{"gene":"PRKDC","run_date":"2026-06-10T06:43:35","timeline":{"discoveries":[{"year":1994,"finding":"The DNA-PK holoenzyme (Ku protein + p350/DNA-PKcs) assembles on double-stranded DNA: Ku binds first to free DNA ends, then recruits p350 to form a catalytically active complex. Reconstitution experiments showed the complex forms only in the presence of double-stranded DNA.","method":"Immunoprecipitation with anti-Ku antibodies, reconstitution with purified components, catalytic activity assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution with purified components plus functional activity assay; foundational biochemical demonstration","pmids":["8041718"],"is_preprint":false},{"year":1992,"finding":"DNA-PKcs is a nuclear serine/threonine protein kinase activated by direct interaction with DNA (not using DNA as template/substrate) and phosphorylates DNA-binding proteins including transcription factors, suggesting a role in modulating transcriptional activity.","method":"Biochemical kinase assays, nuclear fractionation","journal":"Critical reviews in eukaryotic gene expression","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — early biochemical characterization with kinase assays; single lab review summarizing foundational work","pmids":["1486241"],"is_preprint":false},{"year":1995,"finding":"DNA-PKcs (p350) is the gene responsible for the murine SCID defect: p350 and the SCID-complementing gene co-localize to human chromosome 8q11, chromosomal fragments expressing p350 complement the SCID phenotype, and p350 protein is greatly reduced in SCID mouse cells.","method":"Chromosomal complementation, co-localization mapping, western blot of SCID-derived cells","journal":"Science (New York, N.Y.)","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic complementation plus protein-level analysis; independently confirmed in companion paper (PMID:7855602)","pmids":["7855601","7855602"],"is_preprint":false},{"year":1995,"finding":"Absence of the p350 subunit of DNA-PK in the radiosensitive human glioma cell line M059J is associated with defective DNA double-strand break repair, demonstrating that DNA-PK kinase activity is required for DSB repair.","method":"Immunoblot for p350 expression, DNA-PK kinase activity assay, DSB repair assay in human cell lines","journal":"Science (New York, N.Y.)","confidence":"High","confidence_rationale":"Tier 2 / Strong — natural loss-of-function human cell line with direct protein-level and activity measurements, replicated alongside PMID:7855601","pmids":["7855602"],"is_preprint":false},{"year":2005,"finding":"The Artemis:DNA-PKcs complex cleaves a wide range of DNA substrates containing single-to-double-strand transitions (heterologous loops, stem-loops, flaps, gapped substrates) near the transition region; this versatile endonuclease activity is activated by DNA-PKcs binding and phosphorylation of Artemis.","method":"In vitro endonuclease assays with purified Artemis:DNA-PKcs complex on defined DNA substrates","journal":"DNA repair","confidence":"High","confidence_rationale":"Tier 1 / Moderate — reconstituted in vitro endonuclease assay with purified proteins on multiple substrate types; single lab","pmids":["15936993"],"is_preprint":false},{"year":2007,"finding":"DNA-PKcs accumulates at DSB sites in a Ku80-dependent manner. Kinase activity and autophosphorylation status do not affect initial recruitment but do regulate the stability of DNA-PKcs binding to DNA ends; impaired autophosphorylation leads to prolonged retention at unrepaired DSBs, suggesting autophosphorylation facilitates NHEJ by destabilizing the interaction with DNA ends.","method":"Laser-induced DSB formation in live cells, FRAP (fluorescence recovery after photobleaching), live-cell imaging of GFP-DNA-PKcs","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — live-cell FRAP with kinase-dead and phospho-mutant constructs; multiple orthogonal approaches in single study","pmids":["17438073"],"is_preprint":false},{"year":2010,"finding":"DNA-PKcs autophosphorylation at the ABCDE cluster regulates access of repair factors to DNA ends. Kinase-dead DNA-PKcs (DNA-PKcs-KR) cells show hyperrecombination 2-3 fold above DNA-PKcs null, and ATM-dependent phosphorylation of DNA-PKcs-KR contributes to this hyperrecombination phenotype; DNA-PKcs and ATM coordinately regulate the NHEJ/HR pathway choice.","method":"Direct repeat HR assay with I-SceI nuclease, epistasis analysis with kinase-dead and null mutants, RAD51 foci analysis","journal":"DNA repair","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic epistasis with multiple defined mutant cell lines, two orthogonal phenotypic readouts (HR assay and RAD51 foci)","pmids":["19535303"],"is_preprint":false},{"year":2010,"finding":"DNA-PKcs regulates Artemis single-stranded DNA endonuclease activity: purified Artemis has intrinsic ssDNA endonuclease activity that is stimulated by DNA-PKcs; the divalent cation and sequence dependence matches that of the Artemis:DNA-PKcs double-stranded endonuclease activity.","method":"In vitro endonuclease assay with highly purified Artemis, addition of DNA-PKcs, antibody modulation","journal":"DNA repair","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution assay with purified proteins; single lab but multiple orthogonal methods","pmids":["20117966"],"is_preprint":false},{"year":2010,"finding":"DNA-PKcs selectively stimulates WRN helicase activity (but not WRN exonuclease) on telomere D-loop model substrates in vitro, and DNA-PKcs knockdown reduces telomeric G-tail length in vivo; this effect is reversed by WRN helicase overexpression, indicating cooperative roles of DNA-PKcs and WRN in maintaining telomeric G-tails.","method":"In vitro helicase/exonuclease assay with purified DNA-PKcs and WRN, siRNA knockdown, telomere G-tail length measurement","journal":"Aging","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro stimulation assay plus in vivo knockdown rescue; single lab","pmids":["20519774"],"is_preprint":false},{"year":2014,"finding":"DNA damage triggers DNA-PK-dependent phosphorylation of GOLPH3, which increases GOLPH3 interaction with MYO18A, applying tensile force to the Golgi and resulting in Golgi dispersal throughout the cytoplasm. Depletion of DNA-PK, GOLPH3, or MYO18A reduces survival after DNA damage.","method":"siRNA knockdown, phospho-specific antibodies, co-immunoprecipitation, fluorescence microscopy of Golgi morphology, cell survival assays","journal":"Cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal co-IP, functional knockdown with specific phenotypic readout, multiple orthogonal methods in single rigorous study","pmids":["24485452"],"is_preprint":false},{"year":2016,"finding":"DNA-PKcs phosphorylates ATM at specific sites, inhibiting ATM activity. Chemical inhibition or genetic deletion of DNA-PKcs leads to hyperactive ATM; pre-incubation of ATM with active DNA-PKcs reduces ATM activity in vitro. Phospho-mimetic mutations at DNA-PKcs target sites in ATM inhibit ATM activity and impair ATM signaling after DNA damage.","method":"In vitro kinase assay with purified proteins, mutagenesis of ATM phosphorylation sites, DNA damage signaling assays in human cells with DNA-PKcs KO/inhibition","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro kinase assay plus mutagenesis plus genetic epistasis; multiple orthogonal methods","pmids":["27939942"],"is_preprint":false},{"year":2002,"finding":"DNA-PKcs functionally interacts with telomerase in telomere length maintenance: Terc(-/-)/DNA-PKcs(-/-) double-knockout mice show accelerated telomere shortening compared to Terc(-/-) alone. DNA-PKcs is also required for end-to-end chromosome fusions and apoptosis triggered by critically short telomeres.","method":"Double-knockout mouse model, telomere length analysis (FISH/TRF), cytogenetic analysis","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic epistasis in mouse model with quantitative telomere length measurements and chromosome fusion analysis","pmids":["12426399"],"is_preprint":false},{"year":2009,"finding":"Phosphorylation of DNA-PKcs at the Thr-2609 cluster is a critical event for proper telomere end-processing; DNA-PKcs deficiency leads to uncapped telomeres that are inappropriately processed as DSBs, participating in spontaneous and radiation-induced telomere-DSB fusions requiring ligase IV.","method":"Cytogenetic analysis in DNA-PKcs-deficient mouse and human cells, phospho-specific immunoblot, irradiation experiments","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — defined phosphorylation cluster linked to telomere capping phenotype; single lab with multiple methods","pmids":["19244120"],"is_preprint":false},{"year":2017,"finding":"Cryo-EM structure of the human DNA-PK holoenzyme shows DNA-PKcs, KU70, KU80, and DNA forming a 650-kDa heterotetramer; DNA-PKcs N-terminal α-solenoid adopts a double-ring fold; together with KU70/80, DNA-PKcs forms a DNA-binding tunnel that protects ~30-bp DNA end. KU70/80 and DNA coordinately induce conformational changes in DNA-PKcs and allosterically stimulate its kinase activity.","method":"Cryo-electron microscopy (6.6 Å resolution), biochemical activity assays","journal":"Cell research","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure with functional validation; independently confirmed by companion structure paper (PMID:28652322)","pmids":["28840859","28652322"],"is_preprint":false},{"year":2017,"finding":"Cryo-EM structures of human DNA-PKcs (4.4 Å) and DNA-PK holoenzyme (5.8 Å) reveal that DNA-PKcs has three segments: N-terminal region (arm and bridge), circular cradle, and head with kinase domain; the C-terminal globular domain of Ku80 interacts with the arm of DNA-PKcs.","method":"Cryo-electron microscopy structural analysis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Strong — high-resolution cryo-EM structure, replicated by companion paper (PMID:28840859)","pmids":["28652322"],"is_preprint":false},{"year":2020,"finding":"Cryo-EM structures of DNA-PKcs bound to DNA end and in complex with Ku70/80-DNA in inactive and activated states (3.7 Å overall; 3.2 Å FATKIN) reveal that kinase activation involves stretching and twisting within solenoid segments, loosening DNA-end binding. This structural plasticity of HEAT repeats represents a regulatory mechanism for PIKK family kinases.","method":"Cryo-electron microscopy, structural comparison of inactive vs. activated states","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — high-resolution cryo-EM with multiple conformational states; single rigorous study","pmids":["33385326"],"is_preprint":false},{"year":2020,"finding":"DNA-PKcs has a KU-dependent role in rRNA processing and haematopoiesis: KU drives assembly of DNA-PKcs on cellular RNAs including U3 snRNA; U3 snRNA activates purified DNA-PK and triggers DNA-PKcs phosphorylation at T2609. DNA-PK resides in nucleoli in an rRNA-dependent manner and co-purifies with the small subunit processome. Blocking T2609 cluster phosphorylation (but not S2056 cluster) causes defects in 18S rRNA processing and bone marrow failure.","method":"Mouse knockin models (kinase-dead, T2609A), purified protein activation assays with U3 snRNA, nucleolar fractionation, co-purification with SSU processome, ribosome