast NSR1 is required for pre-rRNA processing and ribosome biogenesis: nsr1 deletion strains accumulate 35S pre-rRNA, show slow processing of 35S, impaired 18S rRNA methylation, and a reduced 40S:60S ratio; cold shock greatly exacerbates the processing defect.\",\n      \"method\": \"Pulse-labeling, pulse-chase rRNA analysis, Northern blot, NSR1 cold-shock induction assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic deletion with multiple rRNA processing readouts, replicated independently\",\n      \"pmids\": [\"1644811\", \"1644812\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1993,\n      \"finding\": \"Nucleolar localization of NSR1 requires a bipartite NLS and is mediated redundantly by either the N-terminal acidic/serine-rich domain or both RNA recognition motifs (RRMs); point mutations in RNP consensus octamers cause nuclear mislocalization, and the RGG domain is necessary for nucleolar accumulation when one RNP octamer is mutated.\",\n      \"method\": \"Deletion analysis with beta-galactosidase fusion reporter, immunofluorescence, point mutagenesis\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — systematic deletion and point mutation analysis with reporter fusions, single lab\",\n      \"pmids\": [\"8245119\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1994,\n      \"finding\": \"Nucleolin functions as a transcriptional repressor of the acute-phase response gene alpha-1 acid glycoprotein (AGP); nucleolin was purified from mouse hepatoma cells as the B-motif-binding factor and biochemical studies confirmed its repressor activity.\",\n      \"method\": \"Affinity purification, amino acid sequence analysis, transcription assay, DNA-binding assay\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — protein purification combined with functional transcription assay, single lab\",\n      \"pmids\": [\"8065340\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1996,\n      \"finding\": \"Nucleolin (C23) interacts with nucleolar protein B23 (nucleophosmin); co-immunoprecipitation from HeLa nuclear extract with either anti-C23 or anti-B23 monoclonal antibodies confirmed the interaction, which requires residues 540–628 of C23 (the nucleolar localization region) and residues 194–239 of B23.\",\n      \"method\": \"Two-hybrid system, co-immunoprecipitation, deletion mutant binding studies\",\n      \"journal\": \"European journal of biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — orthogonal methods (two-hybrid + reciprocal co-IP + deletion mapping), replicated\",\n      \"pmids\": [\"8620867\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"B23 (nucleophosmin) and nucleolin (C23) interact in vivo during interphase and cytokinesis but not during prometaphase/metaphase; the interaction persists even after actinomycin D-induced translocation to the nucleoplasm, and mitotic phosphorylation of B23 alone does not explain loss of interaction during mitosis.\",\n      \"method\": \"Chemical cross-linking with DSP, co-immunoprecipitation, in vitro Cdc2 kinase phosphorylation of GST-B23 followed by co-IP\",\n      \"journal\": \"Cancer letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — cross-linking co-IP with cell-cycle synchronization and in vitro kinase experiment, single lab\",\n      \"pmids\": [\"10503877\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"Cell-surface nucleolin is transported to the plasma membrane via an active, non-conventional pathway independent of the ER-Golgi complex; surface nucleolin clusters in an actin cytoskeleton-dependent manner upon antibody cross-linking and can be internalized, mediating intracellular import of ligands.\",\n      \"method\": \"Confocal and electron microscopy, antibody cross-linking experiments, inhibitor studies (low temperature, serum-free medium, glycoprotein transport inhibitors), actin disruption\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal localization and functional inhibition methods in a single study, replicated by other groups\",\n      \"pmids\": [\"11112338\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"Nucleolin RBD1 and RBD2 together (RBD12), but neither alone, specifically recognize a stem-loop NRE (nucleolin recognition element) RNA structure; in the complex, the hairpin loop adopts a well-defined conformation distinct from the free RNA, with both loop E motif and hairpin loop interacting specifically with the protein.\",\n      \"method\": \"NMR spectroscopy of free and protein-bound RNA, titration of RBD1, RBD2, and RBD12 with sNRE\",\n      \"journal\": \"Journal of molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — NMR structure determination with functional binding titrations, single lab but rigorous\",\n      \"pmids\": [\"11397095\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Following cell stress (ionizing radiation, camptothecin, heat shock), nucleolin relocalizes from the nucleolus to the nucleoplasm in a p53-dependent manner; the p53 C-terminal regulatory domain is required for nucleolin-p53 complex formation and nucleolin mobilization, which is independent of p53 transactivation; nucleolin and p53 interact directly in vitro.