{"gene":"TOP1","run_date":"2026-06-10T10:51:55","timeline":{"discoveries":[{"year":1985,"finding":"Yeast TOP1 encodes DNA topoisomerase I; deletion mutants (delta top1) are viable, but delta top1 top2-ts double mutants are inviable even when mitotic progression is blocked, demonstrating that Top1 serves an auxiliary role to Top2 in yeast DNA metabolism.","method":"Gene disruption/deletion in S. cerevisiae, complementation, temperature-shift experiments with synchronous cultures","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean loss-of-function genetics with defined viability phenotype, replicated with multiple alleles in foundational study","pmids":["2997777"],"is_preprint":false},{"year":2002,"finding":"In S. cerevisiae, Tdp1 (tyrosyl-DNA phosphodiesterase) and the Rad1-Rad10 structure-specific endonuclease act as two primary, redundant pathways to remove covalent Top1-DNA adducts (Top1 cleavage complexes) at replication forks; tdp1 rad1 double mutants are highly sensitive to camptothecin and show a TOP1-dependent growth delay. Both pathways funnel into RAD52/RAD51/RAD50-dependent recombination for replication restart.","method":"Genetic epistasis analysis (double/triple mutants), camptothecin sensitivity assays in S. cerevisiae","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — extensive epistasis with multiple alleles, rigorous genetic controls, replicated across multiple pathway genes","pmids":["12368472"],"is_preprint":false},{"year":2007,"finding":"Top1 and Top2 act within a ~600-bp region spanning moving replication forks in S. cerevisiae. TOP1 ablation alone does not block fork progression or activate the Rad53 checkpoint, but top1 top2 double mutants exhibit replication fork block, fork processing, and Rad53/γH2AX activation in S phase, demonstrating that Top1 and Top2 act coordinately to relieve torsional stress and prevent sister chromatid entanglement at forks.","method":"Chromatin immunoprecipitation (ChIP) of replicating chromosomes, genetic analysis of top1, top2, and top1 top2 mutants, checkpoint kinase (Rad53) phosphorylation assays, γH2AX detection","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (ChIP, genetics, checkpoint assays) in a single rigorous study","pmids":["17671091"],"is_preprint":false},{"year":2008,"finding":"Top1 binds to and is catalytically active at telomeric DNA repeats in S. cerevisiae; deletion of TOP1 or expression of catalytically inactive Top1-Y727F increases H4 histone acetylation at subtelomeric regions and activates transcription of telomere-proximal genes, indicating that Top1 catalytic activity promotes a repressed chromatin state at telomeres.","method":"Chromatin immunoprecipitation (ChIP) for Top1 binding and histone modifications, expression profiling (microarray), TOP1 deletion and Y727F catalytic-dead mutant","journal":"Journal of molecular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP and expression profiling with catalytic mutant, single lab, two orthogonal methods","pmids":["18272174"],"is_preprint":false},{"year":2011,"finding":"In fission yeast, STUbL (SUMO-targeted ubiquitin ligase), Rad60, and the Nse2 SUMO E3 ligase suppress spontaneous Top1-mediated DNA damage (Top1 cleavage complexes). Cells lacking STUbL activity become dependent on Tdp1, and Top1ccs detected by ChIP-qPCR are elevated when STUbL, Rad60, or Nse2 activity is absent. Tdp1 and Rad16-Swi10 (NER endonuclease) constitute the two major Top1cc repair pathways, with STUbL/Rad60/Nse2 facilitating the Rad16-Swi10 branch.","method":"Genetic epistasis, chromatin immunoprecipitation (ChIP)-qPCR to detect Top1ccs, camptothecin sensitivity assays in S. pombe","journal":"PLoS genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — ChIP-qPCR for direct Top1cc detection combined with extensive epistasis analysis, multiple orthogonal methods","pmids":["21408210"],"is_preprint":false},{"year":2014,"finding":"Top1 cleavage complexes (Top1ccs) are specifically stabilized at the ribosomal replication fork barrier (rRFB) in S. cerevisiae in a strand-specific, replication-independent manner requiring Fob1 and Tof2; these Top1ccs are reversible (not subject to Tdp1 or Mus81 repair) and their presence correlates with Top1-dependent rDNA stability.","method":"ChIP detection of Top1ccs at rRFB, ectopic rRFB constructs, genetic analysis with fob1 and tof2 mutants, Tdp1/Mus81 epistasis in S. cerevisiae","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct ChIP detection of Top1ccs, ectopic construct validation, epistasis analysis — multiple orthogonal methods","pmids":["24574527"],"is_preprint":false},{"year":2015,"finding":"Trapping of the Top1 cleavage complex (Top1cc) in S. cerevisiae (via camptothecin or top1-T722A allele) causes a 5–10-fold increase in reciprocal crossover events and greatly elevated instability within rDNA and CUP1 tandem arrays, with evidence that recombinogenic lesions arise during or after DNA synthesis.","method":"Sectoring assay for loss of heterozygosity, chromosome microarrays, sub-culturing for 250 generations, genetic analysis in S. cerevisiae","journal":"PLoS genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — quantitative genetic assay plus genome-wide microarray, single lab, two orthogonal methods","pmids":["25830313"],"is_preprint":false},{"year":2015,"finding":"ERCC1-XPF endonuclease and RPA participate in NER-mediated repair of Top1-attached nick DNA lesions (3'-phosphotyrosyl bond). ERCC1-XPF cleaves a Tyr-nick DNA substrate in vitro in the presence of RPA, and repair DNA synthesis of these lesions occurs with NER factors (ERCC1-XPF, RPA, DNA pol δ, FEN1, DNA ligase 1).","method":"In vitro nuclease assay on Tyr-nick DNA substrate, co-localization of ERCC1-XPF and RPA in CPT-treated cells, in vitro repair synthesis reconstitution","journal":"Carcinogenesis","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution of repair synthesis plus in vivo co-localization; multiple orthogonal methods","pmids":["26025908"],"is_preprint":false},{"year":2016,"finding":"PARP1 is activated by trapped Top1 cleavage complexes (Top1cc) and PARylates Top1. Using live-cell FRAP of GFP-tagged human Top1 and PARP inhibitors (ABT-888/Veliparib), PARP inhibition was shown to delocalize Top1 from nucleolus to nucleoplasm and to markedly increase the CPT-induced immobile/bound fraction of Top1 (Top1cc) genome-wide, indicating that PAR-modification of Top1 counteracts CPT-induced stabilization of Top1cc. Top1 residues Trp205 and Asn722 are critical for subnuclear dynamics; Top1-N722S is restricted to the nucleolus under CPT due to deficient PARylation.","method":"Live-cell imaging of GFP-tagged Top1, FRAP with kinetic modelling, PARP inhibitor treatment, site-directed mutagenesis (W205, N722S)","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — direct live-cell imaging + FRAP kinetics + mutagenesis, multiple orthogonal methods in single lab","pmids":["27466387"],"is_preprint":false},{"year":2016,"finding":"Top1-dependent ribonucleotide-associated deletions in yeast arise through a sequential double-cleavage mechanism: Top1 cleaves at a ribonucleotide, the resulting intermediate realigns within a tandem repeat, and Top1 ligates the realigned nick, generating a single-repeat deletion. Deletion frequency is inversely related to the distance between two Top1 cleavage sites, and complementarity downstream of the gap promotes deletions more effectively than upstream complementarity.","method":"Parallel in vitro Top1 cleavage/ligation assays and in vivo yeast mutagenesis assays, systematic variation of substrate distances and complementarity","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution with parallel in vivo validation; mechanistic model tested with multiple substrate variants","pmids":["27257064"],"is_preprint":false},{"year":2019,"finding":"Top1cc-dependent large deletions (~100 bp) in yeast are produced by the NHEJ pathway and require Top1cc removal from DNA ends; these events accumulate in quiescent cells, suggesting that causative DSBs arise outside replication context. Joining of ends from distinct Top1-associated DSBs via NHEJ deletes the intervening sequence.","method":"Genetic analysis of NHEJ mutants in S. cerevisiae, characterization of deletion junctions, quiescence experiments","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis with junction characterization, single lab, two orthogonal approaches","pmids":["31636207"],"is_preprint":false},{"year":2019,"finding":"Yeast Apn2 (AP endonuclease 2) resolves Top1-mediated phosphotyrosine-DNA conjugates (Top1cc), terminal 2',3'-cyclic