{"gene":"APEX2","run_date":"2026-06-09T22:02:43","timeline":{"discoveries":[{"year":2001,"finding":"Human APE2 protein localizes predominantly to the nucleus and partially to mitochondria; its N-terminal 15 amino acids function as a mitochondrial targeting sequence (MTS). APE2 contains a functional PCNA-binding motif in its C-terminal region, demonstrated by immunoprecipitation and in vitro pull-down assays, and co-localizes with PCNA in nuclear foci. Treatment with HAT medium containing deoxyuridine increased APE2-PCNA co-localization, suggesting a role in PCNA-dependent base excision repair.","method":"Subcellular fractionation, Western blot, electron microscopic immunocytochemistry, immunoprecipitation, in vitro pull-down, laser scanning immunofluorescence microscopy, GFP-fusion construct expression in HeLa cells","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP and in vitro pulldown for PCNA interaction, direct EM immunocytochemistry and fractionation for localization, multiple orthogonal methods in a single focused study","pmids":["11376153"],"is_preprint":false},{"year":2002,"finding":"Ape2 (human) shares homology with E. coli ExoIII but exhibits comparatively weak AP site-specific endonuclease and 3'-nuclease activities compared to Ape1. Substitutions in the hydrophobic active-site pocket of Ape1 (F266, W280, L282) dramatically reduce abasic incision potency, and introduction of an ExoIII-like pocket into Ape2 enhances its AP endonuclease function. Mutations at F266 and W280 of Ape1 increase 3'-5' exonuclease activity, indicating this pocket governs substrate specificity across the enzyme family.","method":"Homology modeling, site-directed mutagenesis, in vitro endonuclease and exonuclease activity assays","journal":"Journal of molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro enzymatic assays with active-site mutagenesis, multiple mutants tested, single lab","pmids":["11866537"],"is_preprint":false},{"year":2004,"finding":"APEX2-null mice display growth retardation (~80% body size), dyshematopoiesis, and severe lymphopoiesis defects. Both thymocytes and mitogen-stimulated splenocytes from APEX2-null mice accumulate in G2/M phase, demonstrating that APEX2 is required for proper cell cycle progression of proliferating lymphocytes. APEX2 associates with PCNA and its expression peaks in late S phase.","method":"Gene knockout in mice (homologous recombination in ES cells), flow cytometry (cell cycle analysis), immunoprecipitation, Western blot, mRNA expression analysis","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean KO mouse with defined cellular phenotype (G2/M arrest), multiple cell types analyzed, replicated in lymphocyte populations","pmids":["15319281"],"is_preprint":false},{"year":2006,"finding":"Human Ape2 possesses strong 3'-5' exonuclease and 3'-phosphodiesterase activities and only weak AP endonuclease activity. Mutation of the active-site residue Asp277 to Ala inactivates all these activities. Ape2 preferentially acts on mismatched deoxyribonucleotides at the recessed 3'-termini of partial DNA duplexes, suggesting a role as a 3'-5' exonuclease involved in mismatch processing.","method":"In vitro biochemical assays (3'-5' exonuclease, 3'-phosphodiesterase, AP endonuclease), active-site mutagenesis (D277A)","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Moderate — reconstituted in vitro enzymatic assays with catalytic-residue mutagenesis, multiple substrate types tested, single lab","pmids":["16687656"],"is_preprint":false},{"year":2007,"finding":"Both APE1 and APE2 function in immunoglobulin class switch recombination (CSR) to generate the double-strand breaks (DSBs) necessary for CSR in vivo. APE2-deficient mice haploinsufficient for APE1 show reduced CSR and DSBs in switch-region DNA, demonstrating that abasic sites generated by UNG are converted to single-strand breaks by APEs as a step in DSB formation during CSR.","method":"Genetic epistasis using