\",\n      \"method\": \"Immunofluorescence, co-immunoprecipitation, in vitro binding assay with deletion mutants, p53-null and mutant cell lines\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct in vitro interaction confirmed, in vivo relocalization with genetic requirement for p53 C-terminal domain, multiple orthogonal methods\",\n      \"pmids\": [\"12138209\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Activation of nucleolin RNA-binding activity after genotoxic stress (UV or ionizing radiation) is mediated by the stress-activated protein kinase p38; nucleolin was identified as a genotoxic stress-responsive RNA-binding protein that binds stress-responsive mRNAs.\",\n      \"method\": \"Purification, RNA binding assays, p38 inhibitor treatment, identification of 40 mRNA ligands\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biochemical purification with RNA binding assays and kinase inhibitor studies, single lab\",\n      \"pmids\": [\"12000845\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Nucleolin interacts with the telomerase reverse transcriptase subunit hTERT through its RNA-binding domain 4 and RGG domain, with the interaction also involving hTERC (telomerase RNA); this nucleolin-hTERT interaction is critical for nucleolar localization of hTERT.\",\n      \"method\": \"Co-immunoprecipitation, domain deletion mapping, immunofluorescence colocalization\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — domain-mapped co-IP with functional localization readout, single lab\",\n      \"pmids\": [\"15371412\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Nucleolin functions as a macrophage cell-surface receptor for early apoptotic cells bearing polylactosaminyl CD43; anti-nucleolin antibody or deletion of the antibody-binding region abolishes binding to early apoptotic cells; nucleolin-transfected HEK293 cells acquire ability to bind early apoptotic cells.\",\n      \"method\": \"Antibody blocking, nucleolin transfection of HEK293, deletion mutant analysis, competitive inhibition with oligosaccharides\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — gain-of-function transfection plus deletion analysis and competitive inhibition, single lab\",\n      \"pmids\": [\"16135517\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Extranuclear nucleolin undergoes complex N- and O-glycosylations; N-glycosylation sites were mapped to N317 and N492 within RNA-binding domains 1 and 3, respectively, suggesting glycosylation may regulate RNA-binding function.\",\n      \"method\": \"SDS-PAGE, mass spectrometry, tunicamycin treatment, lectin binding, exoglycosidase digestion, MALDI-TOF, monosaccharide composition analysis\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — multiple orthogonal biochemical methods including MS-based glycan characterization with site identification, single lab\",\n      \"pmids\": [\"15823039\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Nucleolin is required for RNA polymerase I transcription of chromatin (but not naked DNA) templates in vitro, and is specifically associated by ChIP with rRNA genes transcribed by RNAP I but not RNAP II or III; siRNA knockdown of nucleolin specifically inhibits RNAP I transcription.\",\n      \"method\": \"In vitro chromatin transcription assay, ChIP, immunofluorescence, siRNA knockdown\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution on chromatin templates, ChIP, and RNAi knockdown with multiple orthogonal readouts\",\n      \"pmids\": [\"17130237\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Nucleolin depletion by RNAi results in disorganized nucleoli at interphase, prolonged cell cycle with misaligned chromosomes, syntelic kinetochore-microtubule attachments with reduced centromere stretching, and defects in spindle organization; during mitosis, CDC2-phosphorylated nucleolin associates with spindle poles from prometaphase to anaphase.\",\n      \"method\": \"RNAi depletion, high-resolution microscopy, antibody specific for CDC2-phosphorylated nucleolin, kinetochore-microtubule attachment analysis\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean RNAi depletion with specific mitotic phenotypes and phospho-specific localization, single lab with multiple orthogonal readouts\",\n      \"pmids\": [\"17535846\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Endostatin specifically binds cell-surface nucleolin with high affinity; blockage or knockdown of nucleolin abolishes endostatin's antiendothelial activity in vitro and antiangiogenic/antitumor activity in vivo; endostatin is internalized into cell nuclei via nucleolin; endostatin inhibits mitosis-related phosphorylation of nucleolin in the nucleus.\",\n      \"method\": \"Binding assay, neutralizing antibody, siRNA knockdown, colocalization, internalization assay, in vivo tumor model\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods including in vivo validation and mechanistic phosphorylation studies, single lab\",\n      \"pmids\": [\"17615292\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Urokinase (uPA) induces formation of a signaling complex on the cell surface containing uPAR, nucleolin, and casein kinase 2; nucleolin and CK2 were isolated by affinity chromatography and co-localized with uPAR; uPA activates CK2 in this complex leading to nucleolin phosphorylation; blocking nucleolin or CK2 inhibits uPA-induced cell proliferation.