phosphates, and their hydrolyzed products at 3' DNA ends, and suppresses 2-bp slippage mutagenesis in RNH201-deficient cells, defining Apn2 as an alternative Top1cc repair enzyme.","method":"Genetic epistasis, biochemical assays with purified Apn2 on Top1cc and cyclic-phosphate substrates in S. cerevisiae","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro biochemical assay with defined substrates plus in vivo genetics; multiple methods, single lab","pmids":["30778235"],"is_preprint":false},{"year":2022,"finding":"Defective ribonucleotide excision repair in mammals leads to accumulation of genome-embedded ribonucleotides, at which TOP1 activity generates short (2–5 bp) deletions (ID4/ID-TOP1 mutation signature) at TNT motifs; this transcription-associated mutagenesis process contributes to cancer somatic mutations and germline de novo indels.","method":"Genome sequencing of cancer and germline samples from ribonucleotide excision repair-deficient mammals, mutational signature analysis, functional TOP1 inhibition experiments","journal":"Nature","confidence":"High","confidence_rationale":"Tier 2 / Strong — mechanistic experiments in mammalian cells combined with human cancer genome analysis; replicated across multiple biological contexts","pmids":["35140396"],"is_preprint":false},{"year":2022,"finding":"Archaeal CsTOP1 (a structural ortholog of eukaryotic TOP1) adopts an open conformation in apo form, with the hinge loop (5-residue junction between CAP and CAT modules) permitting rotation that allows DNA entry. A conserved tyrosine near the hinge mediates transition to the closed conformation upon DNA binding; directed mutagenesis of hinge residues confirmed this flexibility is required for activity and linked to camptothecin sensitivity.","method":"Crystal structure of apo CsTOP1, directed mutagenesis of hinge residues, camptothecin sensitivity assays","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Moderate — crystal structure plus mutagenesis validation; multiple orthogonal methods in single study","pmids":["35013228"],"is_preprint":false},{"year":2022,"finding":"Whole-genome CRISPR screens and co-deficient cell lines demonstrated that MUS81 mediates the generation of excess double-strand breaks (DSBs) in TDP1-KO cells exposed to Top1 poisons; APEX1/2 are synthetically lethal with TDP1 but do not reduce DSB formation in TDP1-KO cells. Top1cc can be resolved directly by TDP1 or converted to DSBs and repaired by homologous recombination.","method":"Genome-wide CRISPR screens, generation of co-deficient cell lines (TDP1/MUS81 and TDP1/APEX KO), DNA damage assays","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR genetic screens plus isogenic KO cell validation, single lab","pmids":["35869071"],"is_preprint":false},{"year":2023,"finding":"TDP1 suppresses chromosomal translocations arising from transcription-associated TOP1-induced DSBs; error-prone repair of these breaks relies on the MRN complex and canonical NHEJ. TDP1 loss leads to increased chromosome rearrangements specifically linked to transcription-coupled TOP1 activity.","method":"Loss-of-function genetics in human cells, cytogenetic analysis of translocations, epistasis with MRN and NHEJ factors","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis with cytogenetic readout, single lab, two orthogonal methods","pmids":["37945566"],"is_preprint":false},{"year":2023,"finding":"MUS81 endonuclease can efficiently cleave DNA substrates bearing proteolytically processed (but not intact) TOP1, indicating TOP1 must be dislodged or partially degraded before MUS81 cleavage. MUS81 and TDP1 play independent, non-redundant roles in repair of CPT-induced TOP1-DPC lesions.","method":"In vitro MUS81 cleavage assays with fluorescein-, streptavidin-, and proteolytically processed TOP1-modified DNA substrates; nuclear extract assays; TDP1/MUS81 double-KO cell sensitivity assays","journal":"BMC biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with multiple substrate types plus cellular genetic validation; multiple orthogonal methods","pmids":["37194054"],"is_preprint":false},{"year":2024,"finding":"CDK1 phosphorylates TDP1 at Ser61 during mitosis; TDP1-S61A (non-phosphorylatable) is trapped on mitotic chromosomes and causes DNA damage and mitotic defects. Top1cc repair in mitosis occurs via a MUS81-dependent mechanism, and TDP1-S61A enrichment at common fragile sites over-stimulates MUS81-dependent chromatid breaks, leading to anaphase bridges, micronuclei, and 53BP1 nuclear bodies in G1.","method":"CDK1 kinase assay identifying TDP1 S61 phosphorylation, TDP1-S61A mutant cell lines, live imaging, γH2AX/53BP1 focus analysis, MUS81 epistasis","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — biochemical kinase assay identifying modification site plus mutant cell functional analysis; multiple orthogonal methods in single lab","pmids":["39014228"],"is_preprint":false},{"year":2024,"finding":"TOP1 and R-loops are enriched at highly expressed genes where transcriptional DSBs occur; depletion of R-loops or TOP1 significantly reduces transcription-associated DSBs at these loci, establishing that TOP1 and R-loops cooperate in transcriptional DSB formation and contribute to early molecular changes driving cancer development.","method":"Genome-wide DSB mapping, R-loop mapping, TOP1cc mapping, TOP1 depletion and R-loop depletion in human cells","journal":"iScience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genome-wide mapping plus loss-of-function experiments, single lab, multiple orthogonal mapping methods","pmids":["38375218"],"is_preprint":false},{"year":2025,"finding":"TOP1 mutations (S57C, R364H, W401C, G359E) arising under selective pressure from ADC therapy in metastatic breast cancer patients exhibit reduced enzymatic activity, attenuated covalent DNA binding, and resistance to TOP1-inhibitor payloads SN38 and deruxtecan, functionally characterizing resistance-associated TOP1 mutations.","method":"Functional characterization of recombinant mutant TOP1 proteins (enzymatic activity assays, covalent DNA binding assays, in vitro drug resistance assays), plasma-based genotyping","journal":"Clinical cancer research","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro biochemical characterization of mutant proteins with multiple assays, single lab","pmids":["39745368"],"is_preprint":false},{"year":2010,"finding":"TOP1 interacts with BTBD1 and BTBD2; this interaction requires human TOP1 residues 236 and 237. The same residues are required for TOP1 to enhance infectivity of HIV-1 virions when expressed in African Green Monkey producer cells. RNAi knockdown of BTBD2 in AGM and human cells increased HIV-1 permissiveness 2–3-fold.","method":"Co-IP/interaction assays with TOP1 point mutants (residues 236/237), RNAi knockdown of BTBD2, HIV-1 infectivity assays","journal":"Virology journal","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single Co-IP and RNAi experiment, single lab, limited mechanistic follow-up","pmids":["21092135"],"is_preprint":false},{"year":2026,"finding":"In vertebrate (chicken DT40) cells, TOP1 depletion alone does not affect replication fork progression, and TOP2 inhibition (ICRF193) alone also has no effect; however, combined TOP1 depletion plus TOP2 inhibition nearly abolishes DNA replication, causing S-phase arrest and apoptosis. TOP1 depletion combined with TOP2 poison (etoposide) impairs replication initiation rather than elongation. This demonstrates complementary roles of TOP1 and TOP2 in both initiation and progression of DNA replication in vertebrates.","method":"Conditional TOP1 depletion in DT40 cells, ICRF193 and etoposide pharmacological inhibition, DNA fiber assays for fork progression, cell cycle analysis","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional genetic depletion plus pharmacological inhibition with DNA fiber assays, single lab, multiple orthogonal methods","pmids":["41791707"],"is_preprint":false},{"year":2002,"finding":"Deletion of TOP1 in S. cerevisiae increases histone H4 acetylation at the rDNA locus and renders rDNA chromatin more accessible, similar to sir2 mutants, indicating that Top1 activity promotes a repressed chromatin state at rDNA.","method":"Chromatin accessibility assays, histone acetylation analysis at rDNA in top1 and sir2 deletion strains of S. cerevisiae","journal":"Journal of molecular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct chromatin modification assays in loss-of-function mutants, single lab","pmids":["12215413"],"is_preprint":false},{"year":1999,"finding":"The t(11;20)(p15;q11) chromosomal translocation