{"gene":"AATF","run_date":"2026-06-09T22:02:36","timeline":{"discoveries":[{"year":1999,"finding":"AATF (apoptosis antagonizing transcription factor) was identified as a binding partner of Dlk/ZIP kinase (a serine/threonine kinase) via yeast two-hybrid and shown to interfere with Dlk-induced apoptosis. AATF contains an acidic domain and putative leucine zipper and exhibits transcriptional transactivation activity as a Gal4-BD fusion protein.","method":"Yeast two-hybrid interaction screen, transactivation assay (Gal4-BD fusion), apoptosis interference assay","journal":"FEBS letters","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — yeast two-hybrid plus functional transactivation assay and apoptosis assay, single lab, two complementary methods","pmids":["10580117"],"is_preprint":false},{"year":2000,"finding":"Che-1/AATF interacts with RNA polymerase II subunit hRPB11 and with the retinoblastoma protein (Rb) via two distinct domains, and functionally represses Rb's growth suppression by counteracting Rb-mediated inhibition of E2F1 transactivation.","method":"Co-immunoprecipitation, GST pulldown, transactivation reporter assay","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal binding assays plus functional reporter assay, single lab, multiple orthogonal methods","pmids":["10783144"],"is_preprint":false},{"year":2002,"finding":"Che-1/AATF contacts the Rb pocket region and competes with HDAC1 for the same Rb binding site, displacing HDAC1 from Rb/E2F complexes in vitro and from E2F target gene promoters in vivo, thereby activating DNA synthesis. Che-1-specific RNA interference reduces E2F activity and cell proliferation in Rb-proficient but not Rb-deficient cells.","method":"Co-immunoprecipitation, chromatin immunoprecipitation (ChIP), RNA interference, flow cytometry (DNA synthesis)","journal":"Cancer cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (Co-IP, ChIP, RNAi with defined phenotype), single lab with rigorous controls including Rb-deficient cell epistasis","pmids":["12450794"],"is_preprint":false},{"year":2003,"finding":"Che-1/AATF overexpression in colon carcinoma cells activates p21WAF1/CIP1 expression in a p53-independent manner by displacing HDAC1 from Sp1 binding sites on the p21 promoter and accumulating acetylated histone H3, causing G1 arrest. Che-1 RNAi reduces p21 transactivation and increases proliferation.","method":"Chromatin immunoprecipitation (ChIP), RNA interference, flow cytometry, reporter assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — ChIP demonstrating direct promoter occupancy and HDAC1 displacement plus RNAi phenotype, multiple orthogonal methods","pmids":["12847090"],"is_preprint":false},{"year":2003,"finding":"AATF colocalizes with Par-4 in cytoplasmic and nuclear compartments and interacts directly with Par-4 via the leucine zipper domain in neural cells. AATF/Par-4 complex formation is required for AATF to block Par-4-induced aberrant production and secretion of amyloid beta peptide 1-42.","method":"Co-immunoprecipitation, co-localization (immunofluorescence), ELISA for Aβ secretion, overexpression/co-expression assays","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — co-IP plus functional Aβ suppression assay, single lab, two complementary methods","pmids":["14627703"],"is_preprint":false},{"year":2003,"finding":"Che-1/AATF directly interacts with Tau protein via Tau's amino-terminal region (not involved in microtubule binding) in rat cerebellar granule neurons, partially colocalizing in the cytoplasm; this interaction is modulated during neuronal apoptosis.","method":"Co-immunoprecipitation from neuronal lysates, FRET analysis, overexpression in COS-7 cells","journal":"Molecular and cellular neurosciences","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — reciprocal Co-IP from neurons plus FRET validation, single lab, two orthogonal methods","pmids":["14697667"],"is_preprint":false},{"year":2004,"finding":"AATF overexpression in PC12 neural cells suppresses superoxide production, inhibits peroxynitrite formation and membrane lipid peroxidation, and protects against amyloid beta-peptide-induced apoptosis. Inhibition of AATF induction sensitizes cortical neurons to Aβ toxicity.","method":"Overexpression/knockdown in PC12 and cortical neurons, ROS assay, apoptosis assay","journal":"Neurobiology of disease","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — loss- and gain-of-function with multiple biochemical readouts, single lab","pmids":["15207272"],"is_preprint":false},{"year":2006,"finding":"Che-1/AATF contributes to the DNA damage response: checkpoint kinases ATM/ATR and Chk2 physically interact with Che-1 and phosphorylate it in response to DNA damage, leading to Che-1 accumulation, its specific recruitment to TP53 and p21 promoters, transcriptional activation of p53, and maintenance of the G2/M checkpoint.","method":"Co-immunoprecipitation, chromatin immunoprecipitation (ChIP), phosphorylation assay, flow cytometry (cell cycle), RNA interference","journal":"Cancer cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP of kinase-substrate interaction, ChIP showing promoter recruitment, cell-cycle phenotype, multiple orthogonal methods in one study","pmids":["17157788"],"is_preprint":false},{"year":2007,"finding":"In response to apoptotic stimuli, Che-1/AATF interacts with the peptidyl-prolyl isomerase Pin1; Pin1-induced conformational changes are required for Che-1 interaction with E3 ubiquitin ligase HDM2, which promotes Che-1 ubiquitination and proteasomal degradation. A Che-1 mutant unable to bind Pin1 has increased half-life and reduced apoptosis.","method":"Co-immunoprecipitation, ubiquitination assay, half-life measurement, site-directed mutagenesis, apoptosis assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (Co-IP, ubiquitination assay, mutagenesis, functional apoptosis readout), single lab with rigorous mechanistic dissection","pmids":["17468107"],"is_preprint":false},{"year":2007,"finding":"NRAGE overexpression inhibits nuclear localization of Che-1/AATF by sequestering it in the cytoplasm, and targets endogenous Che-1 for proteasome-dependent degradation. Che-1 overexpression completely reverts NRAGE-induced cell death, identifying them as functional antagonists.","method":"Co-immunoprecipitation, subcellular fractionation/localization (EGFP fusion), proteasome inhibitor rescue, cell death assay","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP, direct localization experiment with functional consequence, proteasome rescue, single lab","pmids":["17488777"],"is_preprint":false},{"year":2007,"finding":"Che-1/AATF phosphorylation following DNA damage promotes XIAP expression in a NF-κB-dependent manner, and XIAP expression is required for the antiapoptotic activity of Che-1.","method":"Chromatin immunoprecipitation (ChIP), RNA interference, reporter assay, in vivo tumor xenograft with siRNA","journal":"Cell death and differentiation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP plus functional RNAi, single lab, two orthogonal methods","pmids":["18049476"],"is_preprint":false},{"year":2008,"finding":"Che-1/AATF overexpression induces Cdk5 mRNA and protein expression, directly interacts with Cdk5 in vivo without competing with Cdk5/p35 binding (thus associating with the active kinase complex), and is itself a substrate of Cdk5 kinase.","method":"Co-immunoprecipitation, Western blot, in vitro kinase assay","journal":"Neuroreport","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP demonstrating complex formation plus kinase substrate assay, single lab","pmids":["18388733"],"is_preprint":false},{"year":2009,"finding":"AATF is induced by ER stress through the PERK-eIF2α pathway and transcriptionally activates AKT1 via STAT3, sustaining Akt1 activation and promoting cell survival. AATF knockdown sensitizes cells to ER stress-mediated death; ectopic AATF or constitutively active AKT1 confers resistance.","method":"Gene expression profiling, RNA interference, overexpression, Western blot, reporter assay","journal":"Cell death and differentiation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pathway dissection with genetic perturbations (RNAi, OE) and rescue experiments, single lab, multiple methods","pmids":["19911006"],"is_preprint":false},{"year":2010,"finding":"Che-1/AATF is required for sustaining mutant p53 expression in cancer cell lines; Che-1 depletion by siRNA induces apoptosis in vitro and in vivo and activates DNA damage checkpoint response and p73 transactivation in mutant-p53-expressing cells.","method":"siRNA knockdown, ChIP, Western blot, in vivo xenograft","journal":"Cancer cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP shows Che-1 on mutant p53 promoter, combined with in vitro and in vivo phenotype, single lab","pmids":["20708154"],"is_preprint":false},{"year":2011,"finding":"PARP-1 directly interacts with Che-1/AATF, promotes its poly(ADP-ribosyl)ation in vitro and in vivo, and stabilizes Che-1 after DNA damage independently of ATM kinase activity. Reduced PARP activity or PARP-1 knockout impairs Che-1 accumulation and its occupancy at the p21 promoter.","method":"Co-immunoprecipitation, in vitro PARylation assay, ChIP, PARP inhibitor/knockout, epistasis