\",\n      \"method\": \"Affinity chromatography, nano-electrospray MS, immunoblotting, laser scanning and immunoelectron microscopy, co-immunoprecipitation, in vitro kinase assay, cell proliferation assay\",\n      \"journal\": \"Current biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — affinity isolation, MS identification, co-IP, in vitro kinase assay, and functional inhibition studies with multiple orthogonal methods\",\n      \"pmids\": [\"10607589\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Nucleolin interacts with the miRNA microprocessor components DGCR8 and Drosha in the nucleus; nucleolin directly and specifically binds the primary miR-15a/16 transcript; nucleolin is required for primary-to-precursor miRNA processing of miR-15a/16 in vitro; nuclear localization of nucleolin is critical for this function.\",\n      \"method\": \"Overexpression and knockdown studies, in vitro processing assay with cell extracts, co-immunoprecipitation, direct RNA binding assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro processing assay with rescue by nucleolin addition, plus direct binding and co-IP, single lab\",\n      \"pmids\": [\"22049078\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Nucleolin depletion increases H3K9me2 (heterochromatin mark) and decreases H4K12Ac and H3K4me3 (euchromatin marks) at rRNA genes; ChIP-seq shows nucleolin enrichment at rDNA coding and promoter regions, preferentially at unmethylated genes; nucleolin depletion causes RNAP I accumulation at the start of transcription units and decreased UBF along the gene; nucleolin antagonizes binding of TTF-1 at the T0 terminator, thereby inhibiting TIP5 and HDAC1 recruitment and repressive heterochromatin formation.\",\n      \"method\": \"ChIP-seq, ChIP, siRNA knockdown, histone mark analysis, RNAP I and UBF distribution analysis\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome-scale ChIP-seq with functional siRNA knockdown and multiple chromatin mark readouts, single lab with rigorous controls\",\n      \"pmids\": [\"22859736\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Nucleolin suppresses p53 mRNA translation using both 5'- and 3'-UTRs; nucleolin binds the same 5'-3'-UTR interaction region critical for RPL26 recruitment; nucleolin oligomerizes (dimerizes) through its RNA-binding domain; RPL26 disrupts NCL dimerization; NCL's RNA-binding domain mediates both dimerization and translational repression, and is the domain that interacts with RPL26.\",\n      \"method\": \"Translation assays, RNA binding assays, co-immunoprecipitation, domain deletion and point mutation analysis, dimerization assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — domain-level dissection with multiple orthogonal methods and functional rescue experiments, single lab\",\n      \"pmids\": [\"22433872\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Nucleolin binds LINE-1 ORF2 IRES and functions as an IRES trans-acting factor (ITAF) for ORF2 translation; NCL knockdown specifically reduces ORF2 IRES activity, L1 and L1-assisted Alu retrotransposition without altering L1 RNA or ORF1 protein abundance.\",\n      \"method\": \"RNA affinity chromatography, RNAi knockdown, IRES activity assay, retrotransposition assay\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — RNA affinity chromatography plus functional IRES/retrotransposition assays, single lab\",\n      \"pmids\": [\"23161687\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"HuR promotes nucleolin translation via the nucleolin 3'UTR without affecting mRNA levels; miR-494 inhibits nucleolin expression by enhancing NCL mRNA association with Argonaute-containing complexes and promoting NCL mRNA transport to processing bodies (PBs); HuR and miR-494 functionally compete to control nucleolin abundance.\",\n      \"method\": \"MS2-tagged 3'UTR pull-down, immunoprecipitation, HuR silencing, miR-494 transfection, P-body tracking, ribosome loading assay\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods in a single study, single lab\",\n      \"pmids\": [\"21859890\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Cell-surface nucleolin mediates EV71 binding and infection; EV71 interacts directly with nucleolin via the VP1 capsid protein; knockdown of nucleolin reduces EV71 binding, infection, and production; expression of human nucleolin on mouse cells confers EV71 infection.\",\n      \"method\": \"Glycoproteomics, immunoprecipitation with EV71 particles, anti-nucleolin antibody blocking, siRNA knockdown, gain-of-function expression in mouse cells\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods including gain-of-function and knockdown with multiple viral readouts, single lab\",\n      \"pmids\": [\"25673703\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Nucleolin depletion reduces axonal levels of importin β1 mRNA and protein; subcellular sequestration of nucleolin or importin β1 enhances axonal growth and causes a subcellular shift in protein synthesis; nucleolin associates with importin β1 mRNA in axons and with kinesins for anterograde transport.