produces an NUP98-TOP1 chimeric mRNA encoding the NUP98 FXFG repeats fused to the body of DNA topoisomerase I (TOP1), with the breakpoint in TOP1 occurring within the gene; this fusion is associated with therapy-related myelodysplastic syndrome/acute myeloid leukemia.","method":"Cloning and sequencing of t(11;20) translocation breakpoints, RT-PCR characterization of fusion transcript in patient samples","journal":"Blood","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct molecular cloning and characterization of fusion transcript, replicated in subsequent independent cases","pmids":["10556215"],"is_preprint":false}],"current_model":"Human (and orthologous eukaryotic) TOP1 is a type IB topoisomerase that transiently nicks one DNA strand to relieve torsional stress (supercoiling and R-loops) during replication and transcription; it acts coordinately with TOP2 at replication forks, adopts an open-to-closed conformational change upon DNA binding mediated by a conserved hinge, forms covalent TOP1 cleavage complexes (TOP1cc) that are normally transient but can become toxic when stabilized (by camptothecin or ribonucleotides embedded in DNA), and whose repair is initiated by TDP1 (regulated by CDK1 phosphorylation at S61 during mitosis) or alternatively by ERCC1-XPF (NER), MUS81, or NHEJ; TOP1 activity at ribonucleotides generates a characteristic deletion signature (ID4/ID-TOP1) through a sequential double-cleavage mechanism; PAR-ylation by PARP1 modulates TOP1 subnuclear dynamics and Top1cc stability; and TOP1 promotes repressed chromatin at rDNA and telomeres through its catalytic activity."},"narrative":{"mechanistic_narrative":"TOP1 is a type IB DNA topoisomerase that relieves torsional stress during DNA replication and transcription by transiently nicking one DNA strand through a covalent 3'-phosphotyrosyl cleavage intermediate (TOP1 cleavage complex, TOP1cc) [PMID:27257064, PMID:35013228]. Structural and mutagenic analysis of an archaeal ortholog defines an open-to-closed conformational transition gated by a conserved hinge tyrosine that permits DNA entry and underlies camptothecin sensitivity [PMID:35013228]. TOP1 acts coordinately with TOP2 at replication forks: loss of either alone is tolerated, but combined inactivation blocks fork progression and replication initiation and triggers checkpoint activation, establishing complementary topoisomerase roles in DNA replication from yeast to vertebrates [PMID:2997777, PMID:17671091, PMID:41791707]. Beyond replication, TOP1 catalytic activity promotes a repressed chromatin state at rDNA and telomeres, where its loss elevates histone H4 acetylation and derepresses proximal genes [PMID:18272174, PMID:12215413], and it cooperates with R-loops at highly expressed genes to generate transcription-associated double-strand breaks [PMID:38375218]. TOP1ccs are normally transient but become genotoxic when trapped — by camptothecin or by TOP1 action at genome-embedded ribonucleotides — generating recombination, chromosomal instability, and a characteristic short-deletion mutational signature (ID4/ID-TOP1) produced through a sequential double-cleavage mechanism within tandem repeats [PMID:24574527, PMID:25830313, PMID:27257064, PMID:35140396]. Trapped TOP1ccs are removed by redundant pathways: TDP1-mediated hydrolysis of the phosphotyrosyl bond, the ERCC1-XPF/RPA nucleolytic (NER) branch, MUS81 cleavage of proteolytically processed TOP1cc, and NHEJ, with TDP1 activity regulated by CDK1 phosphorylation at Ser61 during mitosis [PMID:26025908, PMID:30778235, PMID:37194054, PMID:39014228]. PARP1 is activated by trapped TOP1ccs and PARylates TOP1, controlling its nucleolar-to-nucleoplasmic dynamics and counteracting camptothecin-induced TOP1cc stabilization [PMID:27466387]. A recurrent NUP98-TOP1 fusion from the t(11;20) translocation is associated with therapy-related myelodysplastic syndrome/AML [PMID:10556215], and clinically acquired TOP1 point mutations confer resistance to TOP1-inhibitor antibody-drug-conjugate payloads [PMID:39745368].","teleology":[{"year":1985,"claim":"Established that TOP1 encodes DNA topoisomerase I and serves an auxiliary, partially redundant role to TOP2 in genome metabolism, resolving whether either enzyme is individually essential.","evidence":"Gene disruption and synthetic-lethality genetics in S. cerevisiae with temperature-sensitive top2 alleles","pmids":["2997777"],"confidence":"High","gaps":["Did not define the molecular reactions TOP1 and TOP2 share","Mechanism of synthetic lethality not resolved at this stage"]},{"year":2002,"claim":"Showed that covalent TOP1-DNA adducts are removed by two redundant pathways feeding recombinational restart, defining the core logic of TOP1cc repair.","evidence":"Genetic epistasis of tdp1 and rad1-rad10 mutants with camptothecin sensitivity in S. cerevisiae","pmids":["12368472"],"confidence":"High","gaps":["Biochemical step performed by each pathway not reconstituted here","Relative pathway contribution in mammals not addressed"]},{"year":2002,"claim":"Linked TOP1 catalytic activity to chromatin repression at rDNA, extending its role beyond torsional relief.","evidence":"Histone acetylation and chromatin accessibility assays in top1 vs sir2 deletion strains of S. cerevisiae","pmids":["12215413"],"confidence":"Medium","gaps":["Mechanism connecting catalytic activity to histone acetylation unclear","Single lab, yeast only"]},{"year":2007,"claim":"Demonstrated that TOP1 and TOP2 act coordinately within a narrow region around moving replication forks to prevent fork block and sister entanglement, defining where the auxiliary relationship operates.","evidence":"ChIP of replicating chromosomes plus checkpoint and γH2AX assays in top1/top2 mutants of S. cerevisiae","pmids":["17671091"],"confidence":"High","gaps":["Did not distinguish supercoil relief from decatenation contributions precisely","Vertebrate generality untested at this point"]},{"year":2008,"claim":"Extended TOP1's chromatin-repressive role to telomeres, showing catalytic activity is required to maintain repressed subtelomeric chromatin.","evidence":"ChIP for TOP1 binding and histone marks plus expression profiling with a Y727F catalytic-dead mutant in S. cerevisiae","pmids":["18272174"],"confidence":"Medium","gaps":["Direct causal link between strand cleavage and acetylation state not established","Single lab"]},{"year":2011,"claim":"Identified SUMO/STUbL signaling (Rad60, Nse2) as suppressors of spontaneous TOP1ccs that channel lesions into the NER-endonuclease repair branch, adding a regulatory layer to TOP1cc tolerance.","evidence":"Genetic epistasis and ChIP-qPCR detection of Top1ccs in S. pombe","pmids":["21408210"],"confidence":"High","gaps":["Direct SUMO/ubiquitin modification of TOP1 not demonstrated here","Mammalian counterpart not tested"]},{"year":2014,"claim":"Showed that TOP1ccs are physiologically stabilized and reversible at the rDNA replication fork barrier in a Fob1/Tof2-dependent manner, linking site-specific TOP1cc to rDNA stability.","evidence":"ChIP detection of Top1ccs at the rRFB, ectopic constructs, and fob1/tof2 epistasis in S. cerevisiae","pmids":["24574527"],"confidence":"High","gaps":["Function of these reversible Top1ccs in rDNA maintenance not fully defined","Relevance to mammalian rDNA unknown"]},{"year":2015,"claim":"Established that trapped TOP1ccs are recombinogenic, driving crossovers and tandem-array instability, connecting TOP1cc trapping to genome instability.","evidence":"Sectoring/LOH assays, chromosome microarrays, and long-term subculturing in S. cerevisiae","pmids":["25830313"],"confidence":"Medium","gaps":["Exact lesion converting Top1cc to a recombination substrate not defined","Single organism"]},{"year":2015,"claim":"Reconstituted an ERCC1-XPF/RPA nucleolytic pathway for repairing the TOP1 3'-phosphotyrosyl lesion, establishing a TDP1-independent NER-based repair route in mammals.","evidence":"In vitro nuclease and repair-synthesis reconstitution on Tyr-nick substrates plus co-localization in CPT-treated cells","pmids":["26025908"],"confidence":"High","gaps":["In vivo relative contribution vs TDP1 not quantified here","Substrate used intact rather than full protein-DNA crosslink"]},{"year":2016,"claim":"Defined the sequential double-cleavage mechanism by which TOP1 activity at ribonucleotides generates short tandem-repeat deletions, explaining a mutagenic consequence of TOP1 catalysis.","evidence":"Parallel in vitro cleavage/ligation assays and in vivo yeast mutagenesis with systematic substrate variation","pmids":["27257064"],"confidence":"High","gaps":["Did