APE2-knockout and APE1-haploinsufficient mice, CSR assay (flow cytometry for Ig isotype switching), DSB measurement (ligation-mediated PCR)","journal":"The Journal of experimental medicine","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo genetic epistasis with isogenic mouse models, CSR and DSB assays providing mechanistic pathway placement","pmids":["18025127"],"is_preprint":false},{"year":2009,"finding":"PCNA strongly stimulates the 3'-5' exonuclease and 3'-phosphodiesterase activities of Ape2 but has no effect on its AP endonuclease activity. Upon hydrogen-peroxide treatment, Ape2 redistributes to nuclear foci and co-localizes with PCNA. Biochemically, Ape2 can remove 3'-adenine opposite 8-oxoG, suggesting PCNA-dependent participation in oxidative DNA damage repair.","method":"In vitro enzymatic assays with and without PCNA, fluorescence microscopy of H2O2-treated cells, co-localization analysis","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with PCNA stimulation of specific activities, paired with direct localization experiments in cells after oxidative stress, single lab with two orthogonal methods","pmids":["19443450"],"is_preprint":false},{"year":2009,"finding":"APE1 and APE2 convert abasic sites generated by UNG into single-strand breaks (SSBs) in immunoglobulin switch-region DNA during CSR. Mismatch repair is additionally required to convert distal SSBs into DSBs. DNA polymerase beta attempts to correctly repair APE-generated SSBs in switching B cells, but the high frequency of AID-instigated breaks results in net DSB and mutation generation. S region DSBs are introduced and resolved during G1 phase.","method":"Genetic epistasis (APE1/APE2-deficient B cells), ligation-mediated PCR for DSB detection, cell-cycle analysis, mutation frequency assays","journal":"Philosophical transactions of the Royal Society of London. Series B, Biological sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — genetic epistasis in primary B cells with DSB assays, single lab, partially overlapping with PMID 18025127","pmids":["19010771"],"is_preprint":false},{"year":2009,"finding":"Apex2 deficiency in mice causes a drastic reduction in somatic hypermutation (SHM) frequency and mutations per clone without affecting the pattern of base substitution, suggesting Apex2 promotes SHM through its 3'-5' exonuclease activity. Unexpectedly, CSR efficiency was not reduced in Apex2-deficient B cells, and Apex1 knockdown in CH12F3-2 cells also did not reduce CSR, indicating neither APE alone is required for CSR in this system.","method":"Apex2-deficient mouse model, SHM frequency quantification by sequencing, CSR assay, Apex1 shRNA knockdown in B lymphoma cells","journal":"International immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean KO mouse with sequencing-based SHM readout, plus complementary knockdown experiment; contradicts PMID 18025127 on CSR role","pmids":["19556307"],"is_preprint":false},{"year":2013,"finding":"APE2 is required for ATR-Chk1 checkpoint activation in response to oxidative stress (H2O2) in Xenopus egg extracts. APE2 is necessary for generation of RPA-bound single-stranded DNA, recruitment of the ATR-ATRIP-Rad9 checkpoint complex to damage sites, and Chk1 phosphorylation. The PCNA-interaction protein (PIP) box of APE2 is essential for its recruitment to H2O2-damaged chromatin. APE2's 3'-phosphodiesterase and 3'-5' exonuclease activities drive 3'-5' SSB end resection to generate ssDNA. APE2 directly associates with Chk1, and Ser86 in its Chk1-binding motif is essential for Chk1 phosphorylation, indicating a Claspin-like mediator role.","method":"Xenopus egg extract system, immunodepletion of APE2, Chk1 phosphorylation assay, RPA-ssDNA formation assay, checkpoint protein recruitment assay, co-immunoprecipitation (APE2-Chk1), domain/motif mutagenesis (PIP box mutant, S86A mutant)","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Strong — biochemical reconstitution in Xenopus extracts