\",\n      \"method\": \"RNAi/siRNA knockdown, subcellular fractionation, in situ hybridization, cell growth assays in neurons and fibroblasts\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple cell types with knockdown and localization experiments, single lab\",\n      \"pmids\": [\"27477284\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"NCL phosphorylation at six CK2 consensus sites in the N-terminus is required for activation of PARN deadenylase activity upon oncogenic stimuli and UV stress; NCL directly interacts with PARN; under non-stress conditions NCL forms complexes with p53 and HuR; phosphorylation state of NCL determines specificity of its protein-protein and protein-RNA interactions governing mRNA deadenylation.\",\n      \"method\": \"In vitro deadenylase assay, co-immunoprecipitation, phospho-mutant (NCL-6/S*A) analysis, RNA binding assay\",\n      \"journal\": \"RNA biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro PARN assay with phosphomutant and co-IP, single lab\",\n      \"pmids\": [\"29168431\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"LINE1 RNA recruits nucleolin and Kap1/Trim28 to repress Dux (master activator of the 2-cell program) in ESCs; LINE1 RNA also mediates binding of nucleolin and Kap1 to rDNA, promoting rRNA synthesis; in pre-implantation embryos, LINE1 RNA is required for Dux silencing, rRNA synthesis, and exit from the 2-cell stage.\",\n      \"method\": \"RNA immunoprecipitation, ChIP, siRNA/shRNA knockdown, RNA FISH, embryo functional assays\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal IP, ChIP, and functional embryo assays with multiple orthogonal methods\",\n      \"pmids\": [\"29937225\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Binding of cellular nucleolin to the HCV core RNA G-quadruplex (G4) structure stabilizes the G4 and suppresses HCV replication; NCL co-localizes with HCV particles; HCV infection upregulates NCL; silencing NCL greatly enhances viral RNA replication.\",\n      \"method\": \"Direct binding assay (in vitro and in-cell), colocalization, NCL knockdown with viral replication assay, G4-mutant virus comparison\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding plus functional knockdown assay with G4-mutant virus control, single lab\",\n      \"pmids\": [\"30462330\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Nucleolin mediates RHDV internalization via clathrin-dependent endocytosis; NCL interacts specifically with RHDV capsid protein VP60 through NCL N-terminal residues 285–318 and VP60's DVN motif (472Asp-Val-Asn474); NCL also interacts with clathrin light chain A C-terminus; blocking the NCL-VP60 interaction with a DVN peptide markedly reduces virus internalization.\",\n      \"method\": \"Biochemical inhibitors, RNAi, co-immunoprecipitation, domain deletion analysis, competitive peptide inhibition, in vivo animal experiments\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — domain-mapped co-IP, functional blocking with specific peptide, and in vivo animal validation\",\n      \"pmids\": [\"30339712\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Shear stress-activated AMPK phosphorylates nucleolin at serine 328 (S328), which sequesters nucleolin in the nucleus and inhibits processing of miR-93 and miR-484 that target KLF2 and eNOS mRNAs; this identifies an AMPK-NCL-miR-93/miR-484 axis in endothelial mechanotransduction.\",\n      \"method\": \"In silico analysis, phosphorylation site mapping (S328), AMPK inhibitor and activator studies, anti-miR rescue experiments, posttranslational modification analysis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — phospho-site identified with functional rescue experiments, single lab\",\n      \"pmids\": [\"31182601\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Nucleolin strongly and preferentially binds G-quadruplex structures with long central loops (5–9 nucleotides) over short-loop variants; photo-cross-linking identified a 15-amino acid fragment in helix α2 of RBD2 as a loop-contact site; the RGG domain participates in terminal guanine quartet recognition; thus NCL recognizes G4 via concerted action of RBD2 (loop) and RGG (quartet) domains.\",\n      \"method\": \"Binding assays, photo-cross-linking with BrU-modified G4 sequences, quantitative proteomics (LC-MS/MS), G4 ligand competition assays\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — photo-cross-linking with MS-based site mapping plus systematic loop-length and ligand competition experiments, single lab\",\n      \"pmids\": [\"32191439\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"A cysteine tRNA-derived fragment (5'-tRFCys) directly binds nucleolin and promotes its oligomerization into a higher-order ribonucleoprotein complex that stabilizes bound metabolic mRNAs (Mthfd1l and Pafah1b1) against exonucleolytic degradation, driving breast cancer metastasis.\",\n      \"method\": \"Small RNA profiling, direct binding assay, oligomerization assay, RNA stability assay, functional metastasis assays\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct binding established, oligomerization and mRNA stability mechanistically linked, functional metastasis readout, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"35654044\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Nucleolin directly activates NFκB signaling; nucleolin overexpression increases NFκB phosphorylation and upregulates DNMT1; NFκB inactivation diminishes DNMT1 promoter activity, establishing a nucleolin-NFκB-DNMT1 axis in leukemia.