not establish frequency of this process in mammalian genomes","Determinants of intermediate realignment beyond complementarity not exhausted"]},{"year":2016,"claim":"Showed that PARP1 PARylates trapped TOP1 to control its subnuclear dynamics and counteract TOP1cc stabilization, defining a post-translational regulatory mechanism for TOP1cc resolution.","evidence":"Live-cell FRAP of GFP-TOP1 with PARP inhibitors and W205/N722S mutagenesis in human cells","pmids":["27466387"],"confidence":"High","gaps":["Identity of PARylated residues on TOP1 not mapped","Downstream repair enzymes coupled to PARylation not defined"]},{"year":2019,"claim":"Identified Apn2 as an additional 3'-end processing enzyme acting on TOP1cc and ribonucleotide-derived termini, broadening the repertoire of TOP1cc-resolving activities.","evidence":"Genetic epistasis plus biochemical assays with purified Apn2 on defined substrates in S. cerevisiae","pmids":["30778235"],"confidence":"High","gaps":["Mammalian ortholog contribution not addressed","Hierarchy relative to TDP1 in vivo unclear"]},{"year":2019,"claim":"Demonstrated that NHEJ produces large TOP1cc-dependent deletions outside replication, including in quiescent cells, showing TOP1 lesions cause DSBs independent of fork passage.","evidence":"Genetic analysis of NHEJ mutants and deletion-junction characterization in quiescent S. cerevisiae","pmids":["31636207"],"confidence":"Medium","gaps":["Origin of the underlying DSBs not directly visualized","Single organism"]},{"year":2022,"claim":"Established that in mammals TOP1 action at unrepaired embedded ribonucleotides generates the ID4/ID-TOP1 indel signature contributing to cancer and germline de novo mutations, linking the yeast mechanism to human disease.","evidence":"Genome sequencing of ribonucleotide-excision-repair-deficient mammals, signature analysis, and TOP1 inhibition","pmids":["35140396"],"confidence":"High","gaps":["Tissue-specific drivers of the signature not fully mapped","Therapeutic exploitation untested"]},{"year":2022,"claim":"Resolved the conformational mechanism of catalysis, showing a hinge tyrosine gates an open-to-closed transition required for activity and camptothecin sensitivity.","evidence":"Crystal structure of apo archaeal CsTOP1 with directed hinge mutagenesis and CPT sensitivity assays","pmids":["35013228"],"confidence":"High","gaps":["Structure is of an archaeal ortholog, not human TOP1","Dynamics during strand cleavage not captured"]},{"year":2022,"claim":"Dissected the human TOP1cc DSB-conversion pathway, showing MUS81 (not APEX) generates excess DSBs in TDP1-deficient cells, clarifying parallel repair routing.","evidence":"Genome-wide CRISPR screens and isogenic TDP1/MUS81 and TDP1/APEX KO cell lines with DNA damage assays","pmids":["35869071"],"confidence":"Medium","gaps":["Did not define how TOP1cc is dislodged before MUS81 cleavage","Single lab"]},{"year":2023,"claim":"Showed MUS81 requires proteolytic processing of TOP1 before cleaving TOP1cc and acts non-redundantly with TDP1, defining substrate requirements for the nucleolytic branch.","evidence":"In vitro MUS81 cleavage assays with intact vs processed TOP1-DNA substrates plus double-KO cell assays","pmids":["37194054"],"confidence":"High","gaps":["Protease responsible for TOP1 processing not identified here","In vivo ordering of processing and cleavage not resolved"]},{"year":2023,"claim":"Established that TDP1 suppresses chromosomal translocations from transcription-associated TOP1 DSBs repaired error-prone by MRN/NHEJ, connecting TOP1cc repair to genome rearrangement.","evidence":"Loss-of-function genetics with cytogenetic translocation analysis and MRN/NHEJ epistasis in human cells","pmids":["37945566"],"confidence":"Medium","gaps":["Genomic loci of recurrent translocations not mapped","Single lab"]},{"year":2024,"claim":"Identified CDK1 phosphorylation of TDP1 at Ser61 as the switch enabling MUS81-dependent mitotic TOP1cc repair, defining cell-cycle regulation of the repair choice.","evidence":"CDK1 kinase assay, TDP1-S61A mutant cells, live imaging, and MUS81 epistasis in human cells","pmids":["39014228"],"confidence":"High","gaps":["Other mitotic substrates and regulators not excluded","Mechanism of S61A chromatin trapping incompletely defined"]},{"year":2024,"claim":"Showed TOP1 and R-loops cooperate at highly expressed genes to generate transcriptional DSBs implicated in early cancer changes, linking TOP1 catalysis to transcription-associated genome instability.","evidence":"Genome-wide DSB, R-loop, and TOP1cc mapping with TOP1 and R-loop depletion in human cells","pmids":["38375218"],"confidence":"Medium","gaps":["Causal step from R-loop/TOP1 to DSB not biochemically reconstituted","Single lab"]},{"year":2025,"claim":"Functionally characterized clinically acquired TOP1 mutations that reduce activity and covalent DNA binding to confer resistance to TOP1-inhibitor ADC payloads, providing a mechanism of acquired therapy resistance.","evidence":"Enzymatic, covalent-binding, and in vitro drug-resistance assays on recombinant mutant TOP1 with plasma genotyping","pmids":["39745368"],"confidence":"Medium","gaps":["In vivo tumor resistance not directly demonstrated","Structural basis of each mutation not resolved"]},{"year":2026,"claim":"Confirmed in vertebrate cells that TOP1 and TOP2 act complementarily in both replication initiation and progression, generalizing the yeast auxiliary relationship to higher eukaryotes.","evidence":"Conditional TOP1 depletion plus TOP2 inhibition with DNA fiber assays and cell cycle analysis in DT40 cells","pmids":["41791707"],"confidence":"Medium","gaps":["Did not distinguish supercoiling vs decatenation contributions at initiation","Single cell-line model"]},{"year":null,"claim":"How TOP1 PARylation, proteolytic processing, and cell-cycle signaling are integrated to select among the multiple competing TOP1cc repair pathways in human cells remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model coordinating TDP1, ERCC1-XPF, MUS81, Apn2, and NHEJ choice","PARylation target residues on human TOP1 unmapped","Structural model of human TOP1cc engagement by each repair enzyme lacking"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140097","term_label":"catalytic activity, acting on DNA","supporting_discovery_ids":[9,13,19]},{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[3,8,19]},{"term_id":"GO:0016853","term_label":"isomerase activity","supporting_discovery_ids":[9,13]}],"localization":[{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[8]},{"term_id":"GO:0005654","term_label":"nucleoplasm","supporting_discovery_ids":[8]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[3,22]}],"pathway":[{"term_id":"R-HSA-69306","term_label":"DNA Replication","supporting_discovery_ids":[2,21]},{"term_id":"R-HSA-73894","term_label":"DNA 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Netherlands)","url":"https://pubmed.ncbi.nlm.nih.gov/34974222","citation_count":5,"is_preprint":false},{"pmid":"31896687","id":"PMC_31896687","title":"Top1 and Top2 promote replication fork arrest at a programmed pause site.","date":"2020","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/31896687","citation_count":5,"is_preprint":false},{"pmid":"36294786","id":"PMC_36294786","title":"Personalized Prescription of Chemotherapy Based on Assessment of mRNA Expression of BRCA1, RRM1, ERCC1, TOP1, TOP2α, TUBβ3, TYMS, and GSTP1 Genes in Tumors Compared to Standard Chemotherapy in the Treatment of Non-Small-Cell Lung Cancer.","date":"2022","source":"Journal of personalized medicine","url":"https://pubmed.ncbi.nlm.nih.gov/36294786","citation_count":5,"is_preprint":false},{"pmid":"40102901","id":"PMC_40102901","title":"MicroRNA-24-3p targeting Top1 in perirenal fat is involved in circulating inflammation and high cardiovascular disease risk in patients with primary 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mutants (delta top1) are viable, but delta top1 top2-ts double mutants are inviable even when mitotic progression is blocked, demonstrating that Top1 serves an auxiliary role to Top2 in yeast DNA metabolism.\",\n      \"method\": \"Gene disruption/deletion in S. cerevisiae, complementation, temperature-shift experiments with synchronous cultures\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean loss-of-function genetics with defined viability phenotype, replicated with multiple alleles in foundational study\",\n      \"pmids\": [\"2997777\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"In S. cerevisiae, Tdp1 (tyrosyl-DNA phosphodiesterase) and the Rad1-Rad10 structure-specific endonuclease act as two primary, redundant pathways to remove covalent Top1-DNA adducts (Top1 cleavage complexes) at replication forks; tdp1 rad1 double mutants are highly sensitive to camptothecin and show a TOP1-dependent growth delay. Both pathways funnel into RAD52/RAD51/RAD50-dependent recombination for replication restart.