with immunodepletion, co-IP, and structure-function mutagenesis; multiple orthogonal methods in a single rigorous study","pmids":["23754435"],"is_preprint":false},{"year":2014,"finding":"APE2 (but not APE1) is highly expressed in germinal center B cells and contributes to somatic hypermutation (SHM) frequency, A:T mutations, insertions, and deletions. In the absence of both UNG and APE2, A:T mutations are dramatically reduced. APE2-generated SSBs serve as entry points for exonuclease recruited by MSH2-MSH6 mismatch repair proteins, which can recruit translesion polymerases to create mutations. APE1 is expressed at low levels in germinal center B cells and has little effect on SHM.","method":"APE2-deficient mice, APE1-haploinsufficient mice, SHM frequency and pattern analysis by sequencing, UNG/APE2 double-deficient mice, immunofluorescence for protein expression","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic epistasis with multiple mouse KO combinations, sequencing-based SHM analysis, mechanistic pathway placement via genetic interaction with UNG and MMR","pmids":["24927551"],"is_preprint":false},{"year":2016,"finding":"APE2's C-terminal Zf-GRF domain is a nucleic acid (particularly ssDNA)-binding module that regulates APE2's 3'-5' resection activity after oxidative DNA damage. X-ray crystallography of the Zf-GRF domain revealed a crescent-shaped ssDNA-binding claw flexibly appended to the EEP catalytic core. Structure-guided Zf-GRF mutations impair APE2 DNA binding and 3'-5' exonuclease processing, and prevent efficient APE2-dependent RPA recruitment to damaged chromatin and ATR-Chk1 DDR activation in Xenopus egg extracts.","method":"X-ray crystallography (Zf-GRF domain structure), NMR (nucleic acid binding), SAXS, site-directed mutagenesis of Zf-GRF, in vitro exonuclease activity assays, Xenopus egg extract checkpoint activation assays, RPA chromatin recruitment assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Strong — X-ray crystal structure combined with NMR, SAXS, mutagenesis, in vitro activity assays, and in vivo functional validation in Xenopus extracts; multiple orthogonal methods","pmids":["28028224"],"is_preprint":false},{"year":2018,"finding":"APE2 promotes ATR-Chk1 DDR signaling from a site-specific single-strand break (SSB). APE2 interacts with PCNA via its PIP box, and a novel mode of APE2-PCNA interaction was identified via the APE2 Zf-GRF domain and PCNA C-terminus. The Zf-GRF-PCNA interaction facilitates 3'-5' SSB end resection, checkpoint protein complex assembly (ATR, ATRIP, TopBP1, Rad9, Claspin), and SSB-induced ATR-Chk1 signaling. SSB-induced ATR DDR is also essential for SSB repair.","method":"Xenopus HSS (high-speed supernatant) system with plasmid-based site-specific SSB, immunodepletion, co-immunoprecipitation, in vitro pull-down assays (Zf-GRF/PCNA), checkpoint activation assay (Chk1 phosphorylation)","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Moderate — biochemical reconstitution in Xenopus extracts, co-IP and pull-down for protein interactions, multiple domain mutants tested, single lab","pmids":["29361157"],"is_preprint":false},{"year":2019,"finding":"BRCA2-deficient cells are synthetically lethal with APEX2 (APE2) loss. BRCA2-deficient cells specifically require the apurinic endonuclease activity and the PCNA-binding domain of APE2 (but not APE1) for viability, placing APE2 in a distinct repair pathway essential when HR is compromised.","method":"shRNA and CRISPR-based genetic screen in BRCA2-isogenic cell lines, domain function analysis (endonuclease-dead mutants, PCNA-binding mutant), cell viability assays","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome-wide genetic screen validated with isogenic pairs, specific domain mutants confirm mechanistic requirement, independently replicated with two library types","pmids":["30686591"],"is_preprint":false},{"year":2020,"finding":"The primary role of APE2 is to reverse blocked 3'-DNA ends (3'-blocking lesions) that preclude DNA synthesis. APE2 deficiency is synthetically lethal with BRCA1/BRCA2 loss because BRCA-deficient cells are exquisitely sensitive to 3'-blocking lesions. TOP1 processing of genomic ribonucleotides is identified as the main source of 3'-blocking lesions relevant to the APEX2-BRCA1/2 synthetic lethality. Structural and biochemical dissection defines APE2's activity on blocked 3' termini.","method":"Genetic interaction profiling (genome-wide screens), biochemical assays on 3'-blocked DNA substrates, structural analysis of APE2, genetic epistasis with TDP1, TOP1, and RNaseH2","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — structure-guided biochemical dissection combined with genome-wide genetic interaction profiling and epistasis analysis, multiple orthogonal methods, independent replication of synthetic lethality","pmids":["32516598"],"is_preprint":false},{"year":2020,"finding":"Cisplatin treatment upregulates APE2 in proximal tubule cells; APE2 binds to myosin heavy-chain 9 (MYH9) in mitochondria, leading to MYH9 dysfunction and mitochondrial fragmentation contributing to cisplatin-induced acute kidney injury (AKI). APE2-knockout mice are protected from cisplatin-induced AKI. APE2 transgenic mice recapitulate AKI pathophysiology in the absence of cisplatin.","method":"APE2 transgenic and knockout mouse models, cisplatin treatment, APE2 pulldown-mass spectrometry (identification of MYH9 as binding partner), co-immunoprecipitation (APE2-MYH9), histopathology, kidney function assays","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pulldown-MS and co-IP for APE2-MYH9 interaction, transgenic and KO mouse phenotypes, but mechanistic link between APE2-MYH9 binding and mitochondrial fragmentation is not fully reconstituted in vitro","pmids":["33288657"],"is_preprint":false},{"year":2021,"finding":"APE2 is a general regulator of the ATR-Chk1 DDR pathway in human pancreatic cancer cells in response to oxidative stress, DNA replication stress, and DNA double-strand breaks. APE2 knockdown enhances γH2AX foci and micronuclei formation. Celastrol was identified as an APE2 inhibitor that specifically blocks APE2 (but not APE1) binding to ssDNA and inhibits APE2 3'-5' exonuclease activity. Celastrol impairs ATR-Chk1 DDR in both Xenopus egg extracts and human pancreatic cancer cells.","method":"APE2 siRNA knockdown in pancreatic cancer cells, γH2AX foci and micronuclei assay (fluorescence microscopy), Xenopus egg extract DDR assay, in vitro ssDNA-binding and exonuclease activity assays with Celastrol, cell viability assays","journal":"Frontiers in cell and developmental biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KD with defined cellular phenotype, in vitro biochemical validation of inhibitor mechanism, two orthogonal systems (human cells + Xenopus), single lab","pmids":["34796173"],"is_preprint":false},{"year":2022,"finding":"X-ray crystal structures of yeast Apn2 (ortholog of human APE2) in complex with DNA reveal that Apn2 frays and cleaves 3'-DNA termini via a wedging mechanism that facilitates 1-6 nucleotide endonucleolytic cleavages. APN2 deletion or DNA-wedge mutant strains display mutator phenotypes, cell growth defects, and genotoxic stress sensitivity in a ribonucleotide excision repair (RER)-defective background, demonstrating that Apn2 processes Top1-generated complex DNA lesions at ribonucleotides via a wedge-and-cut mechanism.","method":"X-ray crystallography (Apn2-DNA complex structures), biochemical endonuclease/exonuclease assays, site-directed mutagenesis (DNA-wedge mutants), yeast genetic assays (mutation rate, growth, genotoxin sensitivity in RER-defective backgrounds)","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 1 / Strong — X-ray crystal structures of enzyme-DNA complexes combined with mutagenesis, biochemical assays, and in vivo genetic validation; multiple orthogonal methods in a single study","pmids":["36198268"],"is_preprint":false},{"year":2022,"finding":"APE2 (and its yeast ortholog Apn2) are clients of the Hsp70-Hsp90 chaperone axis. Apn2 physically interacts with Ssa1 (Hsp70) and Hsp82 (Hsp90) and the co-chaperone Ydj1 in yeast. Human APE2 also binds to Hsp70 and Hsp90 in mammalian cells. Pharmacological inhibition of Hsp70/Hsp90 leads to rapid loss of APE2 protein in cancer cell lines, demonstrating chaperone-dependent APE2 stability.","method":"Co-immunoprecipitation (Apn2-Ssa1, Apn2-Hsp82, APE2-Hsp70/Hsp90), small molecule chaperone inhibitor treatment, Western blot for protein stability across cancer cell lines","journal":"Biomolecules","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP in yeast and mammalian cells, pharmacological perturbation of chaperones with protein stability readout, conserved interaction validated across species; single lab","pmids":["35883419"],"is_preprint":false},{"year":2023,"finding":"APE2 is an effector of microhomology-mediated end joining (MMEJ). Loss of APE2 inhibits MMEJ at deprotected telomeres and at intra-chromosomal DSBs and is epistatic with Pol Theta (PolQ) for MMEJ activity. APE2 possesses intrinsic flap-cleaving nuclease activity, and its MMEJ function in cells depends on nuclease activity. An uncharacterized domain is required for APE2 recruitment to DSBs.","method":"MMEJ reporter assay, telomere deprotection assay, APE2 loss-of-function (knockout/knockdown), epistasis with Pol Theta, in vitro flap-cleavage assays, domain-deletion and nuclease-dead mutants, DSB recruitment assay","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro flap-cleavage assay combined with genetic epistasis (APE2-PolQ), multiple loss-of-function approaches, specific domain/activity mutants; rigorous mechanistic dissection","pmids":["37044098"],"is_preprint":false},{"year":2023,"finding":"In primary murine B cell cultures, APE2 promotes AID-dependent somatic hypermutation (SHM), while APE1 suppresses SHM. When GC-level APE1/APE2 expression is engineered (reduced APE1 genetically + overexpressed APE2), AID-dependent VDJH4 intron SHM becomes detectable. The C-terminus of APE2 that interacts with PCNA promotes both SHM and CSR, but the Zf-GRF domain (required for ATR-Chk1 interaction) is not required for SHM. APE1 downregulation in GC is required permissively for SHM.","method":"Primary murine B cell cultures, genetic manipulation (apex1+/- mice, APE2 overexpression), SHM frequency and pattern analysis by sequencing, domain/motif mutants (PCNA-binding mutant, Zf-GRF mutant)","journal":"Journal of immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic engineering in primary B cells with domain mutants and sequencing-based SHM readout; dissects domain requirements mechanistically, single lab","pmids":["37074207"],"is_preprint":false},{"year":2024,"finding":"APE2 protein abundance is regulated by ubiquitin-mediated proteasomal degradation. APE2 is poly-ubiquitinated via K48-linked chains, with K371 identified as the key ubiquitination site. MKRN3 was identified and validated as the E3 ubiquitin ligase responsible for APE2 ubiquitination both in cells and in vitro.","method":"Ubiquitination assay (K48-linkage-specific antibodies, proteasome inhibitor treatment), site-directed mutagenesis (K371R mutant), in vitro ubiquitination reconstitution assay with MKRN3, co-immunoprecipitation, Western blot for protein stability","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution of ubiquitination with MKRN3 as E3 ligase, plus mutagenesis (K371R) and in-cell ubiquitination assays; multiple orthogonal methods in a single study","pmids":["38705397"],"is_preprint":false},{"year":2025,"finding":"APE2 directly interacts with MYH9 in cochlear outer hair cells following cisplatin treatment. APE2 overexpression alone (using an inducible transgenic mouse model) is sufficient to cause high-frequency hearing loss with hair cell loss and stereocilia disorganization. Critical MYH9-binding domains of APE2 were