with ATM inhibitor","journal":"DNA repair","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — in vitro enzymatic assay demonstrating direct PARylation plus Co-IP, ChIP, genetic epistasis; multiple orthogonal methods","pmids":["21317046"],"is_preprint":false},{"year":2012,"finding":"Upon genotoxic stress, AATF/Che-1 is phosphorylated by MK2 (downstream of p38), which releases AATF from cytoplasmic MRLC3 and triggers nuclear translocation. Nuclear AATF binds PUMA, BAX, and BAK promoter regions to repress p53-driven transcription of pro-apoptotic genes. Phospho-mimicking AATF confers adriamycin resistance in vivo, while AATF-depleted tumors show enhanced drug response.","method":"Kinase assay (MK2), co-immunoprecipitation (MRLC3), subcellular fractionation, ChIP, site-directed mutagenesis, xenograft experiments","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — kinase substrate identification, Co-IP, ChIP, mutagenesis, and in vivo xenograft validation; multiple orthogonal methods, comprehensive mechanistic dissection","pmids":["22909821"],"is_preprint":false},{"year":2012,"finding":"AATF (nucleolar) acts as a cofactor required for c-Jun-mediated apoptosis. UV irradiation triggers AATF translocation from the nucleolus to the nucleus, where it physically associates with c-Jun. AATF overexpression or knockout levels proportionally alter c-Jun phosphorylation and expression of FasL and TNF-α. AATF mutants defective in c-Jun binding are also defective in AP-1 activity and apoptosis induction.","method":"Co-immunoprecipitation, subcellular localization (translocation assay), deletion mutagenesis, reporter assay, knockout MEFs","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, direct localization experiment with functional consequence, domain mutagenesis, and genetic knockout controls","pmids":["22933572"],"is_preprint":false},{"year":2013,"finding":"Che-1/AATF localizes to interphase centrosomes and accumulates there following DNA damage or spindle poisons. Che-1 depletion generates supernumerary centrosomes, multinucleated cells, and multipolar spindles. Che-1 is required for Chk1 binding to pericentrin and centrosomal localization of Chk1, which regulates centrosomal cyclin B-Cdk1 activation and mitotic entry timing.","method":"Immunofluorescence localization, siRNA knockdown, co-immunoprecipitation (Chk1-pericentrin), flow cytometry, centrosome counting","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct localization with functional consequence, Co-IP defining molecular pathway, multiple defined cellular phenotypes with RNAi","pmids":["23798705"],"is_preprint":false},{"year":2014,"finding":"HIPK2 interacts with Che-1/AATF and phosphorylates it at specific residues upon genotoxic stress, strongly enhancing HDM2/Che-1 interaction and subsequent ubiquitin-mediated proteasomal degradation. HIPK2 depletion reduces Che-1 ubiquitylation and degradation; Che-1 overexpression counteracts HIPK2-induced apoptosis.","method":"Co-immunoprecipitation, in vitro kinase assay, ubiquitylation assay, siRNA knockdown, apoptosis assay","journal":"Cell death & disease","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — in vitro kinase assay identifying Che-1 as HIPK2 substrate, Co-IP, ubiquitylation assay, and functional rescue; multiple orthogonal methods","pmids":["25210797"],"is_preprint":false},{"year":2015,"finding":"Under stress conditions, Che-1/AATF inhibits mTOR activity by inducing expression of the mTOR inhibitors Redd1 and Deptor, and this activity is required for stress-induced autophagy.","method":"siRNA knockdown, gene expression analysis, Western blot, autophagy assays","journal":"The EMBO journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — gene expression induction linked to functional autophagy phenotype via RNAi, single lab","pmids":["25770584"],"is_preprint":false},{"year":2015,"finding":"Che-1/AATF directly binds p53 protein, and this interaction occurs early in DNA damage response but is lost when cells undergo apoptosis. Che-1 forms a ternary complex with p53 and Brca1. Genome-wide ChIP analysis shows that Che-1/p53 interaction results in preferential transactivation of growth-arrest p53 target genes over pro-apoptotic targets. Che-1(+/-) mice show enhanced thymocyte apoptosis after ionizing radiation.","method":"Co-immunoprecipitation, genome-wide ChIP-seq, heterozygous knockout mouse model, irradiation assay","journal":"Cell death & disease","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome-wide ChIP-seq, Co-IP, and in vivo genetic mouse model with defined phenotype; multiple orthogonal methods","pmids":["25996291"],"is_preprint":false},{"year":2016,"finding":"AATF forms a salt-stable protein complex with neuroguidin (NGDN) and NOL10 (the ANN complex). All three members localize to nucleoli with mutual dependence for protein stability. The ANN complex is required for 18S rRNA maturation and nucleolar cleavage steps in the 5'ETS and ITS1 regions, supporting 40S ribosomal subunit biosynthesis.","method":"Immunoprecipitation (mass spectrometry), protein-protein interaction domain mapping, siRNA knockdown, rRNA processing analysis (Northern blot/pulse-chase), immunofluorescence","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 2 / Strong — MS-identified complex with domain mapping, rRNA processing analysis with individual depletions showing same cleavage defects; multiple orthogonal methods","pmids":["27599843"],"is_preprint":false},{"year":2017,"finding":"Che-1/AATF is required for HIF-1α stabilization under hypoxia; Che-1 depletion downregulates SIAH-2 expression (E3 ubiquitin ligase that degrades PHD3, the master regulator of HIF-1α stability), thereby reducing hypoxia-responsive gene expression and affecting glucose metabolism.","method":"siRNA knockdown, Western blot, NMR spectroscopy (metabolic), RNA-seq, ChIP","journal":"Journal of experimental & clinical cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pathway placement via RNAi with multiple readouts and ChIP, single lab","pmids":["28214471"],"is_preprint":false},{"year":2018,"finding":"In Kras-driven murine lung adenocarcinomas, AATF deletion delayed lung cancer formation predominantly in a p53-dependent manner, and targeting Aatf in existing tumors halted tumor progression. This identifies AATF as a key molecule sustaining proliferative tissue and tumor progression partly by inhibiting p53-driven apoptosis in vivo.","method":"Autochthonous Kras-driven mouse lung cancer model, dual recombinase conditional deletion, histology, survival analysis","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo genetic epistasis in autochthonous cancer model with p53-dependence established, rigorous dual-recombinase strategy","pmids":["29321668"],"is_preprint":false},{"year":2018,"finding":"c-Myc directly binds the Che-1/AATF promoter to regulate its expression, and Che-1 acts as a downstream effector of c-Myc. Genome-wide ChIP-seq shows overlapping genomic occupancy; RNA-seq upon depletion of either protein reveals strong overlap of controlled pathways. Che-1 depletion inhibits BCP-ALL cell proliferation.","method":"ChIP-seq, RNA-seq, siRNA knockdown, promoter binding assay, proliferation assay","journal":"EMBO reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome-wide ChIP-seq demonstrates direct promoter binding, RNA-seq epistasis, multiple orthogonal methods","pmids":["29367285"],"is_preprint":false},{"year":2020,"finding":"Che-1/AATF interacts with RNA polymerase I and the nucleolar upstream binding factor UBF, promotes RNA polymerase I-dependent transcription of rRNA genes (rDNA), and binds the rDNA promoter. Che-1 depletion reduces RNA Pol I and UBF recruitment on rDNA, decreases rDNA promoter activity and 47S pre-rRNA production, and induces abnormal nucleolar morphology. Upon DNA damage, Che-1 re-localizes from rDNA to the TP53 promoter.","method":"Co-immunoprecipitation (RNA Pol I, UBF), ChIP, reporter assay, siRNA knockdown, pre-rRNA Northern blot, immunofluorescence","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (Co-IP, ChIP, reporter, rRNA processing, localization); comprehensive mechanistic study in one lab","pmids":["32421830"],"is_preprint":false},{"year":2020,"finding":"Che-1/AATF directly interacts with histones and displaces HDAC class I members from histones in multiple myeloma cells, promoting global histone acetylation and active chromatin. Che-1 depletion leads to global histone deacetylation and reduced BRD4 chromatin accumulation. Transgenic mice expressing human Che-1 in plasma cells develop multiple myeloma.","method":"Co-immunoprecipitation (histones), HDAC displacement assay, ChIP-seq, Western blot, transgenic mouse model","journal":"Blood advances","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP of histone interaction, HDAC displacement, genome-wide ChIP-seq, and in vivo transgenic mouse model confirming oncogenic role","pmids":["33186461"],"is_preprint":false},{"year":2021,"finding":"CK2 protein kinase phosphorylates Che-1/AATF at Ser316, Ser320, and Ser321. These phosphorylation events are required for Che-1/histone H3 interaction and for Che-1's pro-proliferative activity. Mutation of these serines (Che-1 3S mutant) abolishes