\",\n      \"method\": \"Forced expression and RNAi knockdown, NFκB reporter assay, DNMT1 promoter activity assay, Western blot, in vitro and in vivo proliferation/tumorigenesis assays\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional assays with promoter reporter and genetic knockdown/overexpression, single lab\",\n      \"pmids\": [\"25015109\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Nucleolin identifies nucleolin as binding partner of topoisomerase II-alpha (TopIIA); this interaction is mapped to RNA-binding domain 3 of nucleolin and is essential for blocking DNA damage and apoptosis; nucleolin silencing decreases TopIIA decatenation activity and enhances formation of TopIIA-DNA cleavable complexes in the presence of etoposide.\",\n      \"method\": \"Co-immunoprecipitation, domain deletion mapping, TopIIA decatenation assay, cleavable complex assay, siRNA knockdown with DNA damage assays\",\n      \"journal\": \"Leukemia\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — domain-mapped co-IP with enzymatic activity assay, single lab\",\n      \"pmids\": [\"28690315\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Nucleolin is lactylated predominantly at lysine K477 by the acyltransferase P300 in response to glycolytic hyperactivity; K477 lactylation promotes efficient translation of MADD mRNA by preventing alternative splicing that generates a premature termination codon; the resulting MADD protein activates ERK signaling and drives iCCA tumor growth.\",\n      \"method\": \"Proteomics, mass spectrometry, macromolecule interaction studies, P300 acyltransferase assay, alternative splicing analysis, xenograft tumor model, clinical sample validation\",\n      \"journal\": \"Journal of hepatology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — site-specific PTM identified by MS with P300 as writer enzyme, mechanistic link to splicing and ERK signaling validated in vivo, single lab with multiple methods\",\n      \"pmids\": [\"38679071\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Nucleolin acts as a cell-surface receptor for C1QTNF4; interaction is mediated by the second C1q-like domain of C1QTNF4 and the C-terminus of nucleolin; upon cell binding, C1QTNF4 is actively internalized in monocytes; nucleolin may act as a docking molecule with context-dependent coreceptors.\",\n      \"method\": \"Mass spectrometric analysis of cell-surface proteins, co-immunoprecipitation, domain deletion analysis, imaging flow cytometry internalization assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — domain-mapped MS identification plus co-IP and internalization imaging, single lab\",\n      \"pmids\": [\"33676896\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Crystal structure (2.6 Å) of nucleolin bound to the MYC promoter G-quadruplex (MycG4) reveals: MycG4 adopts a folded parallel three-tetrad structure with two K+ ions; RBD1, RBD2, and Linker12 interact with the 6-nt central loop and 5' flanking region; RBD3 and RBD4 bind the 1-nt loops (confirmed by NMR); all four RBDs are required for high-affinity MycG4 binding; CUT&Tag sequencing confirmed nucleolin binding to MycG4 in cells, suggesting G4s are primary cellular substrates of nucleolin.\",\n      \"method\": \"X-ray crystallography (2.6 Å), NMR spectroscopy, CUT&Tag sequencing, binding affinity measurements\",\n      \"journal\": \"Science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — atomic-resolution crystal structure plus NMR validation plus in-cell chromatin binding, multiple orthogonal methods in a single rigorous study\",\n      \"pmids\": [\"40245140\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"The E3 ubiquitin ligase TRIM21 mediates K48-linked polyubiquitination and proteasomal degradation of nucleolin; circRNA circ0006646 protects nucleolin from TRIM21-mediated ubiquitination by preventing the TRIM21-NCL interaction.\",\n      \"method\": \"Multi-omics proteomics, co-immunoprecipitation, ubiquitination assay, proteasome inhibitor treatment, in vivo xenograft and lentivirus models\",\n      \"journal\": \"Advanced science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ubiquitination assay with TRIM21 as E3 ligase identified, K48-linkage specified, but findings come from a circRNA-focused study; single lab\",\n      \"pmids\": [\"38357830\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Cell-surface nucleolin on alveolar macrophages directly binds LPS via affinity chromatography; LPS co-localizes with C23 on cell surface and in cytoplasm; siRNA knockdown of C23 reduces LPS internalization, LPS-induced NF-κB DNA binding, and TNF-α and IL-6 protein expression.\",\n      \"method\": \"LPS affinity chromatography, immunofluorescence colocalization, siRNA knockdown, NF-κB DNA binding assay, cytokine measurement\",\n      \"journal\": \"Cell biology international\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding demonstrated with functional siRNA knockdown, single lab\",\n      \"pmids\": [\"21309751\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Nucleolin on the plasma membrane of cardiomyocytes binds mitochondrial DNA (mtDNA) with ~10-fold greater affinity than nuclear DNA; blocking nucleolin with midkine reduces IL-1β/TNF-α expression and inhibits cellular uptake of CpG-DNA; nucleolin inhibition with AS1411 reduces IL-6 release; nucleolin facilitates CpG-DNA internalization independently of classical endocytosis.