profiling","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple mouse genetic models plus in vitro reconstitution with RNA; multiple orthogonal methods in single rigorous study","pmids":["32103174"],"is_preprint":false},{"year":2020,"finding":"Human DNA-PK functions as a sensor in a STING-independent DNA sensing pathway (SIDSP): DNA-PK drives a broad antiviral response; HSPA8/HSC70 is identified as a target of inducible phosphorylation downstream of DNA-PK in this pathway. Viral proteins E1A (adenovirus) and ICP0 (HSV-1) block this response.","method":"Genetic deletion and inhibition of DNA-PK, phosphoproteomics identifying HSPA8/HSC70 as substrate, viral infection assays, antiviral response measurement","journal":"Science immunology","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic sensor identification, specific substrate phosphorylation, viral antagonist characterization; multiple orthogonal methods","pmids":["31980485"],"is_preprint":false},{"year":2020,"finding":"DNA-PK phosphorylates cGAS and suppresses its enzymatic activity. DNA-PK deficiency reduces cGAS phosphorylation and promotes antiviral innate immune responses. Cells from DNA-PKcs-deficient mice or patients with PRKDC missense mutations exhibit an inflammatory gene expression signature.","method":"In vitro kinase assay with DNA-PKcs and cGAS, co-immunoprecipitation, genetic mouse models, patient-derived cells","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro kinase assay plus genetic models plus patient cells; multiple orthogonal methods","pmids":["33273464"],"is_preprint":false},{"year":2021,"finding":"Cryo-EM structures of DNA-PK bound to DNA ends before and after autophosphorylation, and in complex with Artemis and a DNA hairpin, reveal a functional switch: open DNA ends inhibit cis-autophosphorylation of the ABCDE cluster but activate phosphorylation of other targets; hairpin ends promote ABCDE cis-autophosphorylation. Phosphorylation of four Thr residues in ABCDE causes gross structural rearrangement widening the DNA-binding groove for Artemis recruitment and hairpin cleavage. Artemis locks DNA-PK in a kinase-inactive state.","method":"Cryo-EM structural analysis of multiple states, in vitro kinase and nuclease assays","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structures of multiple functional states combined with biochemical validation; single rigorous study with multiple orthogonal methods","pmids":["34936881"],"is_preprint":false},{"year":2019,"finding":"DNA-PKcs kinase activity is required for initiation of the DDR immediately after DSB induction: it drives phosphorylation of chromatin factors H2AX and KAP1, promotes local chromatin decondensation near DSB sites, and facilitates recruitment of DDR machinery. Loss of DNA-PKcs kinase activity markedly decreases DDR factor recruitment to DSBs.","method":"Kinase-domain inactivating human cell line, ionizing radiation, γH2AX and KAP1 phosphorylation assays, chromatin decondensation measurement, recruitment kinetics of DDR factors","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean kinase-dead cell line with multiple downstream readouts; multiple orthogonal methods","pmids":["31396623"],"is_preprint":false},{"year":2020,"finding":"DNA-PKcs is neddylated at its kinase domain by the E2-conjugating enzyme UBE2M and E3 ligase HUWE1; inhibition of HUWE1-dependent neddylation impairs DNA-PKcs autophosphorylation at Ser2056 and reduces NHEJ efficiency.","method":"Immunoprecipitation, co-immunoprecipitation, HUWE1/UBE2M knockdown, NHEJ reporter assay, phospho-Ser2056 immunoblot","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP plus functional NHEJ assay; single lab with multiple methods but no structural validation of neddylation site","pmids":["32457294"],"is_preprint":false},{"year":2022,"finding":"Cryo-EM structural analysis of the basal (pre-activated) Artemis:DNA-PKcs complex shows the Artemis catalytic domain is positioned externally to DNA-PKcs prior to ABCDE autophosphorylation; both Artemis catalytic and regulatory domains interact with the N-HEAT and FAT domains of DNA-PKcs. A mutually exclusive binding site for Artemis and XRCC4 on DNA-PKcs was defined, and an XRCC4 peptide disrupts the Artemis:DNA-PKcs complex.","method":"Cryo-EM structural analysis, agarose-acrylamide gel complex stabilization, peptide competition assay","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Moderate — cryo-EM structure plus biochemical interaction mapping; single lab","pmids":["35801871"],"is_preprint":false},{"year":2022,"finding":"Cryo-EM structures of DNA-PKcs in three distinct dimeric conformations represent transition states during NHEJ: upon autophosphorylation, the long-range synaptic complex undergoes conformational change with both Ku and DNA-PKcs rotating outward to promote DNA break exposure and DNA-PKcs dissociation.","method":"Single-particle cryo-electron microscopy of NHEJ complexes at different stages","journal":"Science advances","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structures of multiple conformational states; consistent with companion structural studies","pmids":["37256947"],"is_preprint":false},{"year":2022,"finding":"Cryo-EM structures of human DNA-PKcs with ATPγS and four inhibitors (wortmannin, NU7441, AZD7648, M3814) reveal the ATP binding mode and show that inhibitor binding causes movement of the PIKK regulatory domain (PRD), revealing a connection between the p-loop and PRD conformations. Inhibitors function through direct ATP competition and do not negatively allosterically affect holoenzyme assembly.","method":"Cryo-EM structural analysis, electrophoretic mobility shift assay (EMSA) for holoenzyme assembly","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 / Strong — high-resolution cryo-EM with four inhibitors plus biochemical EMSA validation; single rigorous study","pmids":["34987222"],"is_preprint":false},{"year":2022,"finding":"DNA-PKcs promotes fork reversal at stressed replication forks in a manner independent of its NHEJ role; cells lacking DNA-PKcs activity show increased DNA damage during S-phase and sensitivity to replication stress. Prevention of fork reversal by DNA-PKcs inhibition restores chemotherapy sensitivity in BRCA2-deficient tumors with acquired PARP inhibitor resistance.","method":"Electron microscopy of replication intermediates, DNA fiber assay, DNA-PKcs inhibitor/knockout, BRCA2-deficient tumor model","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct visualization of replication forks by EM combined with genetic/pharmacological perturbation; multiple orthogonal methods","pmids":["36130596"],"is_preprint":false},{"year":2022,"finding":"DNA-PKcs interacts with and phosphorylates Fis1 at Thr34 in its TQ motif, increasing Fis1 affinity for Drp1 and inducing mitochondrial fragmentation; knockin mice with non-phosphorylatable T34A Fis1 show improved renal function and reduced mitochondrial fragmentation in acute kidney injury. Cytoplasmic localization of DNA-PKcs was detected in injured kidney tissues.","method":"Co-immunoprecipitation, in vitro kinase assay, Fis1-T34A knockin mice, mitochondrial morphology analysis, human patient urinary sediment analysis","journal":"Science signaling","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro kinase assay with identified phosphorylation site, knockin mouse validation, and human patient correlation; multiple orthogonal methods","pmids":["35290083"],"is_preprint":false},{"year":2022,"finding":"Physical ARTEMIS:DNA-PKcs interaction is necessary for V(D)J recombination: the L3062R pathogenic mutation in DNA-PKcs impairs physical interaction with Artemis; specific mutations in Artemis (in two conserved regions) that disrupt interaction with DNA-PKcs impair V(D)J recombination. Minimal interaction fragments were mapped: 42 aa from FAT region 2 of DNA-PKcs (PKcs3041-3082) and 26 aa from Artemis (ARM378-403).","method":"Mutagenesis, co-immunoprecipitation, V(D)J recombination assay, domain mapping with minimal fragments","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 2 / Strong — mutagenesis with functional V(D)J assay and precise domain mapping; multiple orthogonal methods","pmids":["35150269"],"is_preprint":false},{"year":2023,"finding":"DNA-PKcs directly interacts with mitochondrial proteins ANT2 and VDAC2 forming the DAV complex, which supports ADP-ATP exchange across mitochondrial membranes to sustain oxidative phosphorylation and membrane potential. The DAV complex dissociates in response to oxidative stress, attenuating ADP-ATP exchange; dissociation is mediated by ATM-dependent phosphorylation of DNA-PKcs at the Thr2609 cluster.","method":"Co-immunoprecipitation, mitochondrial fractionation, membrane potential assays, Seahorse metabolic analysis, DNA-PKcs-deficient cell lines, ATM kinase assay","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — co-IP identifying novel complex, functional metabolic readouts, ATM-mediated phosphorylation mechanism; multiple orthogonal methods in single study","pmids":["36727301"],"is_preprint":false},{"year":2023,"finding":"DNA-PK and TRF2 cooperate to repress MRN-initiated resection at leading-end (blunt) telomeres: DNA-PK represses MRN-dependent long-range resection, while the iDDR of TRF2 inhibits MRN-CtIP endonuclease activity that would otherwise cleave DNA-PK off blunt telomere ends. AlphaFold-Multimer predicts conserved iDDR association with Rad50, potentially interfering with CtIP binding.","method":"In vitro resection assays, in vivo telomere resection analysis, AlphaFold-Multimer structural prediction with experimental validation","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vitro and in vivo functional assays demonstrating repression mechanism; multiple orthogonal approaches","pmids":["37653239"],"is_preprint":false},{"year":2021,"finding":"DNA-PKcs (DNA-PK catalytic subunit) phosphorylates SOX2 at S251, stabilizing SOX2 by preventing WWP2-mediated ubiquitination and promoting glioma stem cell maintenance. Upon DNA damage, the DNA-PK complex dissociates from SOX2, allowing WWP2 interaction and SOX2 degradation, triggering differentiation.","method":"Mass spectrometry of SOX2-binding proteins, co-immunoprecipitation, site-directed mutagenesis of S251, ubiquitination assays, in vitro and in vivo (xenograft) DNA-PKcs inhibition","journal":"Science translational medicine","confidence":"High","confidence_rationale":"Tier 1 / Strong — MS identification of interaction, mutagenesis of phosphorylation site, ubiquitination assay, in vivo xenograft; multiple orthogonal methods","pmids":["34193614"],"is_preprint":false},{"year":2020,"finding":"DNA-PKcs kinase activity and autophosphorylation regulate kinase complex conformation and dissociation during NHEJ; expression of catalytically inactive DNA-PKcs causes more genomic instability than loss of the protein itself (structural function), because kinase-dead DNA-PKcs persists at DNA lesions and alters repair pathway choice.","method":"Mouse models expressing kinase-dead DNA-PKcs, genomic instability assays, comparison to protein-null models","journal":"Cell & bioscience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple mouse genetic models compared; review synthesizing