histone H3 binding.","method":"In vitro kinase assay (CK2), site-directed mutagenesis, 2D gel electrophoresis, mass spectrometry, co-immunoprecipitation, proliferation assay","journal":"Journal of experimental & clinical cancer research","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — in vitro kinase assay plus mutagenesis plus MS identification of phospho-sites, Co-IP, functional proliferation assay; multiple orthogonal methods","pmids":["34266450"],"is_preprint":false},{"year":2022,"finding":"AATF/Che-1 localizes to paraspeckles via interaction with the lncRNA NEAT1_2 and directly localizes on R-loops. Depletion of Che-1 causes marked accumulation of RNA:DNA hybrids (R-loops) and triggers sustained interferon gene expression signature through failure to resolve R-loops.","method":"Immunoprecipitation (NEAT1 lncRNA interaction), R-loop immunofluorescence/dot blot (S9.6 antibody), siRNA knockdown, RNA-seq (IFN signature), ChIP","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct localization to paraspeckles and R-loops established, functional R-loop accumulation and IFN activation upon depletion; multiple orthogonal methods","pmids":["35929179"],"is_preprint":false},{"year":2022,"finding":"In ischemic neurons, AATF competitively interacts with nuclear apoptosis-inducing factor (AIF) and inhibits AIF from binding DNA, thereby suppressing parthanatos (AIF-mediated cell death) without affecting PAR accumulation or AIF nuclear translocation.","method":"Co-immunoprecipitation (AATF-AIF), OGD/R model, dMCAO/R in vivo model, DNA fragmentation assay, overexpression","journal":"Journal of molecular neuroscience","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP demonstrating AATF-AIF interaction plus defined mechanistic consequence (DNA binding competition) in vitro and in vivo, single lab","pmids":["36058992"],"is_preprint":false},{"year":2023,"finding":"HAX1 is a novel binding partner of Che-1/AATF; both proteins colocalize extensively in mitochondria and their association is strengthened after oxidative stress. Che-1 depletion correlates with decreased HAX1 mRNA and protein levels in MCF-7 cells.","method":"Co-immunoprecipitation, co-localization (immunofluorescence), siRNA knockdown, Western blot","journal":"Biochimica et biophysica acta. Molecular cell research","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single Co-IP plus co-localization, no mechanistic dissection of functional consequence, single lab","pmids":["37742722"],"is_preprint":false},{"year":2025,"finding":"AATF interacts with XRCC4 (a core NHEJ subunit) and prevents its ubiquitin-mediated proteasomal degradation. Upon DNA damage, ATM phosphorylates AATF at Ser189, causing its dissociation from XRCC4 and rapid recruitment of XRCC4 to DNA break sites for NHEJ repair. AATF depletion or phosphorylation-deficient mutant impedes NHEJ in glioma stem-like cells.","method":"Co-immunoprecipitation (AATF-XRCC4), ATM kinase assay (Ser189 phosphorylation), site-directed mutagenesis, NHEJ assay, ubiquitination assay, xenograft model","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — kinase substrate identification, Co-IP, mutagenesis of phospho-site, NHEJ functional assay, and in vivo xenograft; multiple orthogonal methods","pmids":["40436899"],"is_preprint":false},{"year":2017,"finding":"Che-1/AATF interacts with subunits of the HAT module of SAGA complexes (ADA2A, ADA2B, and GCN5) in human cells, as demonstrated by co-immunoprecipitation and co-localization, with specific domains in ADA2 and GCN5 required for these interactions.","method":"Co-immunoprecipitation, yeast two-hybrid (domain mapping), co-localization","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP plus yeast two-hybrid domain mapping, single lab, two complementary methods","pmids":["29232376"],"is_preprint":false},{"year":2003,"finding":"The Che-1/AATF promoter is TATA-less and contains a negative feedback element: Che-1 protein binds its own promoter and represses its own transcription.","method":"Reporter assay, chromatin immunoprecipitation (ChIP), promoter deletion analysis","journal":"Gene","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — ChIP demonstrating auto-occupancy plus functional reporter assay, single lab","pmids":["14636992"],"is_preprint":false},{"year":2025,"finding":"In pancreatic neuroendocrine tumors, ACSS2 activity induces histone H3/H4 hyperacetylation, which recruits AATF to co-regulate FasL transcription, specifically enhancing soluble FasL secretion. Secreted FasL triggers CD8+ T cell apoptosis via caspase-8/3 cascade, promoting immune evasion. ACSS2 and AATF act non-redundantly and synergistically in modulating FasL expression.","method":"ChIP, Western blot, ELISA (sFasL), co-immunoprecipitation, siRNA/overexpression, flow cytometry (T cell apoptosis)","journal":"Advanced science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP linking AATF to FasL promoter, functional sFasL secretion and T cell apoptosis assays, single lab","pmids":["40791180"],"is_preprint":false}],"current_model":"AATF/Che-1 is a multifunctional nuclear phosphoprotein that binds RNA polymerase II (and I), Rb, and multiple transcription factors; in unstressed cells it promotes proliferation by displacing HDAC1 from Rb/E2F and Sp1 complexes to activate E2F target genes and p21, while upon genotoxic stress it is phosphorylated by ATM/ATR, Chk2, MK2, CK2, and HIPK2 at distinct residues—triggering nuclear translocation, promoter switching from E2F targets to p53/p21 loci to activate cell-cycle arrest, or (when phosphorylated by MK2) binding PUMA/BAX/BAK promoters to repress p53-driven apoptosis—and is ultimately degraded via Pin1/HDM2/HIPK2-mediated ubiquitylation during apoptosis; AATF also forms the ANN complex (with NGDN and NOL10) in nucleoli to support 40S ribosomal subunit biogenesis and rRNA processing, localizes to paraspeckles via NEAT1_2 to resolve R-loops and suppress interferon activation, stabilizes XRCC4 for NHEJ repair (released by ATM-mediated Ser189 phosphorylation at DNA breaks), and interacts with partners including Par-4, Tau, Cdk5, c-Jun, AIF, SAGA subunits, and histones to regulate apoptosis, neuronal function, and chromatin acetylation across multiple cellular contexts."},"narrative":{"mechanistic_narrative":"AATF/Che-1 is a multifunctional nuclear phosphoprotein that integrates transcriptional control of proliferation, the DNA damage response, and apoptosis [PMID:12450794, PMID:17157788]. It binds RNA polymerase II subunit hRPB11 and the retinoblastoma protein, and in proliferating cells it competes with HDAC1 for the Rb pocket—displacing HDAC1 from Rb/E2F complexes and from Sp1 sites to activate E2F targets and p21WAF1/CIP1 with accumulation of acetylated histone H3 [PMID:10783144, PMID:12450794, PMID:12847090]. This pro-proliferative chromatin activity depends on direct histone binding, which requires CK2 phosphorylation at Ser316/Ser320/Ser321 and is reinforced by displacement of class I HDACs to sustain global histone acetylation and BRD4 recruitment [PMID:33186461, PMID:34266450]. Upon genotoxic stress, AATF is phosphorylated and stabilized by ATM/ATR and Chk2 and PARylated by PARP-1, accumulating and switching its promoter occupancy from rDNA and E2F loci to the TP53 and p21 promoters to activate growth-arrest p53 target genes and maintain cell-cycle checkpoints [PMID:17157788, PMID:21317046, PMID:25996291, PMID:32421830]. AATF restrains apoptosis through multiple routes: MK2 phosphorylation releases it to the nucleus where it represses p53-driven PUMA/BAX/BAK transcription, and it sustains XIAP and mutant p53 expression [PMID:18049476, PMID:20708154, PMID:22909821]. Its own abundance is controlled by Pin1- and HIPK2-dependent priming of HDM2-mediated ubiquitylation and proteasomal degradation during apoptosis [PMID:17468107, PMID:25210797]. In the nucleolus AATF supports ribosome biogenesis, forming the ANN complex with NGDN and NOL10 required for 18S rRNA maturation and 40S subunit synthesis and promoting RNA Pol I/UBF-dependent rDNA transcription [PMID:27599843, PMID:32421830]. It additionally stabilizes the NHEJ factor XRCC4 until ATM phosphorylation at Ser189 releases it for recruitment to DNA breaks, and localizes to paraspeckles via NEAT1_2 to resolve R-loops and suppress interferon activation [PMID:35929179, PMID:40436899]. These activities make AATF a driver of tumor proliferation in vivo, including Kras-driven lung adenocarcinoma in a p53-dependent manner and plasma-cell multiple myeloma [PMID:29321668, PMID:33186461].","teleology":[{"year":1999,"claim":"Establishing AATF's existence and its first functional theme—antagonism of apoptosis—anchored all later work by linking it to a death kinase and to transcription.","evidence":"Yeast two-hybrid against Dlk/ZIP kinase plus Gal4-BD transactivation and apoptosis-interference assays","pmids":["10580117"],"confidence":"Medium","gaps":["No direct DNA targets identified","Domain requirements for transactivation undefined"]},{"year":2003,"claim":"Defining AATF as an HDAC1-displacing cofactor at Rb/E2F and Sp1 sites explained how it drives proliferation and p53-independent p21 activation through chromatin acetylation.","evidence":"Co-IP, ChIP, and