mapped. APE2 depletion preserved mitochondrial metabolism and protected cochlear cells from cisplatin-induced apoptosis, and activated an ATR-p53 signaling axis promoting nuclear p53 localization.","method":"Inducible outer hair cell-specific APE2 transgenic mouse model, electron microscopy (stereocilia ultrastructure), co-immunoprecipitation (APE2-MYH9), domain mapping, APE2 knockdown in cochlear cells, mitochondrial metabolism assay, apoptosis assay, p53 localization by immunofluorescence","journal":"Cancer research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — transgenic mouse and co-IP with domain mapping, but mechanistic link between APE2-MYH9 interaction and mitochondrial/apoptotic effects not fully reconstituted in vitro; single lab","pmids":["40464565"],"is_preprint":false},{"year":2025,"finding":"Human APE2 (along with TREX2) can repair 3'-DNA-peptide cross-links (3'-histone-DPCs) derived from abasic (AP) sites. APE2 cleaves chemically synthesized adducts resembling proteolyzed Schiff base 3'-histone-DPCs, extending the known substrate repertoire of APE2 to include these complex 3'-blocking lesions.","method":"In vitro cleavage assays using chemically synthesized 3'-DPC substrates with purified human APE2 and TREX2","journal":"Chemical research in toxicology","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro reconstitution with chemically defined substrates, single lab, no mutagenesis or in vivo validation reported","pmids":["41257340"],"is_preprint":false}],"current_model":"APE2 (APEX2) is a multifunctional DNA repair nuclease with PCNA-dependent 3'-5' exonuclease, 3'-phosphodiesterase, and weak AP endonuclease activities; it localizes to both nucleus and mitochondria, is recruited to oxidative DNA damage sites via its PIP box, and uses its C-terminal Zf-GRF domain to bind ssDNA and drive 3'-5' SSB end resection that generates RPA-coated ssDNA, recruits the ATR-ATRIP-Rad9 complex, and directly mediates Chk1 phosphorylation (acting as a Claspin-like mediator); APE2 also participates in immunoglobulin diversification (CSR and SHM) by generating SSBs at AID-induced abasic sites, acts as an effector of microhomology-mediated end joining (MMEJ) via intrinsic flap-cleaving activity epistatic with Pol Theta, reverses 3'-blocking DNA lesions arising from TOP1 processing of genomic ribonucleotides (making it synthetically lethal with BRCA1/BRCA2 loss), interacts with MYH9 in mitochondria to contribute to cisplatin-induced organ toxicity, and is itself regulated by MKRN3-mediated K48-linked ubiquitination at K371 and by Hsp70-Hsp90 chaperone-dependent stabilization."},"narrative":{"mechanistic_narrative":"APE2 (APEX2) is a multifunctional DNA repair nuclease that processes 3'-blocking DNA lesions and links damage processing to checkpoint signaling [PMID:32516598, PMID:23754435]. Biochemically it carries strong 3'-5' exonuclease and 3'-phosphodiesterase activities together with only weak AP endonuclease activity, all dependent on the active-site residue Asp277, and it preferentially acts on mismatched or blocked recessed 3'-termini [PMID:16687656]; these activities are governed by an ExoIII-like active-site pocket that distinguishes APE2 from the more potent AP endonuclease APE1 [PMID:15319281]. APE2 docks onto PCNA through a C-terminal PIP box, which strongly stimulates its 3'-5' exonuclease and 3'-phosphodiesterase activities and recruits it to oxidative damage foci [PMID:11376153, PMID:19443450], while a second PCNA contact and a C-terminal Zf-GRF zinc-finger domain — a crescent-shaped ssDNA-binding claw appended to the catalytic core — drives 3'-5' single-strand-break end resection to generate RPA-coated ssDNA [PMID:28028224, PMID:29361157]. This resected ssDNA recruits the ATR-ATRIP-Rad9-TopBP1-Claspin complex, and APE2 directly binds Chk1 through a motif requiring Ser86, acting as a Claspin-like mediator of ATR-Chk1 checkpoint activation following oxidative stress, replication stress, and