results from multiple labs but mechanistic model from primary mouse model data","pmids":["32015826"],"is_preprint":false},{"year":2019,"finding":"DNA-PARylation of DNA-PKcs by PARP1 regulates DNA-PK activity: DNA-PKcs is PARylated after DNA damage, PARP inhibition (olaparib) prevents DNA-PKcs detachment from chromatin and maintains DNA-PKcs Ser2056 autophosphorylation; olaparib and DNA-PK inhibition synergize to suppress cell survival.","method":"Immunoprecipitation, immunofluorescence, chromatin fractionation, phospho-Ser2056 immunoblot, cell survival assays","journal":"Molecular medicine reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP and functional chromatin dissociation assay; single lab, multiple methods","pmids":["31485633"],"is_preprint":false},{"year":2020,"finding":"Activation of DNA-PK by hairpinned DNA ends is substrate-specific: hairpinned DNA ends do not activate DNA-PK toward p53, XRCC4, XLF, or HSP90, but robustly stimulate ABCDE cluster autophosphorylation, which is required for Artemis activation. This reveals a multi-step mechanism of kinase activation.","method":"In vitro kinase assays with defined DNA substrates (hairpinned vs open ends), comparison across multiple substrates","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro kinase assay with multiple substrates revealing substrate-specific activation; single lab","pmids":["32716029"],"is_preprint":false},{"year":2021,"finding":"The CRL4A-DTL ubiquitin ligase complex targets DNA-PKcs for nuclear proteasomal degradation; overexpression of CUL4A or DTL reduces NHEJ repair efficiency and increases DSB accumulation, leading to genomic instability and malignant transformation.","method":"Co-immunoprecipitation, ubiquitination assay, NHEJ reporter assay, γH2AX measurement, overexpression in normal cells","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP with functional NHEJ assay and cell transformation; single lab, multiple methods","pmids":["33627782"],"is_preprint":false},{"year":2024,"finding":"PRKDC recruits GDE2 to enhance stability of GNAS protein, which activates AKT phosphorylation, conferring doxorubicin resistance in osteosarcoma. The PRKDC-GDE2-GNAS-AKT regulatory axis was identified by a kinome-wide CRISPR screen.","method":"CRISPR kinome screen, co-immunoprecipitation of PRKDC and GDE2, GNAS stability assays, AKT phosphorylation analysis, xenograft and organoid models","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR screen plus co-IP and in vivo validation; single lab, multiple methods","pmids":["38900943"],"is_preprint":false},{"year":2018,"finding":"DNA-PKcs has a kinase-dependent function in suppressing microhomology-mediated end joining (MMEJ) during class switch recombination (CSR) and a structural (kinase-independent) role in orientation of CSR; kinase-dead DNA-PKcs severely compromises CSR to IgG1 while DNA-PKcs deletion does not, revealing distinct structural and catalytic roles.","method":"Mouse B cell models with kinase-dead vs. null DNA-PKcs, high-throughput sequencing of CSR junctions, translocation analysis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — comparative genetic mouse models with high-throughput junction sequencing; multiple orthogonal methods","pmids":["30072430"],"is_preprint":false},{"year":2001,"finding":"BCR-ABL down-regulates DNA-PKcs via proteasome-dependent degradation that requires BCR-ABL tyrosine kinase activity, resulting in marked DNA repair deficiency and increased sensitivity to ionizing radiation.","method":"Stable and inducible BCR-ABL expression in hematopoietic cells, proteasome inhibitor experiments, western blot, irradiation sensitivity assays","journal":"Blood","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pharmacological dissection of proteasome dependence and tyrosine kinase requirement; single lab, multiple methods","pmids":["11264175"],"is_preprint":false},{"year":2012,"finding":"Upon ionizing radiation, nuclear EGFR associates with DNA-PK, which phosphorylates PNPase (polynucleotide phosphorylase) at Ser-776; phospho-mimetic S776D PNPase has impaired ribonuclease activity, while the non-phosphorylatable S776A mutant retains ribonuclease activity and degrades c-MYC mRNA, affecting radioresistance.","method":"Co-immunoprecipitation, site-directed mutagenesis of PNPase, in vitro ribonuclease assays, knockdown experiments","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro kinase assay with mutagenesis of substrate residue, functional ribonuclease assay; single lab, multiple orthogonal methods","pmids":["22815474"],"is_preprint":false},{"year":2020,"finding":"DNA-PKcs promotes cardiac ischemia-reperfusion injury through direct interaction with BI-1 (Bax inhibitor-1), promoting BI-1 degradation without affecting BI-1 transcription. Loss of DNA-PKcs stabilizes BI-1, protecting mitochondria; concurrent BI-1 knockout abrogates the cardioprotection of DNA-PKcs deletion.","method":"Cardiomyocyte-specific DNA-PKcs knockout mice, co-immunoprecipitation, double-knockout epistasis, mitochondrial function assays","journal":"Basic research in cardiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP plus genetic epistasis in mouse model; single lab","pmids":["31919590"],"is_preprint":false}],"current_model":"DNA-PKcs (PRKDC) is a large nuclear serine/threonine PIKK-family kinase that, upon recruitment to DNA double-strand breaks by the Ku70/80 heterodimer, forms the active DNA-PK holoenzyme; it autophosphorylates at distinct clusters (ABCDE at T2609, S2056) to undergo sequential conformational changes that regulate DNA-end protection versus processing, recruits and activates the Artemis nuclease for hairpin opening in V(D)J recombination, negatively regulates ATM activity through direct phosphorylation, and has additional non-canonical roles including phosphorylating cGAS to suppress innate immunity, phosphorylating GOLPH3 to trigger Golgi dispersal after DNA damage, phosphorylating Fis1 to control mitochondrial fragmentation, acting in an RNA-dependent capacity in ribosome biogenesis (18S rRNA processing) via KU-directed assembly on U3 snRNA, stabilizing SOX2 in glioma stem cells, promoting replication fork reversal during replication stress, and regulating oxidative phosphorylation through a mitochondrial ANT2/VDAC2 complex whose dissociation is controlled by ATM-mediated phosphorylation of the T2609 cluster."},"narrative":{"mechanistic_narrative":"PRKDC encodes DNA-PKcs, a nuclear PIKK-family serine/threonine kinase that serves as the catalytic engine of non-homologous end joining (NHEJ), the principal pathway for repairing DNA double-strand breaks (DSBs) [PMID:8041718, PMID:7855602]. Recruited to free DNA ends by the Ku70/Ku80 heterodimer, DNA-PKcs assembles the catalytically active DNA-PK holoenzyme only in the presence of double-stranded DNA, with Ku and DNA coordinately inducing conformational changes that allosterically stimulate the kinase and protect ~30 bp of the DNA end within a binding tunnel [PMID:8041718, PMID:28840859, PMID:28652322, PMID:33385326]. Activation is governed by autophosphorylation: hairpinned versus open DNA ends differentially trigger phosphorylation of the ABCDE (T2609) cluster, driving a structural rearrangement that widens the DNA-binding groove, destabilizes end binding to promote NHEJ progression, and licenses recruitment of the Artemis nuclease, which DNA-PKcs activates to open hairpins and cleave single-to-double-strand transitions during V(D)J recombination [PMID:17438073, PMID:34936881, PMID:32457294, PMID:32716029, PMID:15936993, PMID:35150269]. DNA-PKcs initiates the broader DNA damage response by phosphorylating chromatin factors H2AX and KAP1 and promoting chromatin decondensation at breaks, and it shapes repair-pathway choice by negatively regulating ATM through direct phosphorylation and by suppressing hyper-recombination [PMID:31396623, PMID:27939942, PMID:19535303]; catalytically inactive DNA-PKcs persists at lesions and causes greater genomic instability than protein loss, revealing distinct structural and catalytic functions [PMID:32015826, PMID:30072430]. Loss of DNA-PKcs underlies the murine SCID immunodeficiency phenotype [PMID:7855601, PMID:7855602]. Beyond canonical repair, DNA-PKcs has KU-directed roles in 18S rRNA processing within the small-subunit processome via U3 snRNA-driven assembly and T2609 phosphorylation [PMID:32103174], acts as a cytosolic DNA sensor driving STING-independent antiviral responses while phosphorylating and suppressing cGAS [PMID:31980485, PMID:33273464], and executes signaling functions at the Golgi (GOLPH3-MYO18A-mediated dispersal after damage) [PMID:24485452], mitochondria (Fis1-driven fragmentation and an ANT2/VDAC2 oxidative-phosphorylation complex) [PMID:35290083, PMID:36727301], telomeres [PMID:12426399, PMID:19244120, PMID:37653239], and replication forks (promoting fork reversal independent of NHEJ) [PMID:36130596].","teleology":[{"year":1994,"claim":"Established that DNA-PK is an assembled holoenzyme requiring Ku and double-stranded DNA, defining the order of complex formation that underlies break recognition.","evidence":"Anti-Ku immunoprecipitation and reconstitution with purified components plus catalytic activity assay","pmids":["8041718"],"confidence":"High","gaps":["Did not resolve the structural basis of activation","Did not identify physiological substrates beyond the complex itself"]},{"year":1995,"claim":"Linked DNA-PKcs genetically to immunodeficiency and DSB repair, showing the gene is responsible for the murine SCID defect and required for repair in human cells.","evidence":"Chromosomal complementation, co-localization mapping, and immunoblot/repair assays in SCID and M059J cells","pmids":["7855601","7855602"],"confidence":"High","gaps":["Did not define the molecular step of NHEJ requiring DNA-PKcs","Catalytic versus structural contribution unresolved"]},{"year":2005,"claim":"Defined a downstream effector function by showing the Artemis:DNA-PKcs complex is a versatile endonuclease cleaving single-to-double-strand transitions, explaining hairpin/overhang processing.","evidence":"In vitro endonuclease assays with purified Artemis:DNA-PKcs on defined substrates","pmids":["15936993"],"confidence":"High","gaps":["Did not define how phosphorylation gates the activity in cells","Structural arrangement of Artemis on DNA-PKcs not resolved"]},{"year":2007,"claim":"Resolved how autophosphorylation regulates NHEJ by showing it controls stability of DNA-PKcs on DNA ends rather than initial recruitment.","evidence":"Laser microirradiation and FRAP with kinase-dead and phospho-mutant GFP-DNA-PKcs in live cells","pmids":["17438073"],"confidence":"High","gaps":["Did not identify the conformational basis of destabilization","Did not connect retention to specific pathway outcomes"]},{"year":2010,"claim":"Mapped DNA-PKcs into pathway-choice control by showing ABCDE autophosphorylation regulates repair-factor access and that DNA-PKcs and ATM coordinately govern NHEJ versus HR.","evidence":"I-SceI HR assays, epistasis with kinase-dead/null mutants, RAD51 foci, plus in vitro Artemis ssDNA endonuclease stimulation","pmids":["19535303","20117966"],"confidence":"High","gaps":["Direct ATM phosphorylation of DNA-PKcs not