RNAi in Rb-proficient vs Rb-deficient cells; ChIP and reporter assays on the p21/Sp1 promoter","pmids":["10783144","12450794","12847090"],"confidence":"High","gaps":["Direct vs indirect promoter recruitment mechanism not fully resolved","Phospho-regulation of HDAC1 competition not yet known at this stage"]},{"year":2007,"claim":"Identifying ATM/ATR/Chk2 phosphorylation as a switch that stabilizes AATF and redirects it to TP53/p21 promoters established its role as a damage-responsive checkpoint activator.","evidence":"Co-IP of kinase-substrate, ChIP of promoter recruitment, cell-cycle flow cytometry, RNAi","pmids":["17157788"],"confidence":"High","gaps":["Exact phospho-residues not mapped","Mechanism of promoter selectivity unresolved"]},{"year":2007,"claim":"Mapping Pin1-primed, HDM2-mediated ubiquitylation showed how AATF protein levels are downregulated to permit apoptosis, defining its turnover control.","evidence":"Co-IP, ubiquitination assay, half-life measurement, Pin1-binding mutant, apoptosis assay","pmids":["17468107"],"confidence":"High","gaps":["Phospho-sites recognized by Pin1 not defined here","Other E3 ligases not excluded"]},{"year":2011,"claim":"Demonstrating PARP-1-dependent PARylation as an ATM-independent route to AATF stabilization revealed a parallel damage signal converging on its promoter occupancy.","evidence":"Co-IP, in vitro PARylation, ChIP at p21 promoter, PARP inhibitor/knockout and ATM-inhibitor epistasis","pmids":["21317046"],"confidence":"High","gaps":["PARylation acceptor residues not mapped","Interplay with phosphorylation marks unresolved"]},{"year":2012,"claim":"Identifying MK2 phosphorylation as the trigger for cytoplasm-to-nucleus translocation and repression of PUMA/BAX/BAK promoters explained how AATF actively blocks p53-driven apoptosis and confers chemoresistance.","evidence":"MK2 kinase assay, MRLC3 Co-IP, fractionation, ChIP, phospho-mimic mutagenesis, xenografts","pmids":["22909821"],"confidence":"High","gaps":["How nuclear AATF selects pro-apoptotic vs growth-arrest p53 loci unclear","Relationship to ATM/Chk2 signaling not integrated"]},{"year":2015,"claim":"Direct p53 binding forming a p53/Brca1 ternary complex that biases transactivation toward growth-arrest genes, confirmed in Che-1(+/-) mice, established AATF as a determinant of the arrest-versus-death p53 decision.","evidence":"Co-IP, genome-wide ChIP-seq, heterozygous knockout mice with irradiation phenotype","pmids":["25996291"],"confidence":"High","gaps":["Structural basis of p53 target selectivity unknown","How interaction is lost during apoptosis not mechanistically defined"]},{"year":2014,"claim":"Adding HIPK2 as a stress kinase that primes AATF for HDM2-dependent degradation completed the picture of how genotoxic signaling tips AATF abundance toward apoptosis.","evidence":"Co-IP, in vitro kinase assay, ubiquitylation assay, RNAi, apoptosis rescue","pmids":["25210797"],"confidence":"High","gaps":["HIPK2 target residues on AATF not specified","Coordination with Pin1/CK2 phosphorylation unresolved"]},{"year":2016,"claim":"Discovery of the ANN complex (AATF-NGDN-NOL10) localized a major non-transcriptional role to nucleolar ribosome biogenesis.","evidence":"IP-MS, domain mapping, RNAi with rRNA processing analysis, immunofluorescence","pmids":["27599843"],"confidence":"High","gaps":["Molecular function of AATF within the complex (catalytic vs scaffold) undefined","Link between ribosome biogenesis and stress signaling not established"]},{"year":2020,"claim":"Linking AATF to RNA Pol I/UBF-driven rDNA transcription, with damage-induced re-localization from rDNA to TP53, unified its nucleolar and checkpoint roles into a stress-responsive promoter switch.","evidence":"Co-IP with Pol I/UBF, ChIP at rDNA, reporter assay, pre-rRNA Northern, immunofluorescence","pmids":["32421830"],"confidence":"High","gaps":["Signal driving rDNA-to-TP53 relocalization not defined","Relationship to ANN complex function unclear"]},{"year":2021,"claim":"Mapping CK2 phosphorylation at Ser316/320/321 as a requirement for histone H3 binding provided the molecular basis for AATF's pro-proliferative chromatin acetylation activity.","evidence":"In vitro CK2 kinase assay, mutagenesis, 2D gel/MS phospho-site mapping, Co-IP, proliferation assay","pmids":["33186461","34266450"],"confidence":"High","gaps":["Structural mode of histone engagement undefined","Whether other kinases regulate this interaction unknown"]},{"year":2022,"claim":"Localization to NEAT1_2 paraspeckles and R-loops, with R-loop accumulation and interferon induction upon depletion, revealed a genome-stability role coupling AATF to innate immune suppression.","evidence":"NEAT1 RNA-IP, S9.6 R-loop dot blot/IF, RNAi, RNA-seq IFN signature, ChIP","pmids":["35929179"],"confidence":"High","gaps":["Direct enzymatic activity on R-loops not demonstrated","Mechanism of R-loop resolution by AATF unresolved"]},{"year":2025,"claim":"Identifying ATM-Ser189 phosphorylation as the release signal that frees stabilized XRCC4 for recruitment to breaks placed AATF directly within NHEJ repair.","evidence":"Co-IP, ATM kinase assay, phospho-site mutagenesis, NHEJ and ubiquitination assays, xenograft","pmids":["40436899"],"confidence":"High","gaps":["Generality across cell types beyond glioma stem-like cells untested","Coordination with AATF's transcriptional damage response unresolved"]},{"year":null,"claim":"How AATF's many post-translational modifications (ATM/ATR, Chk2, MK2, CK2, HIPK2 phosphorylation; PARylation; ubiquitylation) are hierarchically integrated to dictate its localization and the choice among proliferation, arrest, apoptosis suppression, and repair remains unresolved.","evidence":"","pmids":[],"confidence":"High","gaps":["No unified model of combinatorial PTM control","No structural framework for its distinct binding interfaces","Tissue-specific determinants of functional output undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[0,2,3,7,20,25]},{"term_id":"GO:0042393","term_label":"histone binding","supporting_discovery_ids":[26,27]},{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[28]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[2,3,21,31]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[2,25,31]}],"localization":[{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[16,21,25]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[2,7,9,20]},{"term_id":"GO:0005654","term_label":"nucleoplasm","supporting_discovery_ids":[28]},{"term_id":"GO:0005815","term_label":"microtubule organizing center","supporting_discovery_ids":[17]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[4,15]}],"pathway":[{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[7,14,31]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[2,7,17,20]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[8,15,16,18,29]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[2,3,25,26]},{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[21,25]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[23,24,26]},{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[12,19,22,28]}],"complexes":["ANN complex (AATF-NGDN-NOL10)","SAGA HAT module","RNA polymerase I/UBF rDNA transcription complex"],"partners":["RB1","TP53","HDAC1","HDM2","PIN1","PARP1","XRCC4","NGDN"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9NY61","full_name":"Protein AATF","aliases":["Apoptosis-antagonizing transcription factor","Rb-binding protein Che-1"],"length_aa":560,"mass_kda":63.1,"function":"Part of the small subunit (SSU) processome, first precursor of the small eukaryotic ribosomal subunit. 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Also antagonizes PAWR mediated induction of aberrant amyloid peptide production in Alzheimer disease (presenile and senile dementia), although the molecular basis for this phenomenon has not been described to date","subcellular_location":"Nucleus, nucleolus","url":"https://www.uniprot.org/uniprotkb/Q9NY61/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/AATF","classification":"Common 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Modeling Reveals the Importance of the DED Filament Composition in the Effects of Small Molecules Targeting Caspase-8/c-FLIPL Heterodimer.","date":"2020","source":"Biochemistry. 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Implications in oxidative stress cell response.","date":"2023","source":"Biochimica et biophysica acta. Molecular cell research","url":"https://pubmed.ncbi.nlm.nih.gov/37742722","citation_count":2,"is_preprint":false},{"pmid":"29943642","id":"PMC_29943642","title":"A new baby in the c-Myc-directed transcriptional machinery: Che-1/AATF.","date":"2018","source":"Cell cycle (Georgetown, Tex.)","url":"https://pubmed.ncbi.nlm.nih.gov/29943642","citation_count":2,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":46793,"output_tokens":8661,"usd":0.135147,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":18414,"output_tokens":4883,"usd":0.107073,"stage2_stop_reason":"end_turn"},"total_usd":0.24222,"stage1_batch_id":"msgbatch_0179Z2pxRYBsXi2xADneafRw","stage2_batch_id":"msgbatch_01LaY9ZaVdgjWArcZo9eqQE5","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1999,\n      \"finding\": \"AATF (apoptosis antagonizing transcription factor) was identified as a binding partner of Dlk/ZIP kinase (a serine/threonine kinase) via yeast two-hybrid and shown to interfere with Dlk-induced apoptosis. AATF contains an acidic domain and putative leucine zipper and exhibits transcriptional transactivation activity as a Gal4-BD fusion protein.