\",\n      \"method\": \"Microscale thermophoresis (direct binding), immunofluorescence, nucleolin inhibitor (midkine and AS1411) studies, CpG-DNA uptake assay, cytokine measurement\",\n      \"journal\": \"British journal of pharmacology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding measured with MST plus functional inhibition studies, single lab\",\n      \"pmids\": [\"31412132\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"G-quadruplex structures in MALAT1 lncRNA (in the 3' region) are specifically bound by nucleolin and nucleophosmin; G4-disrupting G-to-A mutations in MALAT1 abolish localization of both NCL and NPM to nuclear speckles; truncated NCL (ΔNCL) binds all three MALAT1 rG4s with high affinity in vitro.\",\n      \"method\": \"rG4 domain-specific RNA pull-down, mass spectrometry, RNA immunoprecipitation, imaging, G-to-A mutagenesis, in vitro biophysical binding studies\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — RNA pull-down with MS identification, mutagenesis functional readout, in vitro binding; single lab\",\n      \"pmids\": [\"37558241\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"Nucleolin (NCL/C23) is a multidomain nucleolar phosphoprotein whose tripartite structure (N-terminal acidic/serine domain, four RNA-binding domains RBD1–4, and C-terminal RGG domain) enables it to: (1) bind pre-rRNA and chromatin at rDNA loci to facilitate RNA polymerase I transcriptional elongation and maintain a euchromatic rDNA state by antagonizing TTF-1/TIP5/HDAC1-mediated repression; (2) participate in pre-rRNA processing and ribosome assembly (conserved from yeast NSR1 to mammals); (3) undergo cell-cycle-regulated phosphorylation—by CKII at serine residues during interphase (promoting rDNA transcription) and by Cdc2/CDK1 at threonine TPXK motifs during mitosis (linked to nucleolar chromatin reorganization); (4) recognize G-quadruplex structures (including the MYC promoter G4, revealed by a 2.6-Å crystal structure) through concerted action of RBD1–4 and the RGG domain; (5) suppress p53 mRNA translation by binding 5'–3'-UTR interaction regions and forming homodimers that are disrupted by RPL26 after DNA damage; (6) interact with LINE1 RNA as a nuclear scaffold to repress Dux and promote rRNA synthesis in ESCs; (7) translocate to the cell surface via a non-conventional, ER-Golgi-independent, actin-dependent pathway where it acts as a receptor/co-receptor for diverse ligands (endostatin, EV71, RHDV VP60, C1QTNF4, LPS, apoptotic cells) mediating internalization; and (8) be post-translationally regulated by CKII-mediated phosphorylation (controlling PARN deadenylase activation and mRNA stability), p38-mediated activation of RNA-binding after genotoxic stress, AMPK-mediated S328 phosphorylation (sequestering NCL in the nucleus to inhibit miRNA processing), P300-catalyzed K477 lactylation (redirecting RNA splicing to upregulate MADD/ERK), and TRIM21-mediated K48-ubiquitination leading to proteasomal degradation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"Nucleolin (NCL/C23) is a multidomain nucleolar phosphoprotein that couples ribosomal DNA transcription, pre-rRNA processing, and ribosome assembly to a broad set of nucleic-acid recognition activities through its tandem RNA-binding domains (RBD1–4) and C-terminal RGG domain [#2, #15, #43]. It localizes to the fibrillar regions of the nucleolus during interphase and stays associated with NOR/rDNA structures throughout mitosis [#0, #1], where it binds pre-rRNA in preribosomal RNP particles [#2]; the conserved yeast ortholog NSR1 is required for 35S pre-rRNA processing and 40S subunit formation, establishing this ribosome-biogenesis role across eukaryotes [#7, #8, #9]. At rDNA, nucleolin is selectively required for RNA polymerase I transcription of chromatin templates and maintains a euchromatic, transcriptionally active state by antagonizing TTF-1 binding and the recruitment of TIP5/HDAC1-mediated repressive heterochromatin [#21, #26]. Its activity is governed by cell-cycle-coupled phosphorylation—by casein kinase II on N-terminal acidic-domain serines during interphase and by Cdc2/CDK1 on TPXK threonine motifs during mitosis, the latter directing nucleolin to spindle poles and proper kinetochore-microtubule attachment [#3, #4, #6, #22]. Through concerted action of its RBD and RGG domains, nucleolin recognizes G-quadruplex structures, demonstrated at atomic resolution for the MYC promoter G4, where all four RBDs engage the folded quadruplex [#37, #43]. Beyond rRNA, nucleolin regulates gene expression post-transcriptionally: it represses p53 mRNA translation by binding a 5'–3'-UTR interaction region and homodimerizing through its RNA-binding domain, an interaction disrupted by RPL26 [#27]; it participates in microprocessor-dependent miRNA maturation with Drosha/DGCR8 [#25]; and it stabilizes bound mRNAs as part of higher-order ribonucleoprotein assemblies [#38]. A second functional pool is trafficked to the cell surface by a non-conventional, ER-Golgi-independent, actin-dependent route, where it serves as a receptor or co-receptor mediating ligand internalization for endostatin, viral capsids (EV71, RHDV), C1QTNF4, LPS, and apoptotic cells [#14, #23, #30, #35, #42, #45]. Nucleolin abundance and activity are further tuned by additional post-translational modifications, including AMPK-mediated S328 phosphorylation that sequesters it in the nucleus to inhibit miRNA processing, P300-catalyzed K477 lactylation that redirects splicing, and TRIM21-mediated K48 ubiquitination targeting it for proteasomal degradation [#36, #41, #44].