\",\n      \"method\": \"Genetic epistasis analysis (double/triple mutants), camptothecin sensitivity assays in S. cerevisiae\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — extensive epistasis with multiple alleles, rigorous genetic controls, replicated across multiple pathway genes\",\n      \"pmids\": [\"12368472\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Top1 and Top2 act within a ~600-bp region spanning moving replication forks in S. cerevisiae. TOP1 ablation alone does not block fork progression or activate the Rad53 checkpoint, but top1 top2 double mutants exhibit replication fork block, fork processing, and Rad53/γH2AX activation in S phase, demonstrating that Top1 and Top2 act coordinately to relieve torsional stress and prevent sister chromatid entanglement at forks.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP) of replicating chromosomes, genetic analysis of top1, top2, and top1 top2 mutants, checkpoint kinase (Rad53) phosphorylation assays, γH2AX detection\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (ChIP, genetics, checkpoint assays) in a single rigorous study\",\n      \"pmids\": [\"17671091\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Top1 binds to and is catalytically active at telomeric DNA repeats in S. cerevisiae; deletion of TOP1 or expression of catalytically inactive Top1-Y727F increases H4 histone acetylation at subtelomeric regions and activates transcription of telomere-proximal genes, indicating that Top1 catalytic activity promotes a repressed chromatin state at telomeres.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP) for Top1 binding and histone modifications, expression profiling (microarray), TOP1 deletion and Y727F catalytic-dead mutant\",\n      \"journal\": \"Journal of molecular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP and expression profiling with catalytic mutant, single lab, two orthogonal methods\",\n      \"pmids\": [\"18272174\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"In fission yeast, STUbL (SUMO-targeted ubiquitin ligase), Rad60, and the Nse2 SUMO E3 ligase suppress spontaneous Top1-mediated DNA damage (Top1 cleavage complexes). Cells lacking STUbL activity become dependent on Tdp1, and Top1ccs detected by ChIP-qPCR are elevated when STUbL, Rad60, or Nse2 activity is absent. Tdp1 and Rad16-Swi10 (NER endonuclease) constitute the two major Top1cc repair pathways, with STUbL/Rad60/Nse2 facilitating the Rad16-Swi10 branch.\",\n      \"method\": \"Genetic epistasis, chromatin immunoprecipitation (ChIP)-qPCR to detect Top1ccs, camptothecin sensitivity assays in S. pombe\",\n      \"journal\": \"PLoS genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — ChIP-qPCR for direct Top1cc detection combined with extensive epistasis analysis, multiple orthogonal methods\",\n      \"pmids\": [\"21408210\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Top1 cleavage complexes (Top1ccs) are specifically stabilized at the ribosomal replication fork barrier (rRFB) in S. cerevisiae in a strand-specific, replication-independent manner requiring Fob1 and Tof2; these Top1ccs are reversible (not subject to Tdp1 or Mus81 repair) and their presence correlates with Top1-dependent rDNA stability.\",\n      \"method\": \"ChIP detection of Top1ccs at rRFB, ectopic rRFB constructs, genetic analysis with fob1 and tof2 mutants, Tdp1/Mus81 epistasis in S. cerevisiae\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct ChIP detection of Top1ccs, ectopic construct validation, epistasis analysis — multiple orthogonal methods\",\n      \"pmids\": [\"24574527\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Trapping of the Top1 cleavage complex (Top1cc) in S. cerevisiae (via camptothecin or top1-T722A allele) causes a 5–10-fold increase in reciprocal crossover events and greatly elevated instability within rDNA and CUP1 tandem arrays, with evidence that recombinogenic lesions arise during or after DNA synthesis.\",\n      \"method\": \"Sectoring assay for loss of heterozygosity, chromosome microarrays, sub-culturing for 250 generations, genetic analysis in S. cerevisiae\",\n      \"journal\": \"PLoS genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — quantitative genetic assay plus genome-wide microarray, single lab, two orthogonal methods\",\n      \"pmids\": [\"25830313\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"ERCC1-XPF endonuclease and RPA participate in NER-mediated repair of Top1-attached nick DNA lesions (3'-phosphotyrosyl bond). ERCC1-XPF cleaves a Tyr-nick DNA substrate in vitro in the presence of RPA, and repair DNA synthesis of these lesions occurs with NER factors (ERCC1-XPF, RPA, DNA pol δ, FEN1, DNA ligase 1).\",\n      \"method\": \"In vitro nuclease assay on Tyr-nick DNA substrate, co-localization of ERCC1-XPF and RPA in CPT-treated cells, in vitro repair synthesis reconstitution\",\n      \"journal\": \"Carcinogenesis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution of repair synthesis plus in vivo co-localization; multiple orthogonal methods\",\n      \"pmids\": [\"26025908\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"PARP1 is activated by trapped Top1 cleavage complexes (Top1cc) and PARylates Top1. Using live-cell FRAP of GFP-tagged human Top1 and PARP inhibitors (ABT-888/Veliparib), PARP inhibition was shown to delocalize Top1 from nucleolus to nucleoplasm and to markedly increase the CPT-induced immobile/bound fraction of Top1 (Top1cc) genome-wide, indicating that PAR-modification of Top1 counteracts CPT-induced stabilization of Top1cc. Top1 residues Trp205 and Asn722 are critical for subnuclear dynamics; Top1-N722S is restricted to the nucleolus under CPT due to deficient PARylation.\",\n      \"method\": \"Live-cell imaging of GFP-tagged Top1, FRAP with kinetic modelling, PARP inhibitor treatment, site-directed mutagenesis (W205, N722S)\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — direct live-cell imaging + FRAP kinetics + mutagenesis, multiple orthogonal methods in single lab\",\n      \"pmids\": [\"27466387\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Top1-dependent ribonucleotide-associated deletions in yeast arise through a sequential double-cleavage mechanism: Top1 cleaves at a ribonucleotide, the resulting intermediate realigns within a tandem repeat, and Top1 ligates the realigned nick, generating a single-repeat deletion. Deletion frequency is inversely related to the distance between two Top1 cleavage sites, and complementarity downstream of the gap promotes deletions more effectively than upstream complementarity.\",\n      \"method\": \"Parallel in vitro Top1 cleavage/ligation assays and in vivo yeast mutagenesis assays, systematic variation of substrate distances and complementarity\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution with parallel in vivo validation; mechanistic model tested with multiple substrate variants\",\n      \"pmids\": [\"27257064\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Top1cc-dependent large deletions (~100 bp) in yeast are produced by the NHEJ pathway and require Top1cc removal from DNA ends; these events accumulate in quiescent cells, suggesting that causative DSBs arise outside replication context. Joining of ends from distinct Top1-associated DSBs via NHEJ deletes the intervening sequence.\",\n      \"method\": \"Genetic analysis of NHEJ mutants in S. cerevisiae, characterization of deletion junctions, quiescence experiments\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis with junction characterization, single lab, two orthogonal approaches\",\n      \"pmids\": [\"31636207\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Yeast Apn2 (AP endonuclease 2) resolves Top1-mediated phosphotyrosine-DNA conjugates (Top1cc), terminal 2',3'-cyclic phosphates, and their hydrolyzed products at 3' DNA ends, and suppresses 2-bp slippage mutagenesis in RNH201-deficient cells, defining Apn2 as an alternative Top1cc repair enzyme.