\",\n      \"method\": \"Yeast two-hybrid interaction screen, transactivation assay (Gal4-BD fusion), apoptosis interference assay\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — yeast two-hybrid plus functional transactivation assay and apoptosis assay, single lab, two complementary methods\",\n      \"pmids\": [\"10580117\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"Che-1/AATF interacts with RNA polymerase II subunit hRPB11 and with the retinoblastoma protein (Rb) via two distinct domains, and functionally represses Rb's growth suppression by counteracting Rb-mediated inhibition of E2F1 transactivation.\",\n      \"method\": \"Co-immunoprecipitation, GST pulldown, transactivation reporter assay\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal binding assays plus functional reporter assay, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"10783144\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Che-1/AATF contacts the Rb pocket region and competes with HDAC1 for the same Rb binding site, displacing HDAC1 from Rb/E2F complexes in vitro and from E2F target gene promoters in vivo, thereby activating DNA synthesis. Che-1-specific RNA interference reduces E2F activity and cell proliferation in Rb-proficient but not Rb-deficient cells.\",\n      \"method\": \"Co-immunoprecipitation, chromatin immunoprecipitation (ChIP), RNA interference, flow cytometry (DNA synthesis)\",\n      \"journal\": \"Cancer cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (Co-IP, ChIP, RNAi with defined phenotype), single lab with rigorous controls including Rb-deficient cell epistasis\",\n      \"pmids\": [\"12450794\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Che-1/AATF overexpression in colon carcinoma cells activates p21WAF1/CIP1 expression in a p53-independent manner by displacing HDAC1 from Sp1 binding sites on the p21 promoter and accumulating acetylated histone H3, causing G1 arrest. Che-1 RNAi reduces p21 transactivation and increases proliferation.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP), RNA interference, flow cytometry, reporter assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — ChIP demonstrating direct promoter occupancy and HDAC1 displacement plus RNAi phenotype, multiple orthogonal methods\",\n      \"pmids\": [\"12847090\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"AATF colocalizes with Par-4 in cytoplasmic and nuclear compartments and interacts directly with Par-4 via the leucine zipper domain in neural cells. AATF/Par-4 complex formation is required for AATF to block Par-4-induced aberrant production and secretion of amyloid beta peptide 1-42.\",\n      \"method\": \"Co-immunoprecipitation, co-localization (immunofluorescence), ELISA for Aβ secretion, overexpression/co-expression assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — co-IP plus functional Aβ suppression assay, single lab, two complementary methods\",\n      \"pmids\": [\"14627703\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Che-1/AATF directly interacts with Tau protein via Tau's amino-terminal region (not involved in microtubule binding) in rat cerebellar granule neurons, partially colocalizing in the cytoplasm; this interaction is modulated during neuronal apoptosis.\",\n      \"method\": \"Co-immunoprecipitation from neuronal lysates, FRET analysis, overexpression in COS-7 cells\",\n      \"journal\": \"Molecular and cellular neurosciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — reciprocal Co-IP from neurons plus FRET validation, single lab, two orthogonal methods\",\n      \"pmids\": [\"14697667\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"AATF overexpression in PC12 neural cells suppresses superoxide production, inhibits peroxynitrite formation and membrane lipid peroxidation, and protects against amyloid beta-peptide-induced apoptosis. Inhibition of AATF induction sensitizes cortical neurons to Aβ toxicity.\",\n      \"method\": \"Overexpression/knockdown in PC12 and cortical neurons, ROS assay, apoptosis assay\",\n      \"journal\": \"Neurobiology of disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — loss- and gain-of-function with multiple biochemical readouts, single lab\",\n      \"pmids\": [\"15207272\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Che-1/AATF contributes to the DNA damage response: checkpoint kinases ATM/ATR and Chk2 physically interact with Che-1 and phosphorylate it in response to DNA damage, leading to Che-1 accumulation, its specific recruitment to TP53 and p21 promoters, transcriptional activation of p53, and maintenance of the G2/M checkpoint.\",\n      \"method\": \"Co-immunoprecipitation, chromatin immunoprecipitation (ChIP), phosphorylation assay, flow cytometry (cell cycle), RNA interference\",\n      \"journal\": \"Cancer cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP of kinase-substrate interaction, ChIP showing promoter recruitment, cell-cycle phenotype, multiple orthogonal methods in one study\",\n      \"pmids\": [\"17157788\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"In response to apoptotic stimuli, Che-1/AATF interacts with the peptidyl-prolyl isomerase Pin1; Pin1-induced conformational changes are required for Che-1 interaction with E3 ubiquitin ligase HDM2, which promotes Che-1 ubiquitination and proteasomal degradation. A Che-1 mutant unable to bind Pin1 has increased half-life and reduced apoptosis.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, half-life measurement, site-directed mutagenesis, apoptosis assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (Co-IP, ubiquitination assay, mutagenesis, functional apoptosis readout), single lab with rigorous mechanistic dissection\",\n      \"pmids\": [\"17468107\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"NRAGE overexpression inhibits nuclear localization of Che-1/AATF by sequestering it in the cytoplasm, and targets endogenous Che-1 for proteasome-dependent degradation. Che-1 overexpression completely reverts NRAGE-induced cell death, identifying them as functional antagonists.\",\n      \"method\": \"Co-immunoprecipitation, subcellular fractionation/localization (EGFP fusion), proteasome inhibitor rescue, cell death assay\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP, direct localization experiment with functional consequence, proteasome rescue, single lab\",\n      \"pmids\": [\"17488777\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Che-1/AATF phosphorylation following DNA damage promotes XIAP expression in a NF-κB-dependent manner, and XIAP expression is required for the antiapoptotic activity of Che-1.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP), RNA interference, reporter assay, in vivo tumor xenograft with siRNA\",\n      \"journal\": \"Cell death and differentiation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP plus functional RNAi, single lab, two orthogonal methods\",\n      \"pmids\": [\"18049476\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Che-1/AATF overexpression induces Cdk5 mRNA and protein expression, directly interacts with Cdk5 in vivo without competing with Cdk5/p35 binding (thus associating with the active kinase complex), and is itself a substrate of Cdk5 kinase.\",\n      \"method\": \"Co-immunoprecipitation, Western blot, in vitro kinase assay\",\n      \"journal\": \"Neuroreport\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP demonstrating complex formation plus kinase substrate assay, single lab\",\n      \"pmids\": [\"18388733\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"AATF is induced by ER stress through the PERK-eIF2α pathway and transcriptionally activates AKT1 via STAT3, sustaining Akt1 activation and promoting cell survival. AATF knockdown sensitizes cells to ER stress-mediated death; ectopic AATF or constitutively active AKT1 confers resistance.\",\n      \"method\": \"Gene expression profiling, RNA interference, overexpression, Western blot, reporter assay\",\n      \"journal\": \"Cell death and differentiation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pathway dissection with genetic perturbations (RNAi, OE) and rescue experiments, single lab, multiple methods\",\n      \"pmids\": [\"19911006\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Che-1/AATF is required for sustaining mutant p53 expression in cancer cell lines; Che-1 depletion by siRNA induces apoptosis in vitro and in vivo and activates DNA damage checkpoint response and p73 transactivation in mutant-p53-expressing cells.\",\n      \"method\": \"siRNA knockdown, ChIP, Western blot, in vivo xenograft\",\n      \"journal\": \"Cancer cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP shows Che-1 on mutant p53 promoter, combined with in vitro and in vivo phenotype, single lab\",\n      \"pmids\": [\"20708154\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"PARP-1 directly interacts with Che-1/AATF, promotes its poly(ADP-ribosyl)ation in vitro and in vivo, and stabilizes Che-1 after DNA damage independently of ATM kinase activity. Reduced PARP activity or PARP-1 knockout impairs Che-1 accumulation and its occupancy at the p21 promoter.