yet defined","Site-level basis of pathway switching incomplete"]},{"year":2014,"claim":"Extended DNA-PK signaling beyond the nucleus by showing damage-induced phosphorylation of GOLPH3 drives MYO18A-dependent Golgi dispersal affecting survival.","evidence":"siRNA knockdown, phospho-specific antibodies, reciprocal co-IP, Golgi imaging, and survival assays","pmids":["24485452"],"confidence":"High","gaps":["Phosphorylation sites on GOLPH3 not mapped here","Connection to nuclear DSB sensing unclear"]},{"year":2016,"claim":"Defined the molecular basis of DNA-PKcs/ATM crosstalk by showing DNA-PKcs directly phosphorylates and inhibits ATM, establishing reciprocal kinase regulation.","evidence":"In vitro kinase assays with purified proteins, ATM site mutagenesis, and DNA-PKcs KO/inhibition signaling assays","pmids":["27939942"],"confidence":"High","gaps":["In vivo stoichiometry of ATM inhibition unresolved","Temporal regulation across DDR phases unclear"]},{"year":2017,"claim":"Provided the first cryo-EM structures of DNA-PKcs and the holoenzyme, revealing the α-solenoid architecture, the DNA-binding tunnel protecting end DNA, and allosteric stimulation by Ku/DNA.","evidence":"Cryo-EM (4.4–6.6 Å) of DNA-PKcs and DNA-PK holoenzyme with activity assays","pmids":["28840859","28652322"],"confidence":"High","gaps":["Activated-state conformational changes not yet captured","Autophosphorylation-driven rearrangements not visualized"]},{"year":2019,"claim":"Established DNA-PKcs kinase activity as initiator of the DDR by showing it phosphorylates H2AX and KAP1 and drives chromatin decondensation and DDR factor recruitment at breaks.","evidence":"Kinase-domain-inactivated human cells, irradiation, phospho-readouts, and recruitment kinetics; PARylation by PARP1 regulating chromatin retention","pmids":["31396623","31485633"],"confidence":"High","gaps":["Quantitative hierarchy with ATM-driven DDR signaling unresolved","PARylation site mapping incomplete (Medium-confidence)"]},{"year":2020,"claim":"Uncovered non-canonical RNA-, immune-, and metabolism-linked functions: KU-directed assembly on U3 snRNA for 18S rRNA processing, DNA-PK as a STING-independent DNA sensor, and cGAS phosphorylation suppressing innate immunity.","evidence":"Mouse knockin models, U3 snRNA activation of purified DNA-PK, SSU processome co-purification, phosphoproteomics (HSPA8), in vitro cGAS kinase assay, patient-derived cells","pmids":["32103174","31980485","33273464"],"confidence":"High","gaps":["Mechanistic separation of nuclear repair versus nucleolar/immune roles incomplete","How RNA versus DNA selects functional output unclear"]},{"year":2021,"claim":"Revealed the autophosphorylation-driven functional switch structurally: open versus hairpin ends differentially trigger ABCDE phosphorylation, widening the DNA groove for Artemis recruitment and hairpin cleavage, with Artemis locking the kinase inactive.","evidence":"Cryo-EM of DNA-PK pre/post-autophosphorylation and with Artemis/hairpin, plus kinase and nuclease assays; additional substrate-specific activation by hairpin ends","pmids":["34936881","32716029"],"confidence":"High","gaps":["Order of events in the full synaptic complex not fully defined","Substrate selection rules beyond Artemis incomplete"]},{"year":2021,"claim":"Demonstrated a substrate-stabilizing role in cancer stem cells: DNA-PKcs phosphorylates SOX2 at S251 to block WWP2-mediated degradation, maintaining glioma stem cells.","evidence":"Mass spectrometry, co-IP, S251 mutagenesis, ubiquitination assays, and xenograft DNA-PKcs inhibition","pmids":["34193614"],"confidence":"High","gaps":["Whether nuclear DNA-PK pool serves this function during damage unclear","Generality across other transcription factors untested"]},{"year":2022,"claim":"Defined mitochondrial and conformational dimensions: DNA-PKcs phosphorylates Fis1-T34 to drive fragmentation, structural studies mapped the Artemis/XRCC4 mutually exclusive interface, and dimeric synaptic conformations captured NHEJ transition states.","evidence":"In vitro kinase assays and Fis1-T34A knockin mice; cryo-EM of basal Artemis:DNA-PKcs and dimeric complexes; structures with ATPγS and inhibitors; V(D)J domain-mapping mutagenesis; fork-reversal EM","pmids":["35290083","35801871","37256947","34987222","35150269","36130596"],"confidence":"High","gaps":["How cytoplasmic versus nuclear pools are partitioned unresolved","Coupling of fork-reversal role to canonical kinase signaling unclear"]},{"year":2023,"claim":"Extended DNA-PKcs into mitochondrial energetics and telomere protection: it forms the ANT2/VDAC2 (DAV) complex sustaining ADP-ATP exchange, dissociated by ATM-mediated T2609 phosphorylation, and cooperates with TRF2 to repress MRN-initiated resection at blunt telomeres.","evidence":"Co-IP, mitochondrial fractionation, Seahorse/membrane-potential assays, ATM kinase assays; in vitro/in vivo resection assays with AlphaFold-Multimer prediction","pmids":["36727301","37653239"],"confidence":"High","gaps":["Structural basis of DAV complex assembly not resolved","How a nuclear kinase is partitioned to mitochondria mechanistically unclear"]},{"year":null,"claim":"How DNA-PKcs is spatially and functionally partitioned among its nuclear repair, nucleolar rRNA-processing, cytosolic sensing, Golgi, mitochondrial, and replication-fork roles—and what signals route the same kinase to each—remains unresolved.","evidence":"","pmids":[],"confidence":"High","gaps":["No unifying model of subcellular trafficking","Substrate selection rules across compartments undefined","Relative physiological importance of non-canonical roles unquantified"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[10,18,26,30,38,9]},{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[1,10,26,38]},{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[0,1,13]},{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[16]},{"term_id":"GO:0140657","term_label":"ATP-dependent activity","supporting_discovery_ids":[24]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[1,5]},{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[16]},{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[26,28]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[17,26]},{"term_id":"GO:0005794","term_label":"Golgi apparatus","supporting_discovery_ids":[9]}],"pathway":[{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[0,3,6,20]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[2,17,18,27]},{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[16]},{"term_id":"R-HSA-69306","term_label":"DNA Replication","supporting_discovery_ids":[25]},{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[28]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[27,36]}],"complexes":["DNA-PK holoenzyme (DNA-PKcs–Ku70/Ku80)","Artemis:DNA-PKcs complex","DAV complex (DNA-PKcs–ANT2–VDAC2)","small-subunit (SSU) processome"],"partners":["XRCC6 (KU70)","XRCC5 (KU80)","DCLRE1C (ARTEMIS)","ATM","GOLPH3","FIS1","CGAS","SOX2"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P78527","full_name":"DNA-dependent protein kinase catalytic subunit","aliases":["DNPK1","Ser-473 kinase","S473K","p460"],"length_aa":4128,"mass_kda":469.1,"function":"Serine/threonine-protein kinase that acts as a molecular sensor for DNA damage (PubMed:11955432, PubMed:12649176, PubMed:14734805, PubMed:33854234). Involved in DNA non-homologous end joining (NHEJ) required for double-strand break (DSB) repair and V(D)J recombination (PubMed:11955432, PubMed:12649176, PubMed:14734805, PubMed:33854234, PubMed:34352203). Must be bound to DNA to express its catalytic properties (PubMed:11955432). Promotes processing of hairpin DNA structures in V(D)J recombination by activation of the hairpin endonuclease artemis (DCLRE1C) (PubMed:11955432). Recruited by XRCC5 and XRCC6 to DNA ends and is required to (1) protect and align broken ends of DNA, thereby preventing their degradation, (2) and sequester the DSB for repair by NHEJ (PubMed:11955432, PubMed:12649176, PubMed:14734805, PubMed:15574326, PubMed:33854234). Acts as a scaffold protein to aid the localization of DNA repair proteins to the site of damage (PubMed:11955432, PubMed:12649176, PubMed:14734805, PubMed:15574326). The assembly of the DNA-PK complex at DNA ends is also required for the NHEJ ligation step (PubMed:11955432, PubMed:12649176, PubMed:14734805, PubMed:15574326). Found at the ends of chromosomes, suggesting a further role in the maintenance of telomeric stability and the prevention of chromosomal end fusion (By similarity). Also involved in modulation of transcription (PubMed:11955432, PubMed:12649176, PubMed:14734805, PubMed:15574326). As part of the DNA-PK complex, involved in the early steps of ribosome assembly by promoting the processing of precursor rRNA into mature 18S rRNA in the small-subunit processome (PubMed:32103174). Binding to U3 small nucleolar RNA, recruits PRKDC and XRCC5/Ku86 to the small-subunit processome (PubMed:32103174). Recognizes the substrate consensus sequence [ST]-Q (PubMed:11955432, PubMed:12649176, PubMed:14734805, PubMed:15574326). Phosphorylates 'Ser-139' of histone variant H2AX, thereby regulating DNA damage response mechanism (PubMed:14627815, PubMed:16046194). Phosphorylates ASF1A, DCLRE1C, c-Abl/ABL1, histone H1, HSPCA, c-jun/JUN, p53/TP53, PARP1, POU2F1, DHX9, FH, SRF, NHEJ1/XLF, XRCC1, XRCC4, XRCC5, XRCC6, WRN, MYC and RFA2 (PubMed:10026262, PubMed:10467406, PubMed:11889123, PubMed:12509254, PubMed:14599745, PubMed:14612514, PubMed:14704337, PubMed:15177042, PubMed:1597196, PubMed:16397295, PubMed:18644470, PubMed:2247066, PubMed:2507541, PubMed:26237645, PubMed:26666690, PubMed:28712728, PubMed:29478807, PubMed:30247612, PubMed:8407951, PubMed:8464713, PubMed:9139719, PubMed:9362500). Can phosphorylate C1D not only in the presence of linear DNA but also in the presence of supercoiled DNA (PubMed:9679063). Ability to phosphorylate p53/TP53 in the presence of supercoiled DNA is dependent on C1D (PubMed:9363941). Acts as a regulator of the phosphatidylinositol 3-kinase/protein kinase B signal transduction by mediating phosphorylation of 'Ser-473' of protein kinase B (PKB/AKT1, PKB/AKT2, PKB/AKT3), promoting their activation (PubMed:15262962). Contributes to the determination of the circadian period length by antagonizing phosphorylation of CRY1 'Ser-588' and increasing CRY1 protein stability, most likely through an indirect mechanism (By similarity). Plays a role in the regulation of DNA virus-mediated innate immune response by assembling into the HDP-RNP complex, a complex that serves as a platform for IRF3 phosphorylation and subsequent innate immune response activation through the cGAS-STING pathway (PubMed:28712728). Also regulates the cGAS-STING pathway by catalyzing phosphorylation of CGAS, thereby impairing CGAS oligomerization and activation (PubMed:33273464). Also regulates the cGAS-STING pathway by mediating phosphorylation of PARP1 (PubMed:35460603)","subcellular_location":"Nucleus; Nucleus, nucleolus; Cytoplasm, cytosol","url":"https://www.uniprot.org/uniprotkb/P78527/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/PRKDC","classification":"Common