\",\n      \"method\": \"Genetic epistasis, biochemical assays with purified Apn2 on Top1cc and cyclic-phosphate substrates in S. cerevisiae\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro biochemical assay with defined substrates plus in vivo genetics; multiple methods, single lab\",\n      \"pmids\": [\"30778235\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Defective ribonucleotide excision repair in mammals leads to accumulation of genome-embedded ribonucleotides, at which TOP1 activity generates short (2–5 bp) deletions (ID4/ID-TOP1 mutation signature) at TNT motifs; this transcription-associated mutagenesis process contributes to cancer somatic mutations and germline de novo indels.\",\n      \"method\": \"Genome sequencing of cancer and germline samples from ribonucleotide excision repair-deficient mammals, mutational signature analysis, functional TOP1 inhibition experiments\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — mechanistic experiments in mammalian cells combined with human cancer genome analysis; replicated across multiple biological contexts\",\n      \"pmids\": [\"35140396\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Archaeal CsTOP1 (a structural ortholog of eukaryotic TOP1) adopts an open conformation in apo form, with the hinge loop (5-residue junction between CAP and CAT modules) permitting rotation that allows DNA entry. A conserved tyrosine near the hinge mediates transition to the closed conformation upon DNA binding; directed mutagenesis of hinge residues confirmed this flexibility is required for activity and linked to camptothecin sensitivity.\",\n      \"method\": \"Crystal structure of apo CsTOP1, directed mutagenesis of hinge residues, camptothecin sensitivity assays\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure plus mutagenesis validation; multiple orthogonal methods in single study\",\n      \"pmids\": [\"35013228\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Whole-genome CRISPR screens and co-deficient cell lines demonstrated that MUS81 mediates the generation of excess double-strand breaks (DSBs) in TDP1-KO cells exposed to Top1 poisons; APEX1/2 are synthetically lethal with TDP1 but do not reduce DSB formation in TDP1-KO cells. Top1cc can be resolved directly by TDP1 or converted to DSBs and repaired by homologous recombination.\",\n      \"method\": \"Genome-wide CRISPR screens, generation of co-deficient cell lines (TDP1/MUS81 and TDP1/APEX KO), DNA damage assays\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR genetic screens plus isogenic KO cell validation, single lab\",\n      \"pmids\": [\"35869071\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"TDP1 suppresses chromosomal translocations arising from transcription-associated TOP1-induced DSBs; error-prone repair of these breaks relies on the MRN complex and canonical NHEJ. TDP1 loss leads to increased chromosome rearrangements specifically linked to transcription-coupled TOP1 activity.\",\n      \"method\": \"Loss-of-function genetics in human cells, cytogenetic analysis of translocations, epistasis with MRN and NHEJ factors\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis with cytogenetic readout, single lab, two orthogonal methods\",\n      \"pmids\": [\"37945566\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"MUS81 endonuclease can efficiently cleave DNA substrates bearing proteolytically processed (but not intact) TOP1, indicating TOP1 must be dislodged or partially degraded before MUS81 cleavage. MUS81 and TDP1 play independent, non-redundant roles in repair of CPT-induced TOP1-DPC lesions.\",\n      \"method\": \"In vitro MUS81 cleavage assays with fluorescein-, streptavidin-, and proteolytically processed TOP1-modified DNA substrates; nuclear extract assays; TDP1/MUS81 double-KO cell sensitivity assays\",\n      \"journal\": \"BMC biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with multiple substrate types plus cellular genetic validation; multiple orthogonal methods\",\n      \"pmids\": [\"37194054\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"CDK1 phosphorylates TDP1 at Ser61 during mitosis; TDP1-S61A (non-phosphorylatable) is trapped on mitotic chromosomes and causes DNA damage and mitotic defects. Top1cc repair in mitosis occurs via a MUS81-dependent mechanism, and TDP1-S61A enrichment at common fragile sites over-stimulates MUS81-dependent chromatid breaks, leading to anaphase bridges, micronuclei, and 53BP1 nuclear bodies in G1.\",\n      \"method\": \"CDK1 kinase assay identifying TDP1 S61 phosphorylation, TDP1-S61A mutant cell lines, live imaging, γH2AX/53BP1 focus analysis, MUS81 epistasis\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — biochemical kinase assay identifying modification site plus mutant cell functional analysis; multiple orthogonal methods in single lab\",\n      \"pmids\": [\"39014228\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"TOP1 and R-loops are enriched at highly expressed genes where transcriptional DSBs occur; depletion of R-loops or TOP1 significantly reduces transcription-associated DSBs at these loci, establishing that TOP1 and R-loops cooperate in transcriptional DSB formation and contribute to early molecular changes driving cancer development.\",\n      \"method\": \"Genome-wide DSB mapping, R-loop mapping, TOP1cc mapping, TOP1 depletion and R-loop depletion in human cells\",\n      \"journal\": \"iScience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genome-wide mapping plus loss-of-function experiments, single lab, multiple orthogonal mapping methods\",\n      \"pmids\": [\"38375218\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"TOP1 mutations (S57C, R364H, W401C, G359E) arising under selective pressure from ADC therapy in metastatic breast cancer patients exhibit reduced enzymatic activity, attenuated covalent DNA binding, and resistance to TOP1-inhibitor payloads SN38 and deruxtecan, functionally characterizing resistance-associated TOP1 mutations.\",\n      \"method\": \"Functional characterization of recombinant mutant TOP1 proteins (enzymatic activity assays, covalent DNA binding assays, in vitro drug resistance assays), plasma-based genotyping\",\n      \"journal\": \"Clinical cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro biochemical characterization of mutant proteins with multiple assays, single lab\",\n      \"pmids\": [\"39745368\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"TOP1 interacts with BTBD1 and BTBD2; this interaction requires human TOP1 residues 236 and 237. The same residues are required for TOP1 to enhance infectivity of HIV-1 virions when expressed in African Green Monkey producer cells. RNAi knockdown of BTBD2 in AGM and human cells increased HIV-1 permissiveness 2–3-fold.\",\n      \"method\": \"Co-IP/interaction assays with TOP1 point mutants (residues 236/237), RNAi knockdown of BTBD2, HIV-1 infectivity assays\",\n      \"journal\": \"Virology journal\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP and RNAi experiment, single lab, limited mechanistic follow-up\",\n      \"pmids\": [\"21092135\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"In vertebrate (chicken DT40) cells, TOP1 depletion alone does not affect replication fork progression, and TOP2 inhibition (ICRF193) alone also has no effect; however, combined TOP1 depletion plus TOP2 inhibition nearly abolishes DNA replication, causing S-phase arrest and apoptosis. TOP1 depletion combined with TOP2 poison (etoposide) impairs replication initiation rather than elongation. This demonstrates complementary roles of TOP1 and TOP2 in both initiation and progression of DNA replication in vertebrates.\",\n      \"method\": \"Conditional TOP1 depletion in DT40 cells, ICRF193 and etoposide pharmacological inhibition, DNA fiber assays for fork progression, cell cycle analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional genetic depletion plus pharmacological inhibition with DNA fiber assays, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"41791707\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Deletion of TOP1 in S. cerevisiae increases histone H4 acetylation at the rDNA locus and renders rDNA chromatin more accessible, similar to sir2 mutants, indicating that Top1 activity promotes a repressed chromatin state at rDNA.\",\n      \"method\": \"Chromatin accessibility assays, histone acetylation analysis at rDNA in top1 and sir2 deletion strains of S. cerevisiae\",\n      \"journal\": \"Journal of molecular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct chromatin modification assays in loss-of-function mutants, single lab\",\n      \"pmids\": [\"12215413\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"The t(11;20)(p15;q11) chromosomal translocation produces an NUP98-TOP1 chimeric mRNA encoding the NUP98 FXFG repeats fused to the body of DNA topoisomerase I (TOP1), with the breakpoint in TOP1 occurring within the gene; this fusion is associated with therapy-related myelodysplastic syndrome/acute myeloid leukemia.