\",\n      \"method\": \"Co-immunoprecipitation, in vitro PARylation assay, ChIP, PARP inhibitor/knockout, epistasis with ATM inhibitor\",\n      \"journal\": \"DNA repair\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — in vitro enzymatic assay demonstrating direct PARylation plus Co-IP, ChIP, genetic epistasis; multiple orthogonal methods\",\n      \"pmids\": [\"21317046\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Upon genotoxic stress, AATF/Che-1 is phosphorylated by MK2 (downstream of p38), which releases AATF from cytoplasmic MRLC3 and triggers nuclear translocation. Nuclear AATF binds PUMA, BAX, and BAK promoter regions to repress p53-driven transcription of pro-apoptotic genes. Phospho-mimicking AATF confers adriamycin resistance in vivo, while AATF-depleted tumors show enhanced drug response.\",\n      \"method\": \"Kinase assay (MK2), co-immunoprecipitation (MRLC3), subcellular fractionation, ChIP, site-directed mutagenesis, xenograft experiments\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — kinase substrate identification, Co-IP, ChIP, mutagenesis, and in vivo xenograft validation; multiple orthogonal methods, comprehensive mechanistic dissection\",\n      \"pmids\": [\"22909821\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"AATF (nucleolar) acts as a cofactor required for c-Jun-mediated apoptosis. UV irradiation triggers AATF translocation from the nucleolus to the nucleus, where it physically associates with c-Jun. AATF overexpression or knockout levels proportionally alter c-Jun phosphorylation and expression of FasL and TNF-α. AATF mutants defective in c-Jun binding are also defective in AP-1 activity and apoptosis induction.\",\n      \"method\": \"Co-immunoprecipitation, subcellular localization (translocation assay), deletion mutagenesis, reporter assay, knockout MEFs\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, direct localization experiment with functional consequence, domain mutagenesis, and genetic knockout controls\",\n      \"pmids\": [\"22933572\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Che-1/AATF localizes to interphase centrosomes and accumulates there following DNA damage or spindle poisons. Che-1 depletion generates supernumerary centrosomes, multinucleated cells, and multipolar spindles. Che-1 is required for Chk1 binding to pericentrin and centrosomal localization of Chk1, which regulates centrosomal cyclin B-Cdk1 activation and mitotic entry timing.\",\n      \"method\": \"Immunofluorescence localization, siRNA knockdown, co-immunoprecipitation (Chk1-pericentrin), flow cytometry, centrosome counting\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct localization with functional consequence, Co-IP defining molecular pathway, multiple defined cellular phenotypes with RNAi\",\n      \"pmids\": [\"23798705\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"HIPK2 interacts with Che-1/AATF and phosphorylates it at specific residues upon genotoxic stress, strongly enhancing HDM2/Che-1 interaction and subsequent ubiquitin-mediated proteasomal degradation. HIPK2 depletion reduces Che-1 ubiquitylation and degradation; Che-1 overexpression counteracts HIPK2-induced apoptosis.\",\n      \"method\": \"Co-immunoprecipitation, in vitro kinase assay, ubiquitylation assay, siRNA knockdown, apoptosis assay\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — in vitro kinase assay identifying Che-1 as HIPK2 substrate, Co-IP, ubiquitylation assay, and functional rescue; multiple orthogonal methods\",\n      \"pmids\": [\"25210797\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Under stress conditions, Che-1/AATF inhibits mTOR activity by inducing expression of the mTOR inhibitors Redd1 and Deptor, and this activity is required for stress-induced autophagy.\",\n      \"method\": \"siRNA knockdown, gene expression analysis, Western blot, autophagy assays\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — gene expression induction linked to functional autophagy phenotype via RNAi, single lab\",\n      \"pmids\": [\"25770584\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Che-1/AATF directly binds p53 protein, and this interaction occurs early in DNA damage response but is lost when cells undergo apoptosis. Che-1 forms a ternary complex with p53 and Brca1. Genome-wide ChIP analysis shows that Che-1/p53 interaction results in preferential transactivation of growth-arrest p53 target genes over pro-apoptotic targets. Che-1(+/-) mice show enhanced thymocyte apoptosis after ionizing radiation.\",\n      \"method\": \"Co-immunoprecipitation, genome-wide ChIP-seq, heterozygous knockout mouse model, irradiation assay\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome-wide ChIP-seq, Co-IP, and in vivo genetic mouse model with defined phenotype; multiple orthogonal methods\",\n      \"pmids\": [\"25996291\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"AATF forms a salt-stable protein complex with neuroguidin (NGDN) and NOL10 (the ANN complex). All three members localize to nucleoli with mutual dependence for protein stability. The ANN complex is required for 18S rRNA maturation and nucleolar cleavage steps in the 5'ETS and ITS1 regions, supporting 40S ribosomal subunit biosynthesis.\",\n      \"method\": \"Immunoprecipitation (mass spectrometry), protein-protein interaction domain mapping, siRNA knockdown, rRNA processing analysis (Northern blot/pulse-chase), immunofluorescence\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — MS-identified complex with domain mapping, rRNA processing analysis with individual depletions showing same cleavage defects; multiple orthogonal methods\",\n      \"pmids\": [\"27599843\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Che-1/AATF is required for HIF-1α stabilization under hypoxia; Che-1 depletion downregulates SIAH-2 expression (E3 ubiquitin ligase that degrades PHD3, the master regulator of HIF-1α stability), thereby reducing hypoxia-responsive gene expression and affecting glucose metabolism.\",\n      \"method\": \"siRNA knockdown, Western blot, NMR spectroscopy (metabolic), RNA-seq, ChIP\",\n      \"journal\": \"Journal of experimental & clinical cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pathway placement via RNAi with multiple readouts and ChIP, single lab\",\n      \"pmids\": [\"28214471\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"In Kras-driven murine lung adenocarcinomas, AATF deletion delayed lung cancer formation predominantly in a p53-dependent manner, and targeting Aatf in existing tumors halted tumor progression. This identifies AATF as a key molecule sustaining proliferative tissue and tumor progression partly by inhibiting p53-driven apoptosis in vivo.\",\n      \"method\": \"Autochthonous Kras-driven mouse lung cancer model, dual recombinase conditional deletion, histology, survival analysis\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo genetic epistasis in autochthonous cancer model with p53-dependence established, rigorous dual-recombinase strategy\",\n      \"pmids\": [\"29321668\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"c-Myc directly binds the Che-1/AATF promoter to regulate its expression, and Che-1 acts as a downstream effector of c-Myc. Genome-wide ChIP-seq shows overlapping genomic occupancy; RNA-seq upon depletion of either protein reveals strong overlap of controlled pathways. Che-1 depletion inhibits BCP-ALL cell proliferation.\",\n      \"method\": \"ChIP-seq, RNA-seq, siRNA knockdown, promoter binding assay, proliferation assay\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome-wide ChIP-seq demonstrates direct promoter binding, RNA-seq epistasis, multiple orthogonal methods\",\n      \"pmids\": [\"29367285\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Che-1/AATF interacts with RNA polymerase I and the nucleolar upstream binding factor UBF, promotes RNA polymerase I-dependent transcription of rRNA genes (rDNA), and binds the rDNA promoter. Che-1 depletion reduces RNA Pol I and UBF recruitment on rDNA, decreases rDNA promoter activity and 47S pre-rRNA production, and induces abnormal nucleolar morphology. Upon DNA damage, Che-1 re-localizes from rDNA to the TP53 promoter.\",\n      \"method\": \"Co-immunoprecipitation (RNA Pol I, UBF), ChIP, reporter assay, siRNA knockdown, pre-rRNA Northern blot, immunofluorescence\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (Co-IP, ChIP, reporter, rRNA processing, localization); comprehensive mechanistic study in one lab\",\n      \"pmids\": [\"32421830\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Che-1/AATF directly interacts with histones and displaces HDAC class I members from histones in multiple myeloma cells, promoting global histone acetylation and active chromatin. Che-1 depletion leads to global histone deacetylation and reduced BRD4 chromatin accumulation. Transgenic mice expressing human Che-1 in plasma cells develop multiple myeloma.