Essential","n_dependent_lines":68,"n_total_lines":74,"dependency_fraction":0.918918918918919},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000253729","cell_line_id":"CID001251","localizations":[{"compartment":"nucleoplasm","grade":3}],"interactors":[{"gene":"XRCC6","stoichiometry":10.0},{"gene":"XRCC5","stoichiometry":10.0},{"gene":"NUCKS1","stoichiometry":4.0},{"gene":"SUPT16H","stoichiometry":4.0},{"gene":"ANKRD28","stoichiometry":0.2},{"gene":"CSNK1A1","stoichiometry":0.2},{"gene":"FKBP5","stoichiometry":0.2},{"gene":"FKBP8","stoichiometry":0.2},{"gene":"H2AFZ","stoichiometry":0.2},{"gene":"HIST2H2BE","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID001251","total_profiled":1310},"omim":[{"mim_id":"621451","title":"SMALL NUCLEOLAR RNA HOST GENE 12; SNHG12","url":"https://www.omim.org/entry/621451"},{"mim_id":"620529","title":"RING FINGER PROTEIN 121; RNF121","url":"https://www.omim.org/entry/620529"},{"mim_id":"616980","title":"CELL CYCLE REGULATOR OF NONHOMOLOGOUS END JOINING; CYREN","url":"https://www.omim.org/entry/616980"},{"mim_id":"616954","title":"YOU-HOOVER-FONG SYNDROME; YHFS","url":"https://www.omim.org/entry/616954"},{"mim_id":"616315","title":"PAXX NONHOMOLOGOUS END JOINING FACTOR; PAXX","url":"https://www.omim.org/entry/616315"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Enhanced","locations":[{"location":"Nucleoplasm","reliability":"Enhanced"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/PRKDC"},"hgnc":{"alias_symbol":["DNPK1","p350","DNAPK","XRCC7","DNA-PKcs","DNAPKc","DNA-PKC","p460"],"prev_symbol":["HYRC","HYRC1"]},"alphafold":{"accession":"P78527","domains":[],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P78527","model_url":"","pae_url":"","plddt_mean":null},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=PRKDC","jax_strain_url":"https://www.jax.org/strain/search?query=PRKDC"},"sequence":{"accession":"P78527","fasta_url":"https://rest.uniprot.org/uniprotkb/P78527.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P78527/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P78527"}},"corpus_meta":[{"pmid":"7855601","id":"PMC_7855601","title":"DNA-dependent kinase (p350) as a candidate gene for the murine SCID defect.","date":"1995","source":"Science (New York, 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Reconstitution experiments showed the complex forms only in the presence of double-stranded DNA.\",\n      \"method\": \"Immunoprecipitation with anti-Ku antibodies, reconstitution with purified components, catalytic activity assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution with purified components plus functional activity assay; foundational biochemical demonstration\",\n      \"pmids\": [\"8041718\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1992,\n      \"finding\": \"DNA-PKcs is a nuclear serine/threonine protein kinase activated by direct interaction with DNA (not using DNA as template/substrate) and phosphorylates DNA-binding proteins including transcription factors, suggesting a role in modulating transcriptional activity.\",\n      \"method\": \"Biochemical kinase assays, nuclear fractionation\",\n      \"journal\": \"Critical reviews in eukaryotic gene expression\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — early biochemical characterization with kinase assays; single lab review summarizing foundational work\",\n      \"pmids\": [\"1486241\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1995,\n      \"finding\": \"DNA-PKcs (p350) is the gene responsible for the murine SCID defect: p350 and the SCID-complementing gene co-localize to human chromosome 8q11, chromosomal fragments expressing p350 complement the SCID phenotype, and p350 protein is greatly reduced in SCID mouse cells.\",\n      \"method\": \"Chromosomal complementation, co-localization mapping, western blot of SCID-derived cells\",\n      \"journal\": \"Science (New York, N.Y.)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic complementation plus protein-level analysis; independently confirmed in companion paper (PMID:7855602)\",\n      \"pmids\": [\"7855601\", \"7855602\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1995,\n      \"finding\": \"Absence of the p350 subunit of DNA-PK in the radiosensitive human glioma cell line M059J is associated with defective DNA double-strand break repair, demonstrating that DNA-PK kinase activity is required for DSB repair.\",\n      \"method\": \"Immunoblot for p350 expression, DNA-PK kinase activity assay, DSB repair assay in human cell lines\",\n      \"journal\": \"Science (New York, N.Y.)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — natural loss-of-function human cell line with direct protein-level and activity measurements, replicated alongside PMID:7855601\",\n      \"pmids\": [\"7855602\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"The Artemis:DNA-PKcs complex cleaves a wide range of DNA substrates containing single-to-double-strand transitions (heterologous loops, stem-loops, flaps, gapped substrates) near the transition region; this versatile endonuclease activity is activated by DNA-PKcs binding and phosphorylation of Artemis.\",\n      \"method\": \"In vitro endonuclease assays with purified Artemis:DNA-PKcs complex on defined DNA substrates\",\n      \"journal\": \"DNA repair\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — reconstituted in vitro endonuclease assay with purified proteins on multiple substrate types; single lab\",\n      \"pmids\": [\"15936993\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"DNA-PKcs accumulates at DSB sites in a Ku80-dependent manner. Kinase activity and autophosphorylation status do not affect initial recruitment but do regulate the stability of DNA-PKcs binding to DNA ends; impaired autophosphorylation leads to prolonged retention at unrepaired DSBs, suggesting autophosphorylation facilitates NHEJ by destabilizing the interaction with DNA ends.\",\n      \"method\": \"Laser-induced DSB formation in live cells, FRAP (fluorescence recovery after photobleaching), live-cell imaging of GFP-DNA-PKcs\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — live-cell FRAP with kinase-dead and phospho-mutant constructs; multiple orthogonal approaches in single study\",\n      \"pmids\": [\"17438073\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"DNA-PKcs autophosphorylation at the ABCDE cluster regulates access of repair factors to DNA ends. Kinase-dead DNA-PKcs (DNA-PKcs-KR) cells show hyperrecombination 2-3 fold above DNA-PKcs null, and ATM-dependent phosphorylation of DNA-PKcs-KR contributes to this hyperrecombination phenotype; DNA-PKcs and ATM coordinately regulate the NHEJ/HR pathway choice.\",\n      \"method\": \"Direct repeat HR assay with I-SceI nuclease, epistasis analysis with kinase-dead and null mutants, RAD51 foci analysis\",\n      \"journal\": \"DNA repair\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic epistasis with multiple defined mutant cell lines, two orthogonal phenotypic readouts (HR assay and RAD51 foci)\",\n      \"pmids\": [\"19535303\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"DNA-PKcs regulates Artemis single-stranded DNA endonuclease activity: purified Artemis has intrinsic ssDNA endonuclease activity that is stimulated by DNA-PKcs; the divalent cation and sequence dependence matches that of the Artemis:DNA-PKcs double-stranded endonuclease activity.\",\n      \"method\": \"In vitro endonuclease assay with highly purified Artemis, addition of DNA-PKcs, antibody modulation\",\n      \"journal\": \"DNA repair\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution assay with purified proteins; single lab but multiple orthogonal methods\",\n      \"pmids\": [\"20117966\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"DNA-PKcs selectively stimulates WRN helicase activity (but not WRN exonuclease) on telomere D-loop model substrates in vitro, and DNA-PKcs knockdown reduces telomeric G-tail length in vivo; this effect is reversed by WRN helicase overexpression, indicating cooperative roles of DNA-PKcs and WRN in maintaining telomeric G-tails.\",\n      \"method\": \"In vitro helicase/exonuclease assay with purified DNA-PKcs and WRN, siRNA knockdown, telomere G-tail length measurement\",\n      \"journal\": \"Aging\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro stimulation assay plus in vivo knockdown rescue; single lab\",\n      \"pmids\": [\"20519774\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"DNA damage triggers DNA-PK-dependent phosphorylation of GOLPH3, which increases GOLPH3 interaction with MYO18A, applying tensile force to the Golgi and resulting in Golgi dispersal throughout the cytoplasm. Depletion of DNA-PK, GOLPH3, or MYO18A reduces survival after DNA damage.\",\n      \"method\": \"siRNA knockdown, phospho-specific antibodies, co-immunoprecipitation, fluorescence microscopy of Golgi morphology, cell survival assays\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal co-IP, functional knockdown with specific phenotypic readout, multiple orthogonal methods in single rigorous study\",\n      \"pmids\": [\"24485452\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"DNA-PKcs phosphorylates ATM at specific sites, inhibiting ATM activity. Chemical inhibition or genetic deletion of DNA-PKcs leads to hyperactive ATM; pre-incubation of ATM with active DNA-PKcs reduces ATM activity in vitro. Phospho-mimetic mutations at DNA-PKcs target sites in ATM inhibit ATM activity and impair ATM signaling after DNA damage.\",\n      \"method\": \"In vitro kinase assay with purified proteins, mutagenesis of ATM phosphorylation sites, DNA damage signaling assays in human cells with DNA-PKcs KO/inhibition\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro kinase assay plus mutagenesis plus genetic epistasis; multiple orthogonal methods\",\n      \"pmids\": [\"27939942\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"DNA-PKcs functionally interacts with telomerase in telomere length maintenance: Terc(-/-)/DNA-PKcs(-/-) double-knockout mice show accelerated telomere shortening compared to Terc(-/-) alone. DNA-PKcs is also required for end-to-end chromosome fusions and apoptosis triggered by critically short telomeres.\",\n      \"method\": \"Double-knockout mouse model, telomere length analysis (FISH/TRF), cytogenetic analysis\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic epistasis in mouse model with quantitative telomere length measurements and chromosome fusion analysis\",\n      \"pmids\": [\"12426399\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Phosphorylation of DNA-PKcs at the Thr-2609 cluster is a critical event for proper telomere end-processing; DNA-PKcs deficiency leads to uncapped telomeres that are inappropriately processed as DSBs, participating in spontaneous and radiation-induced telomere-DSB fusions requiring ligase IV.\",\n      \"method\": \"Cytogenetic analysis in DNA-PKcs-deficient mouse and human cells, phospho-specific immunoblot, irradiation experiments\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — defined phosphorylation cluster linked to telomere capping phenotype; single lab with multiple methods\",\n      \"pmids\": [\"19244120\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Cryo-EM structure of the human DNA-PK holoenzyme shows DNA-PKcs, KU70, KU80, and DNA forming a 650-kDa heterotetramer; DNA-PKcs N-terminal α-solenoid adopts a double-ring fold; together with KU70/80, DNA-PKcs forms a DNA-binding tunnel that protects ~30-bp DNA end. KU70/80 and DNA coordinately induce conformational changes in DNA-PKcs and allosterically stimulate its kinase activity.