\",\n      \"method\": \"Cloning and sequencing of t(11;20) translocation breakpoints, RT-PCR characterization of fusion transcript in patient samples\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct molecular cloning and characterization of fusion transcript, replicated in subsequent independent cases\",\n      \"pmids\": [\"10556215\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"Human (and orthologous eukaryotic) TOP1 is a type IB topoisomerase that transiently nicks one DNA strand to relieve torsional stress (supercoiling and R-loops) during replication and transcription; it acts coordinately with TOP2 at replication forks, adopts an open-to-closed conformational change upon DNA binding mediated by a conserved hinge, forms covalent TOP1 cleavage complexes (TOP1cc) that are normally transient but can become toxic when stabilized (by camptothecin or ribonucleotides embedded in DNA), and whose repair is initiated by TDP1 (regulated by CDK1 phosphorylation at S61 during mitosis) or alternatively by ERCC1-XPF (NER), MUS81, or NHEJ; TOP1 activity at ribonucleotides generates a characteristic deletion signature (ID4/ID-TOP1) through a sequential double-cleavage mechanism; PAR-ylation by PARP1 modulates TOP1 subnuclear dynamics and Top1cc stability; and TOP1 promotes repressed chromatin at rDNA and telomeres through its catalytic activity.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"TOP1 is a type IB DNA topoisomerase that relieves torsional stress during DNA replication and transcription by transiently nicking one DNA strand through a covalent 3'-phosphotyrosyl cleavage intermediate (TOP1 cleavage complex, TOP1cc) [#9, #13]. Structural and mutagenic analysis of an archaeal ortholog defines an open-to-closed conformational transition gated by a conserved hinge tyrosine that permits DNA entry and underlies camptothecin sensitivity [#13]. TOP1 acts coordinately with TOP2 at replication forks: loss of either alone is tolerated, but combined inactivation blocks fork progression and replication initiation and triggers checkpoint activation, establishing complementary topoisomerase roles in DNA replication from yeast to vertebrates [#0, #2, #21]. Beyond replication, TOP1 catalytic activity promotes a repressed chromatin state at rDNA and telomeres, where its loss elevates histone H4 acetylation and derepresses proximal genes [#3, #22], and it cooperates with R-loops at highly expressed genes to generate transcription-associated double-strand breaks [#18]. TOP1ccs are normally transient but become genotoxic when trapped — by camptothecin or by TOP1 action at genome-embedded ribonucleotides — generating recombination, chromosomal instability, and a characteristic short-deletion mutational signature (ID4/ID-TOP1) produced through a sequential double-cleavage mechanism within tandem repeats [#5, #6, #9, #12]. Trapped TOP1ccs are removed by redundant pathways: TDP1-mediated hydrolysis of the phosphotyrosyl bond, the ERCC1-XPF/RPA nucleolytic (NER) branch, MUS81 cleavage of proteolytically processed TOP1cc, and NHEJ, with TDP1 activity regulated by CDK1 phosphorylation at Ser61 during mitosis [#7, #11, #16, #17]. PARP1 is activated by trapped TOP1ccs and PARylates TOP1, controlling its nucleolar-to-nucleoplasmic dynamics and counteracting camptothecin-induced TOP1cc stabilization [#8]. A recurrent NUP98-TOP1 fusion from the t(11;20) translocation is associated with therapy-related myelodysplastic syndrome/AML [#23], and clinically acquired TOP1 point mutations confer resistance to TOP1-inhibitor antibody-drug-conjugate payloads [#19].\",\n  \"teleology\": [\n    {\n      \"year\": 1985,\n      \"claim\": \"Established that TOP1 encodes DNA topoisomerase I and serves an auxiliary, partially redundant role to TOP2 in genome metabolism, resolving whether either enzyme is individually essential.\",\n      \"evidence\": \"Gene disruption and synthetic-lethality genetics in S. cerevisiae with temperature-sensitive top2 alleles\",\n      \"pmids\": [\"2997777\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define the molecular reactions TOP1 and TOP2 share\", \"Mechanism of synthetic lethality not resolved at this stage\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Showed that covalent TOP1-DNA adducts are removed by two redundant pathways feeding recombinational restart, defining the core logic of TOP1cc repair.\",\n      \"evidence\": \"Genetic epistasis of tdp1 and rad1-rad10 mutants with camptothecin sensitivity in S. cerevisiae\",\n      \"pmids\": [\"12368472\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Biochemical step performed by each pathway not reconstituted here\", \"Relative pathway contribution in mammals not addressed\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Linked TOP1 catalytic activity to chromatin repression at rDNA, extending its role beyond torsional relief.\",\n      \"evidence\": \"Histone acetylation and chromatin accessibility assays in top1 vs sir2 deletion strains of S. cerevisiae\",\n      \"pmids\": [\"12215413\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism connecting catalytic activity to histone acetylation unclear\", \"Single lab, yeast only\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Demonstrated that TOP1 and TOP2 act coordinately within a narrow region around moving replication forks to prevent fork block and sister entanglement, defining where the auxiliary relationship operates.\",\n      \"evidence\": \"ChIP of replicating chromosomes plus checkpoint and γH2AX assays in top1/top2 mutants of S. cerevisiae\",\n      \"pmids\": [\"17671091\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not distinguish supercoil relief from decatenation contributions precisely\", \"Vertebrate generality untested at this point\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Extended TOP1's chromatin-repressive role to telomeres, showing catalytic activity is required to maintain repressed subtelomeric chromatin.\",\n      \"evidence\": \"ChIP for TOP1 binding and histone marks plus expression profiling with a Y727F catalytic-dead mutant in S. cerevisiae\",\n      \"pmids\": [\"18272174\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct causal link between strand cleavage and acetylation state not established\", \"Single lab\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Identified SUMO/STUbL signaling (Rad60, Nse2) as suppressors of spontaneous TOP1ccs that channel lesions into the NER-endonuclease repair branch, adding a regulatory layer to TOP1cc tolerance.\",\n      \"evidence\": \"Genetic epistasis and ChIP-qPCR detection of Top1ccs in S. pombe\",\n      \"pmids\": [\"21408210\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct SUMO/ubiquitin modification of TOP1 not demonstrated here\", \"Mammalian counterpart not tested\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Showed that TOP1ccs are physiologically stabilized and reversible at the rDNA replication fork barrier in a Fob1/Tof2-dependent manner, linking site-specific TOP1cc to rDNA stability.\",\n      \"evidence\": \"ChIP detection of Top1ccs at the rRFB, ectopic constructs, and fob1/tof2 epistasis in S. cerevisiae\",\n      \"pmids\": [\"24574527\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Function of these reversible Top1ccs in rDNA maintenance not fully defined\", \"Relevance to mammalian rDNA unknown\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Established that trapped TOP1ccs are recombinogenic, driving crossovers and tandem-array instability, connecting TOP1cc trapping to genome instability.\",\n      \"evidence\": \"Sectoring/LOH assays, chromosome microarrays, and long-term subculturing in S. cerevisiae\",\n      \"pmids\": [\"25830313\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Exact lesion converting Top1cc to a recombination substrate not defined\", \"Single organism\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Reconstituted an ERCC1-XPF/RPA nucleolytic pathway for repairing the TOP1 3'-phosphotyrosyl lesion, establishing a TDP1-independent NER-based repair route in mammals.\",\n      \"evidence\": \"In vitro nuclease and repair-synthesis reconstitution on Tyr-nick substrates plus co-localization in CPT-treated cells\",\n      \"pmids\": [\"26025908\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo relative contribution vs TDP1 not quantified here\", \"Substrate used intact rather than full protein-DNA crosslink\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Defined the sequential double-cleavage mechanism by which TOP1 activity at ribonucleotides generates short tandem-repeat deletions, explaining a mutagenic consequence of TOP1 catalysis.