\",\n      \"method\": \"Co-immunoprecipitation (histones), HDAC displacement assay, ChIP-seq, Western blot, transgenic mouse model\",\n      \"journal\": \"Blood advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP of histone interaction, HDAC displacement, genome-wide ChIP-seq, and in vivo transgenic mouse model confirming oncogenic role\",\n      \"pmids\": [\"33186461\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"CK2 protein kinase phosphorylates Che-1/AATF at Ser316, Ser320, and Ser321. These phosphorylation events are required for Che-1/histone H3 interaction and for Che-1's pro-proliferative activity. Mutation of these serines (Che-1 3S mutant) abolishes histone H3 binding.\",\n      \"method\": \"In vitro kinase assay (CK2), site-directed mutagenesis, 2D gel electrophoresis, mass spectrometry, co-immunoprecipitation, proliferation assay\",\n      \"journal\": \"Journal of experimental & clinical cancer research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — in vitro kinase assay plus mutagenesis plus MS identification of phospho-sites, Co-IP, functional proliferation assay; multiple orthogonal methods\",\n      \"pmids\": [\"34266450\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"AATF/Che-1 localizes to paraspeckles via interaction with the lncRNA NEAT1_2 and directly localizes on R-loops. Depletion of Che-1 causes marked accumulation of RNA:DNA hybrids (R-loops) and triggers sustained interferon gene expression signature through failure to resolve R-loops.\",\n      \"method\": \"Immunoprecipitation (NEAT1 lncRNA interaction), R-loop immunofluorescence/dot blot (S9.6 antibody), siRNA knockdown, RNA-seq (IFN signature), ChIP\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct localization to paraspeckles and R-loops established, functional R-loop accumulation and IFN activation upon depletion; multiple orthogonal methods\",\n      \"pmids\": [\"35929179\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"In ischemic neurons, AATF competitively interacts with nuclear apoptosis-inducing factor (AIF) and inhibits AIF from binding DNA, thereby suppressing parthanatos (AIF-mediated cell death) without affecting PAR accumulation or AIF nuclear translocation.\",\n      \"method\": \"Co-immunoprecipitation (AATF-AIF), OGD/R model, dMCAO/R in vivo model, DNA fragmentation assay, overexpression\",\n      \"journal\": \"Journal of molecular neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP demonstrating AATF-AIF interaction plus defined mechanistic consequence (DNA binding competition) in vitro and in vivo, single lab\",\n      \"pmids\": [\"36058992\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"HAX1 is a novel binding partner of Che-1/AATF; both proteins colocalize extensively in mitochondria and their association is strengthened after oxidative stress. Che-1 depletion correlates with decreased HAX1 mRNA and protein levels in MCF-7 cells.\",\n      \"method\": \"Co-immunoprecipitation, co-localization (immunofluorescence), siRNA knockdown, Western blot\",\n      \"journal\": \"Biochimica et biophysica acta. Molecular cell research\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP plus co-localization, no mechanistic dissection of functional consequence, single lab\",\n      \"pmids\": [\"37742722\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"AATF interacts with XRCC4 (a core NHEJ subunit) and prevents its ubiquitin-mediated proteasomal degradation. Upon DNA damage, ATM phosphorylates AATF at Ser189, causing its dissociation from XRCC4 and rapid recruitment of XRCC4 to DNA break sites for NHEJ repair. AATF depletion or phosphorylation-deficient mutant impedes NHEJ in glioma stem-like cells.\",\n      \"method\": \"Co-immunoprecipitation (AATF-XRCC4), ATM kinase assay (Ser189 phosphorylation), site-directed mutagenesis, NHEJ assay, ubiquitination assay, xenograft model\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — kinase substrate identification, Co-IP, mutagenesis of phospho-site, NHEJ functional assay, and in vivo xenograft; multiple orthogonal methods\",\n      \"pmids\": [\"40436899\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Che-1/AATF interacts with subunits of the HAT module of SAGA complexes (ADA2A, ADA2B, and GCN5) in human cells, as demonstrated by co-immunoprecipitation and co-localization, with specific domains in ADA2 and GCN5 required for these interactions.\",\n      \"method\": \"Co-immunoprecipitation, yeast two-hybrid (domain mapping), co-localization\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP plus yeast two-hybrid domain mapping, single lab, two complementary methods\",\n      \"pmids\": [\"29232376\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"The Che-1/AATF promoter is TATA-less and contains a negative feedback element: Che-1 protein binds its own promoter and represses its own transcription.\",\n      \"method\": \"Reporter assay, chromatin immunoprecipitation (ChIP), promoter deletion analysis\",\n      \"journal\": \"Gene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — ChIP demonstrating auto-occupancy plus functional reporter assay, single lab\",\n      \"pmids\": [\"14636992\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"In pancreatic neuroendocrine tumors, ACSS2 activity induces histone H3/H4 hyperacetylation, which recruits AATF to co-regulate FasL transcription, specifically enhancing soluble FasL secretion. Secreted FasL triggers CD8+ T cell apoptosis via caspase-8/3 cascade, promoting immune evasion. ACSS2 and AATF act non-redundantly and synergistically in modulating FasL expression.\",\n      \"method\": \"ChIP, Western blot, ELISA (sFasL), co-immunoprecipitation, siRNA/overexpression, flow cytometry (T cell apoptosis)\",\n      \"journal\": \"Advanced science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP linking AATF to FasL promoter, functional sFasL secretion and T cell apoptosis assays, single lab\",\n      \"pmids\": [\"40791180\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"AATF/Che-1 is a multifunctional nuclear phosphoprotein that binds RNA polymerase II (and I), Rb, and multiple transcription factors; in unstressed cells it promotes proliferation by displacing HDAC1 from Rb/E2F and Sp1 complexes to activate E2F target genes and p21, while upon genotoxic stress it is phosphorylated by ATM/ATR, Chk2, MK2, CK2, and HIPK2 at distinct residues—triggering nuclear translocation, promoter switching from E2F targets to p53/p21 loci to activate cell-cycle arrest, or (when phosphorylated by MK2) binding PUMA/BAX/BAK promoters to repress p53-driven apoptosis—and is ultimately degraded via Pin1/HDM2/HIPK2-mediated ubiquitylation during apoptosis; AATF also forms the ANN complex (with NGDN and NOL10) in nucleoli to support 40S ribosomal subunit biogenesis and rRNA processing, localizes to paraspeckles via NEAT1_2 to resolve R-loops and suppress interferon activation, stabilizes XRCC4 for NHEJ repair (released by ATM-mediated Ser189 phosphorylation at DNA breaks), and interacts with partners including Par-4, Tau, Cdk5, c-Jun, AIF, SAGA subunits, and histones to regulate apoptosis, neuronal function, and chromatin acetylation across multiple cellular contexts.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"AATF/Che-1 is a multifunctional nuclear phosphoprotein that integrates transcriptional control of proliferation, the DNA damage response, and apoptosis [#2, #7]. It binds RNA polymerase II subunit hRPB11 and the retinoblastoma protein, and in proliferating cells it competes with HDAC1 for the Rb pocket—displacing HDAC1 from Rb/E2F complexes and from Sp1 sites to activate E2F targets and p21WAF1/CIP1 with accumulation of acetylated histone H3 [#1, #2, #3]. This pro-proliferative chromatin activity depends on direct histone binding, which requires CK2 phosphorylation at Ser316/Ser320/Ser321 and is reinforced by displacement of class I HDACs to sustain global histone acetylation and BRD4 recruitment [#26, #27]. Upon genotoxic stress, AATF is phosphorylated and stabilized by ATM/ATR and Chk2 and PARylated by PARP-1, accumulating and switching its promoter occupancy from rDNA and E2F loci to the TP53 and p21 promoters to activate growth-arrest p53 target genes and maintain cell-cycle checkpoints [#7, #14, #20, #25]. AATF restrains apoptosis through multiple routes: MK2 phosphorylation releases it to the nucleus where it represses p53-driven PUMA/BAX/BAK transcription, and it sustains XIAP and mutant p53 expression [#10, #13, #15]. Its own abundance is controlled by Pin1- and HIPK2-dependent priming of HDM2-mediated ubiquitylation and proteasomal degradation during apoptosis [#8, #18]. In the nucleolus AATF supports ribosome biogenesis, forming the ANN complex with NGDN and NOL10 required for 18S rRNA maturation and 40S subunit synthesis and promoting RNA Pol I/UBF-dependent rDNA transcription [#21, #25]. It additionally stabilizes the NHEJ factor XRCC4 until ATM phosphorylation at Ser189 releases it for recruitment to DNA breaks, and localizes to paraspeckles via NEAT1_2 to resolve R-loops and suppress interferon activation [#28, #31]. These activities make AATF a driver of tumor proliferation in vivo, including Kras-driven lung adenocarcinoma in a p53-dependent manner and plasma-cell multiple myeloma [#23, #26].