\",\n      \"method\": \"Cryo-electron microscopy (6.6 Å resolution), biochemical activity assays\",\n      \"journal\": \"Cell research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure with functional validation; independently confirmed by companion structure paper (PMID:28652322)\",\n      \"pmids\": [\"28840859\", \"28652322\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Cryo-EM structures of human DNA-PKcs (4.4 Å) and DNA-PK holoenzyme (5.8 Å) reveal that DNA-PKcs has three segments: N-terminal region (arm and bridge), circular cradle, and head with kinase domain; the C-terminal globular domain of Ku80 interacts with the arm of DNA-PKcs.\",\n      \"method\": \"Cryo-electron microscopy structural analysis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — high-resolution cryo-EM structure, replicated by companion paper (PMID:28840859)\",\n      \"pmids\": [\"28652322\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Cryo-EM structures of DNA-PKcs bound to DNA end and in complex with Ku70/80-DNA in inactive and activated states (3.7 Å overall; 3.2 Å FATKIN) reveal that kinase activation involves stretching and twisting within solenoid segments, loosening DNA-end binding. This structural plasticity of HEAT repeats represents a regulatory mechanism for PIKK family kinases.\",\n      \"method\": \"Cryo-electron microscopy, structural comparison of inactive vs. activated states\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — high-resolution cryo-EM with multiple conformational states; single rigorous study\",\n      \"pmids\": [\"33385326\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"DNA-PKcs has a KU-dependent role in rRNA processing and haematopoiesis: KU drives assembly of DNA-PKcs on cellular RNAs including U3 snRNA; U3 snRNA activates purified DNA-PK and triggers DNA-PKcs phosphorylation at T2609. DNA-PK resides in nucleoli in an rRNA-dependent manner and co-purifies with the small subunit processome. Blocking T2609 cluster phosphorylation (but not S2056 cluster) causes defects in 18S rRNA processing and bone marrow failure.\",\n      \"method\": \"Mouse knockin models (kinase-dead, T2609A), purified protein activation assays with U3 snRNA, nucleolar fractionation, co-purification with SSU processome, ribosome profiling\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple mouse genetic models plus in vitro reconstitution with RNA; multiple orthogonal methods in single rigorous study\",\n      \"pmids\": [\"32103174\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Human DNA-PK functions as a sensor in a STING-independent DNA sensing pathway (SIDSP): DNA-PK drives a broad antiviral response; HSPA8/HSC70 is identified as a target of inducible phosphorylation downstream of DNA-PK in this pathway. Viral proteins E1A (adenovirus) and ICP0 (HSV-1) block this response.\",\n      \"method\": \"Genetic deletion and inhibition of DNA-PK, phosphoproteomics identifying HSPA8/HSC70 as substrate, viral infection assays, antiviral response measurement\",\n      \"journal\": \"Science immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic sensor identification, specific substrate phosphorylation, viral antagonist characterization; multiple orthogonal methods\",\n      \"pmids\": [\"31980485\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"DNA-PK phosphorylates cGAS and suppresses its enzymatic activity. DNA-PK deficiency reduces cGAS phosphorylation and promotes antiviral innate immune responses. Cells from DNA-PKcs-deficient mice or patients with PRKDC missense mutations exhibit an inflammatory gene expression signature.\",\n      \"method\": \"In vitro kinase assay with DNA-PKcs and cGAS, co-immunoprecipitation, genetic mouse models, patient-derived cells\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro kinase assay plus genetic models plus patient cells; multiple orthogonal methods\",\n      \"pmids\": [\"33273464\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Cryo-EM structures of DNA-PK bound to DNA ends before and after autophosphorylation, and in complex with Artemis and a DNA hairpin, reveal a functional switch: open DNA ends inhibit cis-autophosphorylation of the ABCDE cluster but activate phosphorylation of other targets; hairpin ends promote ABCDE cis-autophosphorylation. Phosphorylation of four Thr residues in ABCDE causes gross structural rearrangement widening the DNA-binding groove for Artemis recruitment and hairpin cleavage. Artemis locks DNA-PK in a kinase-inactive state.\",\n      \"method\": \"Cryo-EM structural analysis of multiple states, in vitro kinase and nuclease assays\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structures of multiple functional states combined with biochemical validation; single rigorous study with multiple orthogonal methods\",\n      \"pmids\": [\"34936881\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"DNA-PKcs kinase activity is required for initiation of the DDR immediately after DSB induction: it drives phosphorylation of chromatin factors H2AX and KAP1, promotes local chromatin decondensation near DSB sites, and facilitates recruitment of DDR machinery. Loss of DNA-PKcs kinase activity markedly decreases DDR factor recruitment to DSBs.\",\n      \"method\": \"Kinase-domain inactivating human cell line, ionizing radiation, γH2AX and KAP1 phosphorylation assays, chromatin decondensation measurement, recruitment kinetics of DDR factors\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean kinase-dead cell line with multiple downstream readouts; multiple orthogonal methods\",\n      \"pmids\": [\"31396623\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"DNA-PKcs is neddylated at its kinase domain by the E2-conjugating enzyme UBE2M and E3 ligase HUWE1; inhibition of HUWE1-dependent neddylation impairs DNA-PKcs autophosphorylation at Ser2056 and reduces NHEJ efficiency.\",\n      \"method\": \"Immunoprecipitation, co-immunoprecipitation, HUWE1/UBE2M knockdown, NHEJ reporter assay, phospho-Ser2056 immunoblot\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP plus functional NHEJ assay; single lab with multiple methods but no structural validation of neddylation site\",\n      \"pmids\": [\"32457294\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Cryo-EM structural analysis of the basal (pre-activated) Artemis:DNA-PKcs complex shows the Artemis catalytic domain is positioned externally to DNA-PKcs prior to ABCDE autophosphorylation; both Artemis catalytic and regulatory domains interact with the N-HEAT and FAT domains of DNA-PKcs. A mutually exclusive binding site for Artemis and XRCC4 on DNA-PKcs was defined, and an XRCC4 peptide disrupts the Artemis:DNA-PKcs complex.\",\n      \"method\": \"Cryo-EM structural analysis, agarose-acrylamide gel complex stabilization, peptide competition assay\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — cryo-EM structure plus biochemical interaction mapping; single lab\",\n      \"pmids\": [\"35801871\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Cryo-EM structures of DNA-PKcs in three distinct dimeric conformations represent transition states during NHEJ: upon autophosphorylation, the long-range synaptic complex undergoes conformational change with both Ku and DNA-PKcs rotating outward to promote DNA break exposure and DNA-PKcs dissociation.\",\n      \"method\": \"Single-particle cryo-electron microscopy of NHEJ complexes at different stages\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structures of multiple conformational states; consistent with companion structural studies\",\n      \"pmids\": [\"37256947\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Cryo-EM structures of human DNA-PKcs with ATPγS and four inhibitors (wortmannin, NU7441, AZD7648, M3814) reveal the ATP binding mode and show that inhibitor binding causes movement of the PIKK regulatory domain (PRD), revealing a connection between the p-loop and PRD conformations. Inhibitors function through direct ATP competition and do not negatively allosterically affect holoenzyme assembly.\",\n      \"method\": \"Cryo-EM structural analysis, electrophoretic mobility shift assay (EMSA) for holoenzyme assembly\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — high-resolution cryo-EM with four inhibitors plus biochemical EMSA validation; single rigorous study\",\n      \"pmids\": [\"34987222\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"DNA-PKcs promotes fork reversal at stressed replication forks in a manner independent of its NHEJ role; cells lacking DNA-PKcs activity show increased DNA damage during S-phase and sensitivity to replication stress. Prevention of fork reversal by DNA-PKcs inhibition restores chemotherapy sensitivity in BRCA2-deficient tumors with acquired PARP inhibitor resistance.\",\n      \"method\": \"Electron microscopy of replication intermediates, DNA fiber assay, DNA-PKcs inhibitor/knockout, BRCA2-deficient tumor model\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct visualization of replication forks by EM combined with genetic/pharmacological perturbation; multiple orthogonal methods\",\n      \"pmids\": [\"36130596\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"DNA-PKcs interacts with and phosphorylates Fis1 at Thr34 in its TQ motif, increasing Fis1 affinity for Drp1 and inducing mitochondrial fragmentation; knockin mice with non-phosphorylatable T34A Fis1 show improved renal function and reduced mitochondrial fragmentation in acute kidney injury. Cytoplasmic localization of DNA-PKcs was detected in injured kidney tissues.\",\n      \"method\": \"Co-immunoprecipitation, in vitro kinase assay, Fis1-T34A knockin mice, mitochondrial morphology analysis, human patient urinary sediment analysis\",\n      \"journal\": \"Science signaling\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro kinase assay with identified phosphorylation site, knockin mouse validation, and human patient correlation; multiple orthogonal methods\",\n      \"pmids\": [\"35290083\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Physical ARTEMIS:DNA-PKcs interaction is necessary for V(D)J recombination: the L3062R pathogenic mutation in DNA-PKcs impairs physical interaction with Artemis; specific mutations in Artemis (in two conserved regions) that disrupt interaction with DNA-PKcs impair V(D)J recombination. Minimal interaction fragments were mapped: 42 aa from FAT region 2 of DNA-PKcs (PKcs3041-3082) and 26 aa from Artemis (ARM378-403).\",\n      \"method\": \"Mutagenesis, co-immunoprecipitation, V(D)J recombination assay, domain mapping with minimal fragments\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — mutagenesis with functional V(D)J assay and precise domain mapping; multiple orthogonal methods\",\n      \"pmids\": [\"35150269\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"DNA-PKcs directly interacts with mitochondrial proteins ANT2 and VDAC2 forming the DAV complex, which supports ADP-ATP exchange across mitochondrial membranes to sustain oxidative phosphorylation and membrane potential. The DAV complex dissociates in response to oxidative stress, attenuating ADP-ATP exchange; dissociation is mediated by ATM-dependent phosphorylation of DNA-PKcs at the Thr2609 cluster.