\",\n  \"teleology\": [\n    {\n      \"year\": 1984,\n      \"claim\": \"Established where nucleolin resides and that it remains tethered to rDNA-bearing structures through the cell cycle, anchoring its connection to ribosomal gene loci.\",\n      \"evidence\": \"Silver staining and immunoelectron microscopy across interphase and mitosis in hepatoma and PtK2 cells\",\n      \"pmids\": [\"6206987\", \"6345184\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Localization alone did not define a molecular activity\", \"Did not address what nucleolin binds at NORs\"]\n    },\n    {\n      \"year\": 1986,\n      \"claim\": \"Showed nucleolin physically associates with nascent pre-rRNA in preribosomal particles, providing the first direct molecular link to ribosome biogenesis.\",\n      \"evidence\": \"UV cross-linking and immunoprecipitation with rDNA probe hybridization from purified nucleoli\",\n      \"pmids\": [\"3790520\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define which RNA-binding domains mediate binding\", \"Processing role not yet functionally demonstrated\"]\n    },\n    {\n      \"year\": 1990,\n      \"claim\": \"Resolved nucleolin as a dually phosphorylated protein—CKII on interphase serines and Cdc2 on mitotic TPXK threonines—linking its modification state to cell-cycle context.\",\n      \"evidence\": \"In vitro CKII and M-phase H1 kinase assays with peptide mapping compared to in vivo 32P labeling\",\n      \"pmids\": [\"3427111\", \"3190709\", \"2192260\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional consequence of each phospho-state not established at the time\", \"Did not link phosphorylation to specific binding partners\"]\n    },\n    {\n      \"year\": 1992,\n      \"claim\": \"Genetic disruption of the yeast ortholog NSR1 demonstrated an evolutionarily conserved, essential role in pre-rRNA processing and 40S subunit production.\",\n      \"evidence\": \"nsr1 deletion with pulse-labeling, Northern blot, and ribosomal subunit gradient analysis in yeast\",\n      \"pmids\": [\"1706724\", \"1508189\", \"1644811\", \"1644812\", \"8245119\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism of processing facilitation not defined\", \"Conservation of mammalian processing function inferred rather than directly tested in mammals\"]\n    },\n    {\n      \"year\": 1996,\n      \"claim\": \"Mapped a direct interaction between nucleolin and nucleophosmin/B23, defining a core nucleolar protein-protein contact and its responsible domains.\",\n      \"evidence\": \"Two-hybrid, reciprocal co-IP, and deletion mapping in HeLa extracts; cell-cycle-resolved cross-linking co-IP\",\n      \"pmids\": [\"8620867\", \"10503877\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional output of the NCL-B23 complex not fully defined\", \"Loss of interaction during mitosis not mechanistically explained\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Demonstrated that nucleolin is specifically required for RNA polymerase I transcription on chromatin and physically associates with rDNA, distinguishing a chromatin-dependent transcription role.\",\n      \"evidence\": \"In vitro chromatin transcription, ChIP, and siRNA knockdown\",\n      \"pmids\": [\"17130237\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How nucleolin facilitates elongation on chromatin not resolved\", \"Did not address the histone-mark mechanism\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Defined the chromatin mechanism: nucleolin antagonizes TTF-1/TIP5/HDAC1-mediated repression to keep rDNA euchromatic and permit RNAP I progression.\",\n      \"evidence\": \"ChIP-seq, histone mark and RNAP I/UBF distribution analysis with siRNA knockdown\",\n      \"pmids\": [\"22859736\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct competition with TTF-1 for binding sites not structurally resolved\", \"Selectivity for unmethylated genes mechanism unexplained\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Linked the mitotic Cdc2-phosphorylated pool of nucleolin to spindle organization and chromosome segregation fidelity, extending its role beyond the nucleolus.\",\n      \"evidence\": \"RNAi depletion with high-resolution microscopy and phospho-specific antibody localization\",\n      \"pmids\": [\"17535846\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular partners at spindle poles not identified\", \"Mechanism connecting nucleolin to kinetochore-microtubule attachment unknown\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Established nucleolin as a translational and post-transcriptional regulator beyond rRNA—repressing p53 mRNA via RBD-mediated dimerization reversed by RPL26, and supporting microprocessor-dependent miRNA maturation.