\",\n      \"evidence\": \"Parallel in vitro cleavage/ligation assays and in vivo yeast mutagenesis with systematic substrate variation\",\n      \"pmids\": [\"27257064\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not establish frequency of this process in mammalian genomes\", \"Determinants of intermediate realignment beyond complementarity not exhausted\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Showed that PARP1 PARylates trapped TOP1 to control its subnuclear dynamics and counteract TOP1cc stabilization, defining a post-translational regulatory mechanism for TOP1cc resolution.\",\n      \"evidence\": \"Live-cell FRAP of GFP-TOP1 with PARP inhibitors and W205/N722S mutagenesis in human cells\",\n      \"pmids\": [\"27466387\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Identity of PARylated residues on TOP1 not mapped\", \"Downstream repair enzymes coupled to PARylation not defined\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Identified Apn2 as an additional 3'-end processing enzyme acting on TOP1cc and ribonucleotide-derived termini, broadening the repertoire of TOP1cc-resolving activities.\",\n      \"evidence\": \"Genetic epistasis plus biochemical assays with purified Apn2 on defined substrates in S. cerevisiae\",\n      \"pmids\": [\"30778235\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mammalian ortholog contribution not addressed\", \"Hierarchy relative to TDP1 in vivo unclear\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Demonstrated that NHEJ produces large TOP1cc-dependent deletions outside replication, including in quiescent cells, showing TOP1 lesions cause DSBs independent of fork passage.\",\n      \"evidence\": \"Genetic analysis of NHEJ mutants and deletion-junction characterization in quiescent S. cerevisiae\",\n      \"pmids\": [\"31636207\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Origin of the underlying DSBs not directly visualized\", \"Single organism\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Established that in mammals TOP1 action at unrepaired embedded ribonucleotides generates the ID4/ID-TOP1 indel signature contributing to cancer and germline de novo mutations, linking the yeast mechanism to human disease.\",\n      \"evidence\": \"Genome sequencing of ribonucleotide-excision-repair-deficient mammals, signature analysis, and TOP1 inhibition\",\n      \"pmids\": [\"35140396\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Tissue-specific drivers of the signature not fully mapped\", \"Therapeutic exploitation untested\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Resolved the conformational mechanism of catalysis, showing a hinge tyrosine gates an open-to-closed transition required for activity and camptothecin sensitivity.\",\n      \"evidence\": \"Crystal structure of apo archaeal CsTOP1 with directed hinge mutagenesis and CPT sensitivity assays\",\n      \"pmids\": [\"35013228\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structure is of an archaeal ortholog, not human TOP1\", \"Dynamics during strand cleavage not captured\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Dissected the human TOP1cc DSB-conversion pathway, showing MUS81 (not APEX) generates excess DSBs in TDP1-deficient cells, clarifying parallel repair routing.\",\n      \"evidence\": \"Genome-wide CRISPR screens and isogenic TDP1/MUS81 and TDP1/APEX KO cell lines with DNA damage assays\",\n      \"pmids\": [\"35869071\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not define how TOP1cc is dislodged before MUS81 cleavage\", \"Single lab\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Showed MUS81 requires proteolytic processing of TOP1 before cleaving TOP1cc and acts non-redundantly with TDP1, defining substrate requirements for the nucleolytic branch.\",\n      \"evidence\": \"In vitro MUS81 cleavage assays with intact vs processed TOP1-DNA substrates plus double-KO cell assays\",\n      \"pmids\": [\"37194054\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Protease responsible for TOP1 processing not identified here\", \"In vivo ordering of processing and cleavage not resolved\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Established that TDP1 suppresses chromosomal translocations from transcription-associated TOP1 DSBs repaired error-prone by MRN/NHEJ, connecting TOP1cc repair to genome rearrangement.\",\n      \"evidence\": \"Loss-of-function genetics with cytogenetic translocation analysis and MRN/NHEJ epistasis in human cells\",\n      \"pmids\": [\"37945566\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Genomic loci of recurrent translocations not mapped\", \"Single lab\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identified CDK1 phosphorylation of TDP1 at Ser61 as the switch enabling MUS81-dependent mitotic TOP1cc repair, defining cell-cycle regulation of the repair choice.\",\n      \"evidence\": \"CDK1 kinase assay, TDP1-S61A mutant cells, live imaging, and MUS81 epistasis in human cells\",\n      \"pmids\": [\"39014228\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Other mitotic substrates and regulators not excluded\", \"Mechanism of S61A chromatin trapping incompletely defined\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Showed TOP1 and R-loops cooperate at highly expressed genes to generate transcriptional DSBs implicated in early cancer changes, linking TOP1 catalysis to transcription-associated genome instability.\",\n      \"evidence\": \"Genome-wide DSB, R-loop, and TOP1cc mapping with TOP1 and R-loop depletion in human cells\",\n      \"pmids\": [\"38375218\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Causal step from R-loop/TOP1 to DSB not biochemically reconstituted\", \"Single lab\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Functionally characterized clinically acquired TOP1 mutations that reduce activity and covalent DNA binding to confer resistance to TOP1-inhibitor ADC payloads, providing a mechanism of acquired therapy resistance.\",\n      \"evidence\": \"Enzymatic, covalent-binding, and in vitro drug-resistance assays on recombinant mutant TOP1 with plasma genotyping\",\n      \"pmids\": [\"39745368\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"In vivo tumor resistance not directly demonstrated\", \"Structural basis of each mutation not resolved\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Confirmed in vertebrate cells that TOP1 and TOP2 act complementarily in both replication initiation and progression, generalizing the yeast auxiliary relationship to higher eukaryotes.\",\n      \"evidence\": \"Conditional TOP1 depletion plus TOP2 inhibition with DNA fiber assays and cell cycle analysis in DT40 cells\",\n      \"pmids\": [\"41791707\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not distinguish supercoiling vs decatenation contributions at initiation\", \"Single cell-line model\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How TOP1 PARylation, proteolytic processing, and cell-cycle signaling are integrated to select among the multiple competing TOP1cc repair pathways in human cells remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified model coordinating TDP1, ERCC1-XPF, MUS81, Apn2, and NHEJ choice\", \"PARylation target residues on human TOP1 unmapped\", \"Structural model of human TOP1cc engagement by each repair enzyme lacking\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140097\", \"supporting_discovery_ids\": [9, 13, 19]},\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [3, 8, 19]},\n      {\"term_id\": \"GO:0016853\", \"supporting_discovery_ids\": [9, 13]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [8]},\n      {\"term_id\": \"GO:0005654\", \"supporting_discovery_ids\": [8]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [3, 22]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-69306\", \"supporting_discovery_ids\": [2, 21]},\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [7, 11, 16, 17]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [3, 18, 22]},\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [3, 22]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"TDP1\", \"MUS81\", \"PARP1\", \"ERCC1\", \"XPF\", \"RPA\", \"BTBD1\", \"BTBD2\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"tie","faith_supported":8,"faith_total":8,"faith_pct":100.0}}