\",\n  \"teleology\": [\n    {\n      \"year\": 1999,\n      \"claim\": \"Establishing AATF's existence and its first functional theme—antagonism of apoptosis—anchored all later work by linking it to a death kinase and to transcription.\",\n      \"evidence\": \"Yeast two-hybrid against Dlk/ZIP kinase plus Gal4-BD transactivation and apoptosis-interference assays\",\n      \"pmids\": [\"10580117\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No direct DNA targets identified\", \"Domain requirements for transactivation undefined\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Defining AATF as an HDAC1-displacing cofactor at Rb/E2F and Sp1 sites explained how it drives proliferation and p53-independent p21 activation through chromatin acetylation.\",\n      \"evidence\": \"Co-IP, ChIP, and RNAi in Rb-proficient vs Rb-deficient cells; ChIP and reporter assays on the p21/Sp1 promoter\",\n      \"pmids\": [\"10783144\", \"12450794\", \"12847090\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct vs indirect promoter recruitment mechanism not fully resolved\", \"Phospho-regulation of HDAC1 competition not yet known at this stage\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Identifying ATM/ATR/Chk2 phosphorylation as a switch that stabilizes AATF and redirects it to TP53/p21 promoters established its role as a damage-responsive checkpoint activator.\",\n      \"evidence\": \"Co-IP of kinase-substrate, ChIP of promoter recruitment, cell-cycle flow cytometry, RNAi\",\n      \"pmids\": [\"17157788\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Exact phospho-residues not mapped\", \"Mechanism of promoter selectivity unresolved\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Mapping Pin1-primed, HDM2-mediated ubiquitylation showed how AATF protein levels are downregulated to permit apoptosis, defining its turnover control.\",\n      \"evidence\": \"Co-IP, ubiquitination assay, half-life measurement, Pin1-binding mutant, apoptosis assay\",\n      \"pmids\": [\"17468107\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Phospho-sites recognized by Pin1 not defined here\", \"Other E3 ligases not excluded\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Demonstrating PARP-1-dependent PARylation as an ATM-independent route to AATF stabilization revealed a parallel damage signal converging on its promoter occupancy.\",\n      \"evidence\": \"Co-IP, in vitro PARylation, ChIP at p21 promoter, PARP inhibitor/knockout and ATM-inhibitor epistasis\",\n      \"pmids\": [\"21317046\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"PARylation acceptor residues not mapped\", \"Interplay with phosphorylation marks unresolved\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Identifying MK2 phosphorylation as the trigger for cytoplasm-to-nucleus translocation and repression of PUMA/BAX/BAK promoters explained how AATF actively blocks p53-driven apoptosis and confers chemoresistance.\",\n      \"evidence\": \"MK2 kinase assay, MRLC3 Co-IP, fractionation, ChIP, phospho-mimic mutagenesis, xenografts\",\n      \"pmids\": [\"22909821\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How nuclear AATF selects pro-apoptotic vs growth-arrest p53 loci unclear\", \"Relationship to ATM/Chk2 signaling not integrated\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Direct p53 binding forming a p53/Brca1 ternary complex that biases transactivation toward growth-arrest genes, confirmed in Che-1(+/-) mice, established AATF as a determinant of the arrest-versus-death p53 decision.\",\n      \"evidence\": \"Co-IP, genome-wide ChIP-seq, heterozygous knockout mice with irradiation phenotype\",\n      \"pmids\": [\"25996291\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of p53 target selectivity unknown\", \"How interaction is lost during apoptosis not mechanistically defined\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Adding HIPK2 as a stress kinase that primes AATF for HDM2-dependent degradation completed the picture of how genotoxic signaling tips AATF abundance toward apoptosis.\",\n      \"evidence\": \"Co-IP, in vitro kinase assay, ubiquitylation assay, RNAi, apoptosis rescue\",\n      \"pmids\": [\"25210797\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"HIPK2 target residues on AATF not specified\", \"Coordination with Pin1/CK2 phosphorylation unresolved\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Discovery of the ANN complex (AATF-NGDN-NOL10) localized a major non-transcriptional role to nucleolar ribosome biogenesis.\",\n      \"evidence\": \"IP-MS, domain mapping, RNAi with rRNA processing analysis, immunofluorescence\",\n      \"pmids\": [\"27599843\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular function of AATF within the complex (catalytic vs scaffold) undefined\", \"Link between ribosome biogenesis and stress signaling not established\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Linking AATF to RNA Pol I/UBF-driven rDNA transcription, with damage-induced re-localization from rDNA to TP53, unified its nucleolar and checkpoint roles into a stress-responsive promoter switch.\",\n      \"evidence\": \"Co-IP with Pol I/UBF, ChIP at rDNA, reporter assay, pre-rRNA Northern, immunofluorescence\",\n      \"pmids\": [\"32421830\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Signal driving rDNA-to-TP53 relocalization not defined\", \"Relationship to ANN complex function unclear\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Mapping CK2 phosphorylation at Ser316/320/321 as a requirement for histone H3 binding provided the molecular basis for AATF's pro-proliferative chromatin acetylation activity.\",\n      \"evidence\": \"In vitro CK2 kinase assay, mutagenesis, 2D gel/MS phospho-site mapping, Co-IP, proliferation assay\",\n      \"pmids\": [\"33186461\", \"34266450\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural mode of histone engagement undefined\", \"Whether other kinases regulate this interaction unknown\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Localization to NEAT1_2 paraspeckles and R-loops, with R-loop accumulation and interferon induction upon depletion, revealed a genome-stability role coupling AATF to innate immune suppression.\",\n      \"evidence\": \"NEAT1 RNA-IP, S9.6 R-loop dot blot/IF, RNAi, RNA-seq IFN signature, ChIP\",\n      \"pmids\": [\"35929179\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct enzymatic activity on R-loops not demonstrated\", \"Mechanism of R-loop resolution by AATF unresolved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Identifying ATM-Ser189 phosphorylation as the release signal that frees stabilized XRCC4 for recruitment to breaks placed AATF directly within NHEJ repair.\",\n      \"evidence\": \"Co-IP, ATM kinase assay, phospho-site mutagenesis, NHEJ and ubiquitination assays, xenograft\",\n      \"pmids\": [\"40436899\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Generality across cell types beyond glioma stem-like cells untested\", \"Coordination with AATF's transcriptional damage response unresolved\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How AATF's many post-translational modifications (ATM/ATR, Chk2, MK2, CK2, HIPK2 phosphorylation; PARylation; ubiquitylation) are hierarchically integrated to dictate its localization and the choice among proliferation, arrest, apoptosis suppression, and repair remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No unified model of combinatorial PTM control\", \"No structural framework for its distinct binding interfaces\", \"Tissue-specific determinants of functional output undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [0, 2, 3, 7, 20, 25]},\n      {\"term_id\": \"GO:0042393\", \"supporting_discovery_ids\": [26, 27]},\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [28]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [2, 3, 21, 31]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [2, 25, 31]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [16, 21, 25]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [2, 7, 9, 20]},\n      {\"term_id\": \"GO:0005654\", \"supporting_discovery_ids\": [28]},\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [17]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [4, 15]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [7, 14, 31]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [2, 7, 17, 20]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [8, 15, 16, 18, 29]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [2, 3, 25, 26]},\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [21, 25]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [23, 24, 26]},\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [12, 19, 22, 28]}\n    ],\n    \"complexes\": [\n      \"ANN complex (AATF-NGDN-NOL10)\",\n      \"SAGA HAT module\",\n      \"RNA polymerase I/UBF rDNA transcription complex\"\n    ],\n    \"partners\": [\n      \"RB1\",\n      \"TP53\",\n      \"HDAC1\",\n      \"HDM2\",\n      \"PIN1\",\n      \"PARP1\",\n      \"XRCC4\",\n      \"NGDN\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":9,"faith_total":9,"faith_pct":100.0}}