\",\n      \"method\": \"Co-immunoprecipitation, mitochondrial fractionation, membrane potential assays, Seahorse metabolic analysis, DNA-PKcs-deficient cell lines, ATM kinase assay\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — co-IP identifying novel complex, functional metabolic readouts, ATM-mediated phosphorylation mechanism; multiple orthogonal methods in single study\",\n      \"pmids\": [\"36727301\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"DNA-PK and TRF2 cooperate to repress MRN-initiated resection at leading-end (blunt) telomeres: DNA-PK represses MRN-dependent long-range resection, while the iDDR of TRF2 inhibits MRN-CtIP endonuclease activity that would otherwise cleave DNA-PK off blunt telomere ends. AlphaFold-Multimer predicts conserved iDDR association with Rad50, potentially interfering with CtIP binding.\",\n      \"method\": \"In vitro resection assays, in vivo telomere resection analysis, AlphaFold-Multimer structural prediction with experimental validation\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vitro and in vivo functional assays demonstrating repression mechanism; multiple orthogonal approaches\",\n      \"pmids\": [\"37653239\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"DNA-PKcs (DNA-PK catalytic subunit) phosphorylates SOX2 at S251, stabilizing SOX2 by preventing WWP2-mediated ubiquitination and promoting glioma stem cell maintenance. Upon DNA damage, the DNA-PK complex dissociates from SOX2, allowing WWP2 interaction and SOX2 degradation, triggering differentiation.\",\n      \"method\": \"Mass spectrometry of SOX2-binding proteins, co-immunoprecipitation, site-directed mutagenesis of S251, ubiquitination assays, in vitro and in vivo (xenograft) DNA-PKcs inhibition\",\n      \"journal\": \"Science translational medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — MS identification of interaction, mutagenesis of phosphorylation site, ubiquitination assay, in vivo xenograft; multiple orthogonal methods\",\n      \"pmids\": [\"34193614\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"DNA-PKcs kinase activity and autophosphorylation regulate kinase complex conformation and dissociation during NHEJ; expression of catalytically inactive DNA-PKcs causes more genomic instability than loss of the protein itself (structural function), because kinase-dead DNA-PKcs persists at DNA lesions and alters repair pathway choice.\",\n      \"method\": \"Mouse models expressing kinase-dead DNA-PKcs, genomic instability assays, comparison to protein-null models\",\n      \"journal\": \"Cell & bioscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple mouse genetic models compared; review synthesizing results from multiple labs but mechanistic model from primary mouse model data\",\n      \"pmids\": [\"32015826\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"DNA-PARylation of DNA-PKcs by PARP1 regulates DNA-PK activity: DNA-PKcs is PARylated after DNA damage, PARP inhibition (olaparib) prevents DNA-PKcs detachment from chromatin and maintains DNA-PKcs Ser2056 autophosphorylation; olaparib and DNA-PK inhibition synergize to suppress cell survival.\",\n      \"method\": \"Immunoprecipitation, immunofluorescence, chromatin fractionation, phospho-Ser2056 immunoblot, cell survival assays\",\n      \"journal\": \"Molecular medicine reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP and functional chromatin dissociation assay; single lab, multiple methods\",\n      \"pmids\": [\"31485633\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Activation of DNA-PK by hairpinned DNA ends is substrate-specific: hairpinned DNA ends do not activate DNA-PK toward p53, XRCC4, XLF, or HSP90, but robustly stimulate ABCDE cluster autophosphorylation, which is required for Artemis activation. This reveals a multi-step mechanism of kinase activation.\",\n      \"method\": \"In vitro kinase assays with defined DNA substrates (hairpinned vs open ends), comparison across multiple substrates\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro kinase assay with multiple substrates revealing substrate-specific activation; single lab\",\n      \"pmids\": [\"32716029\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"The CRL4A-DTL ubiquitin ligase complex targets DNA-PKcs for nuclear proteasomal degradation; overexpression of CUL4A or DTL reduces NHEJ repair efficiency and increases DSB accumulation, leading to genomic instability and malignant transformation.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, NHEJ reporter assay, γH2AX measurement, overexpression in normal cells\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP with functional NHEJ assay and cell transformation; single lab, multiple methods\",\n      \"pmids\": [\"33627782\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"PRKDC recruits GDE2 to enhance stability of GNAS protein, which activates AKT phosphorylation, conferring doxorubicin resistance in osteosarcoma. The PRKDC-GDE2-GNAS-AKT regulatory axis was identified by a kinome-wide CRISPR screen.\",\n      \"method\": \"CRISPR kinome screen, co-immunoprecipitation of PRKDC and GDE2, GNAS stability assays, AKT phosphorylation analysis, xenograft and organoid models\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR screen plus co-IP and in vivo validation; single lab, multiple methods\",\n      \"pmids\": [\"38900943\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"DNA-PKcs has a kinase-dependent function in suppressing microhomology-mediated end joining (MMEJ) during class switch recombination (CSR) and a structural (kinase-independent) role in orientation of CSR; kinase-dead DNA-PKcs severely compromises CSR to IgG1 while DNA-PKcs deletion does not, revealing distinct structural and catalytic roles.\",\n      \"method\": \"Mouse B cell models with kinase-dead vs. null DNA-PKcs, high-throughput sequencing of CSR junctions, translocation analysis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — comparative genetic mouse models with high-throughput junction sequencing; multiple orthogonal methods\",\n      \"pmids\": [\"30072430\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"BCR-ABL down-regulates DNA-PKcs via proteasome-dependent degradation that requires BCR-ABL tyrosine kinase activity, resulting in marked DNA repair deficiency and increased sensitivity to ionizing radiation.\",\n      \"method\": \"Stable and inducible BCR-ABL expression in hematopoietic cells, proteasome inhibitor experiments, western blot, irradiation sensitivity assays\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pharmacological dissection of proteasome dependence and tyrosine kinase requirement; single lab, multiple methods\",\n      \"pmids\": [\"11264175\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Upon ionizing radiation, nuclear EGFR associates with DNA-PK, which phosphorylates PNPase (polynucleotide phosphorylase) at Ser-776; phospho-mimetic S776D PNPase has impaired ribonuclease activity, while the non-phosphorylatable S776A mutant retains ribonuclease activity and degrades c-MYC mRNA, affecting radioresistance.\",\n      \"method\": \"Co-immunoprecipitation, site-directed mutagenesis of PNPase, in vitro ribonuclease assays, knockdown experiments\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro kinase assay with mutagenesis of substrate residue, functional ribonuclease assay; single lab, multiple orthogonal methods\",\n      \"pmids\": [\"22815474\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"DNA-PKcs promotes cardiac ischemia-reperfusion injury through direct interaction with BI-1 (Bax inhibitor-1), promoting BI-1 degradation without affecting BI-1 transcription. Loss of DNA-PKcs stabilizes BI-1, protecting mitochondria; concurrent BI-1 knockout abrogates the cardioprotection of DNA-PKcs deletion.\",\n      \"method\": \"Cardiomyocyte-specific DNA-PKcs knockout mice, co-immunoprecipitation, double-knockout epistasis, mitochondrial function assays\",\n      \"journal\": \"Basic research in cardiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP plus genetic epistasis in mouse model; single lab\",\n      \"pmids\": [\"31919590\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"DNA-PKcs (PRKDC) is a large nuclear serine/threonine PIKK-family kinase that, upon recruitment to DNA double-strand breaks by the Ku70/80 heterodimer, forms the active DNA-PK holoenzyme; it autophosphorylates at distinct clusters (ABCDE at T2609, S2056) to undergo sequential conformational changes that regulate DNA-end protection versus processing, recruits and activates the Artemis nuclease for hairpin opening in V(D)J recombination, negatively regulates ATM activity through direct phosphorylation, and has additional non-canonical roles including phosphorylating cGAS to suppress innate immunity, phosphorylating GOLPH3 to trigger Golgi dispersal after DNA damage, phosphorylating Fis1 to control mitochondrial fragmentation, acting in an RNA-dependent capacity in ribosome biogenesis (18S rRNA processing) via KU-directed assembly on U3 snRNA, stabilizing SOX2 in glioma stem cells, promoting replication fork reversal during replication stress, and regulating oxidative phosphorylation through a mitochondrial ANT2/VDAC2 complex whose dissociation is controlled by ATM-mediated phosphorylation of the T2609 cluster.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"PRKDC encodes DNA-PKcs, a nuclear PIKK-family serine/threonine kinase that serves as the catalytic engine of non-homologous end joining (NHEJ), the principal pathway for repairing DNA double-strand breaks (DSBs) [#0, #3]. Recruited to free DNA ends by the Ku70/Ku80 heterodimer, DNA-PKcs assembles the catalytically active DNA-PK holoenzyme only in the presence of double-stranded DNA, with Ku and DNA coordinately inducing conformational changes that allosterically stimulate the kinase and protect ~30 bp of the DNA end within a binding tunnel [#0, #13, #15]. Activation is governed by autophosphorylation: hairpinned versus open DNA ends differentially trigger phosphorylation of the ABCDE (T2609) cluster, driving a structural rearrangement that widens the DNA-binding groove, destabilizes end binding to promote NHEJ progression, and licenses recruitment of the Artemis nuclease, which DNA-PKcs activates to open hairpins and cleave single-to-double-strand transitions during V(D)J recombination [#5, #19, #21, #33, #4, #27]. DNA-PKcs initiates the broader DNA damage response by phosphorylating chromatin factors H2AX and KAP1 and promoting chromatin decondensation at breaks, and it shapes repair-pathway choice by negatively regulating ATM through direct phosphorylation and by suppressing hyper-recombination [#20, #10, #6]; catalytically inactive DNA-PKcs persists at lesions and causes greater genomic instability than protein loss, revealing distinct structural and catalytic functions [#31, #36]. Loss of DNA-PKcs underlies the murine SCID immunodeficiency phenotype [#2]. Beyond canonical repair, DNA-PKcs has KU-directed roles in 18S rRNA processing within the small-subunit processome via U3 snRNA-driven assembly and T2609 phosphorylation [#16], acts as a cytosolic DNA sensor driving STING-independent antiviral responses while phosphorylating and suppressing cGAS [#17, #18], and executes signaling functions at the Golgi (GOLPH3-MYO18A-mediated dispersal after damage) [#9], mitochondria (Fis1-driven fragmentation and an ANT2/VDAC2 oxidative-phosphorylation complex) [#26, #28], telomeres [#11, #12, #29], and replication forks (promoting fork reversal independent of NHEJ) [#25].