\",\n      \"evidence\": \"Translation/RNA-binding/dimerization assays and domain dissection; in vitro miRNA processing with Drosha/DGCR8 co-IP\",\n      \"pmids\": [\"22433872\", \"22049078\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo extent of p53 regulation across stress contexts not defined\", \"Range of miRNAs dependent on nucleolin not comprehensively mapped\"]\n    },\n    {\n      \"year\": 2000,\n      \"claim\": \"Identified a non-conventional, ER-Golgi-independent, actin-dependent route delivering nucleolin to the cell surface where it functions in ligand internalization, defining an extranuclear functional pool.\",\n      \"evidence\": \"Confocal/EM imaging with transport-inhibitor and actin-disruption studies plus antibody cross-linking\",\n      \"pmids\": [\"11112338\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular machinery of the non-conventional export route unknown\", \"How surface targeting is regulated not defined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Showed surface nucleolin serves as a receptor/co-receptor for a structurally diverse ligand set, from angiogenesis regulators to viral capsids and innate-immune ligands, internalizing cargo into cells.\",\n      \"evidence\": \"Binding, knockdown, gain-of-function, domain-mapping, and in vivo studies for endostatin, EV71, RHDV VP60, C1QTNF4, LPS, mtDNA/CpG-DNA, and apoptotic cells\",\n      \"pmids\": [\"17615292\", \"25673703\", \"30339712\", \"33676896\", \"21309751\", \"31412132\", \"16135517\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Identity of obligate coreceptors for most ligands unknown\", \"Whether a single surface conformation accommodates all ligands unresolved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Provided atomic-resolution mechanism for G-quadruplex recognition, showing all four RBDs plus the linker and RGG cooperatively engage the MYC promoter G4 in cells, identifying G4s as primary cellular substrates.\",\n      \"evidence\": \"2.6-Å crystal structure, NMR, binding affinity, and CUT&Tag in cells; preceded by photo-cross-linking loop-length analysis\",\n      \"pmids\": [\"40245140\", \"32191439\", \"11397095\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Genome-wide functional consequences of G4 binding not fully cataloged\", \"How G4 binding integrates with rDNA transcription and mRNA regulation unresolved\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Expanded the post-translational control of nucleolin to lysine lactylation, AMPK phosphorylation, and TRIM21 ubiquitination, showing distinct modifications reroute its splicing, miRNA-processing, and stability functions in disease contexts.\",\n      \"evidence\": \"MS-based PTM mapping with writer-enzyme assays, phospho-mutant rescue, ubiquitination assays, and xenograft models\",\n      \"pmids\": [\"38679071\", \"31182601\", \"38357830\", \"29168431\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Interplay/hierarchy among the different PTMs not defined\", \"Most PTM mechanisms shown in single disease models without broad validation\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How nucleolin's many activities—rDNA transcription, G4 recognition, mRNA regulation, and surface receptor function—are coordinately partitioned and switched by its modification state within a single cell remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified model linking PTM state to subcellular pool and substrate choice\", \"Mechanism routing nucleolin between nuclear and surface pools incompletely defined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [2, 15, 27, 37, 43, 47]},\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [21, 26, 43, 46]},\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [11, 21, 26]},\n      {\"term_id\": \"GO:0045182\", \"supporting_discovery_ids\": [27, 28]},\n      {\"term_id\": \"GO:0001618\", \"supporting_discovery_ids\": [30, 35]},\n      {\"term_id\": \"GO:0038024\", \"supporting_discovery_ids\": [14, 23, 42]},\n      {\"term_id\": \"GO:0060089\", \"supporting_discovery_ids\": [19, 45, 46]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [0, 1, 2]},\n      {\"term_id\": \"GO:0005654\", \"supporting_discovery_ids\": [16, 25]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [14, 23, 30, 35, 42, 45, 46]},\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [22]},\n      {\"term_id\": \"GO:0000228\", \"supporting_discovery_ids\": [21, 26]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [21, 26]},\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [2, 8, 25, 27, 38]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [6, 22]},\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [26]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [45, 46]},\n      {\"term_id\": \"R-HSA-5653656\", \"supporting_discovery_ids\": [14, 35]}\n    ],\n    \"complexes\": [\n      \"microprocessor (Drosha/DGCR8)\",\n      \"uPAR-nucleolin-CK2 surface signaling complex\"\n    ],\n    \"partners\": [\n      \"NPM1/B23\",\n      \"TP53\",\n      \"RPL26\",\n      \"DROSHA\",\n      \"DGCR8\",\n      \"hTERT\",\n      \"PARN\",\n      \"TRIM21\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"faith_supported":7,"faith_total":8,"faith_pct":87.5}}