\",\n  \"teleology\": [\n    {\n      \"year\": 1994,\n      \"claim\": \"Established that DNA-PK is an assembled holoenzyme requiring Ku and double-stranded DNA, defining the order of complex formation that underlies break recognition.\",\n      \"evidence\": \"Anti-Ku immunoprecipitation and reconstitution with purified components plus catalytic activity assay\",\n      \"pmids\": [\"8041718\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve the structural basis of activation\", \"Did not identify physiological substrates beyond the complex itself\"]\n    },\n    {\n      \"year\": 1995,\n      \"claim\": \"Linked DNA-PKcs genetically to immunodeficiency and DSB repair, showing the gene is responsible for the murine SCID defect and required for repair in human cells.\",\n      \"evidence\": \"Chromosomal complementation, co-localization mapping, and immunoblot/repair assays in SCID and M059J cells\",\n      \"pmids\": [\"7855601\", \"7855602\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define the molecular step of NHEJ requiring DNA-PKcs\", \"Catalytic versus structural contribution unresolved\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Defined a downstream effector function by showing the Artemis:DNA-PKcs complex is a versatile endonuclease cleaving single-to-double-strand transitions, explaining hairpin/overhang processing.\",\n      \"evidence\": \"In vitro endonuclease assays with purified Artemis:DNA-PKcs on defined substrates\",\n      \"pmids\": [\"15936993\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define how phosphorylation gates the activity in cells\", \"Structural arrangement of Artemis on DNA-PKcs not resolved\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Resolved how autophosphorylation regulates NHEJ by showing it controls stability of DNA-PKcs on DNA ends rather than initial recruitment.\",\n      \"evidence\": \"Laser microirradiation and FRAP with kinase-dead and phospho-mutant GFP-DNA-PKcs in live cells\",\n      \"pmids\": [\"17438073\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not identify the conformational basis of destabilization\", \"Did not connect retention to specific pathway outcomes\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Mapped DNA-PKcs into pathway-choice control by showing ABCDE autophosphorylation regulates repair-factor access and that DNA-PKcs and ATM coordinately govern NHEJ versus HR.\",\n      \"evidence\": \"I-SceI HR assays, epistasis with kinase-dead/null mutants, RAD51 foci, plus in vitro Artemis ssDNA endonuclease stimulation\",\n      \"pmids\": [\"19535303\", \"20117966\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct ATM phosphorylation of DNA-PKcs not yet defined\", \"Site-level basis of pathway switching incomplete\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Extended DNA-PK signaling beyond the nucleus by showing damage-induced phosphorylation of GOLPH3 drives MYO18A-dependent Golgi dispersal affecting survival.\",\n      \"evidence\": \"siRNA knockdown, phospho-specific antibodies, reciprocal co-IP, Golgi imaging, and survival assays\",\n      \"pmids\": [\"24485452\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Phosphorylation sites on GOLPH3 not mapped here\", \"Connection to nuclear DSB sensing unclear\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Defined the molecular basis of DNA-PKcs/ATM crosstalk by showing DNA-PKcs directly phosphorylates and inhibits ATM, establishing reciprocal kinase regulation.\",\n      \"evidence\": \"In vitro kinase assays with purified proteins, ATM site mutagenesis, and DNA-PKcs KO/inhibition signaling assays\",\n      \"pmids\": [\"27939942\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo stoichiometry of ATM inhibition unresolved\", \"Temporal regulation across DDR phases unclear\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Provided the first cryo-EM structures of DNA-PKcs and the holoenzyme, revealing the α-solenoid architecture, the DNA-binding tunnel protecting end DNA, and allosteric stimulation by Ku/DNA.\",\n      \"evidence\": \"Cryo-EM (4.4–6.6 Å) of DNA-PKcs and DNA-PK holoenzyme with activity assays\",\n      \"pmids\": [\"28840859\", \"28652322\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Activated-state conformational changes not yet captured\", \"Autophosphorylation-driven rearrangements not visualized\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Established DNA-PKcs kinase activity as initiator of the DDR by showing it phosphorylates H2AX and KAP1 and drives chromatin decondensation and DDR factor recruitment at breaks.\",\n      \"evidence\": \"Kinase-domain-inactivated human cells, irradiation, phospho-readouts, and recruitment kinetics; PARylation by PARP1 regulating chromatin retention\",\n      \"pmids\": [\"31396623\", \"31485633\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Quantitative hierarchy with ATM-driven DDR signaling unresolved\", \"PARylation site mapping incomplete (Medium-confidence)\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Uncovered non-canonical RNA-, immune-, and metabolism-linked functions: KU-directed assembly on U3 snRNA for 18S rRNA processing, DNA-PK as a STING-independent DNA sensor, and cGAS phosphorylation suppressing innate immunity.\",\n      \"evidence\": \"Mouse knockin models, U3 snRNA activation of purified DNA-PK, SSU processome co-purification, phosphoproteomics (HSPA8), in vitro cGAS kinase assay, patient-derived cells\",\n      \"pmids\": [\"32103174\", \"31980485\", \"33273464\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanistic separation of nuclear repair versus nucleolar/immune roles incomplete\", \"How RNA versus DNA selects functional output unclear\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Revealed the autophosphorylation-driven functional switch structurally: open versus hairpin ends differentially trigger ABCDE phosphorylation, widening the DNA groove for Artemis recruitment and hairpin cleavage, with Artemis locking the kinase inactive.\",\n      \"evidence\": \"Cryo-EM of DNA-PK pre/post-autophosphorylation and with Artemis/hairpin, plus kinase and nuclease assays; additional substrate-specific activation by hairpin ends\",\n      \"pmids\": [\"34936881\", \"32716029\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Order of events in the full synaptic complex not fully defined\", \"Substrate selection rules beyond Artemis incomplete\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Demonstrated a substrate-stabilizing role in cancer stem cells: DNA-PKcs phosphorylates SOX2 at S251 to block WWP2-mediated degradation, maintaining glioma stem cells.\",\n      \"evidence\": \"Mass spectrometry, co-IP, S251 mutagenesis, ubiquitination assays, and xenograft DNA-PKcs inhibition\",\n      \"pmids\": [\"34193614\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether nuclear DNA-PK pool serves this function during damage unclear\", \"Generality across other transcription factors untested\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Defined mitochondrial and conformational dimensions: DNA-PKcs phosphorylates Fis1-T34 to drive fragmentation, structural studies mapped the Artemis/XRCC4 mutually exclusive interface, and dimeric synaptic conformations captured NHEJ transition states.\",\n      \"evidence\": \"In vitro kinase assays and Fis1-T34A knockin mice; cryo-EM of basal Artemis:DNA-PKcs and dimeric complexes; structures with ATPγS and inhibitors; V(D)J domain-mapping mutagenesis; fork-reversal EM\",\n      \"pmids\": [\"35290083\", \"35801871\", \"37256947\", \"34987222\", \"35150269\", \"36130596\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How cytoplasmic versus nuclear pools are partitioned unresolved\", \"Coupling of fork-reversal role to canonical kinase signaling unclear\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Extended DNA-PKcs into mitochondrial energetics and telomere protection: it forms the ANT2/VDAC2 (DAV) complex sustaining ADP-ATP exchange, dissociated by ATM-mediated T2609 phosphorylation, and cooperates with TRF2 to repress MRN-initiated resection at blunt telomeres.\",\n      \"evidence\": \"Co-IP, mitochondrial fractionation, Seahorse/membrane-potential assays, ATM kinase assays; in vitro/in vivo resection assays with AlphaFold-Multimer prediction\",\n      \"pmids\": [\"36727301\", \"37653239\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of DAV complex assembly not resolved\", \"How a nuclear kinase is partitioned to mitochondria mechanistically unclear\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How DNA-PKcs is spatially and functionally partitioned among its nuclear repair, nucleolar rRNA-processing, cytosolic sensing, Golgi, mitochondrial, and replication-fork roles—and what signals route the same kinase to each—remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No unifying model of subcellular trafficking\", \"Substrate selection rules across compartments undefined\", \"Relative physiological importance of non-canonical roles unquantified\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [10, 18, 26, 30, 38, 9]},\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [1, 10, 26, 38]},\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [0, 1, 13]},\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [16]},\n      {\"term_id\": \"GO:0140657\", \"supporting_discovery_ids\": [24]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [1, 5]},\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [16]},\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [26, 28]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [17, 26]},\n      {\"term_id\": \"GO:0005794\", \"supporting_discovery_ids\": [9]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [0, 3, 6, 20]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [2, 17, 18, 27]},\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [16]},\n      {\"term_id\": \"R-HSA-69306\", \"supporting_discovery_ids\": [25]},\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [28]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [27, 36]}\n    ],\n    \"complexes\": [\n      \"DNA-PK holoenzyme (DNA-PKcs–Ku70/Ku80)\",\n      \"Artemis:DNA-PKcs complex\",\n      \"DAV complex (DNA-PKcs–ANT2–VDAC2)\",\n      \"small-subunit (SSU) processome\"\n    ],\n    \"partners\": [\n      \"XRCC6 (Ku70)\",\n      \"XRCC5 (Ku80)\",\n      \"DCLRE1C (Artemis)\",\n      \"ATM\",\n      \"GOLPH3\",\n      \"FIS1\",\n      \"CGAS\",\n      \"SOX2\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}