double-strand breaks [PMID:23754435, PMID:29361157, PMID:34796173]. The same end-processing biochemistry underlies a defined role in reversing 3'-blocking lesions that arise from TOP1 processing of genomic ribonucleotides, rendering APE2 synthetically lethal with BRCA1/BRCA2 loss [PMID:32516598, PMID:30686591], and APE2 additionally functions as an effector of microhomology-mediated end joining via intrinsic flap-cleaving activity epistatic with Pol Theta [PMID:37044098]. In adaptive immunity, APE2 converts AID/UNG-generated abasic sites into single-strand breaks to promote somatic hypermutation and immunoglobulin diversification, with its PCNA-binding C-terminus required for this activity [PMID:24927551, PMID:37074207, PMID:18025127]. APE2 is itself regulated by MKRN3-mediated K48-linked polyubiquitination at K371 and by Hsp70-Hsp90 chaperone-dependent protein stabilization [PMID:38705397, PMID:35883419], and a mitochondrial pool that binds MYH9 contributes to cisplatin-induced kidney and cochlear toxicity [PMID:33288657, PMID:40464565].","teleology":[{"year":2001,"claim":"Established where APE2 acts and how it is anchored, showing it is a dual nucleus/mitochondrion protein that physically engages PCNA in repair foci.","evidence":"Subcellular fractionation, EM immunocytochemistry, reciprocal Co-IP and in vitro pull-down with PCNA in HeLa cells","pmids":["11376153"],"confidence":"High","gaps":["Functional consequence of PCNA binding not yet defined","Mitochondrial role not characterized"]},{"year":2002,"claim":"Defined why APE2 is a weak AP endonuclease relative to APE1, attributing substrate specificity across the family to an active-site hydrophobic pocket.","evidence":"Homology modeling and active-site mutagenesis with in vitro endonuclease/exonuclease assays","pmids":["11866537"],"confidence":"High","gaps":["Did not establish APE2's preferred cellular substrate","No structure of APE2 itself"]},{"year":2006,"claim":"Reframed APE2 as primarily a 3'-5' exonuclease/3'-phosphodiesterase acting on recessed mismatched termini, identifying Asp277 as the catalytic residue for all activities.","evidence":"Reconstituted in vitro biochemical assays with D277A catalytic mutant and multiple substrates","pmids":["16687656"],"confidence":"High","gaps":["In vivo mismatch-processing role not demonstrated","Regulation of activity not addressed"]},{"year":2004,"claim":"Connected APE2 to organismal proliferation, showing it is required for lymphocyte cell-cycle progression and PCNA-associated late-S-phase function.","evidence":"APEX2-null mouse with flow-cytometric cell-cycle analysis and Co-IP","pmids":["15319281"],"confidence":"High","gaps":["Molecular cause of G2/M accumulation not resolved","Link between repair activity and proliferation defect unclear"]},{"year":2007,"claim":"Placed APE2 in immunoglobulin class switch recombination as an enzyme converting UNG-generated abasic sites to single-strand breaks during DSB formation.","evidence":"Genetic epistasis with APE1-haploinsufficient mice, CSR flow cytometry, LM-PCR DSB detection","pmids":["18025127"],"confidence":"High","gaps":["Relative APE1/APE2 contribution debated in later work","Which catalytic activity is used in vivo not isolated"]},{"year":2009,"claim":"Showed PCNA selectively stimulates APE2's 3'-5' exonuclease/phosphodiesterase activities (not AP endonuclease) and that APE2 redistributes to foci on oxidative stress, tying its enzymology to oxidative damage repair.","evidence":"In vitro assays +/- PCNA, fluorescence microscopy of H2O2-treated cells, 8-oxoG substrate processing","pmids":["19443450"],"confidence":"High","gaps":["Downstream pathway from foci recruitment not defined","Specificity of stimulation mechanism not structurally resolved"]},{"year":2009,"claim":"Clarified the CSR mechanism, showing APE-generated SSBs require MMR to become DSBs and that Pol beta competes to repair them in G1.","evidence":"Genetic epistasis in primary B cells with LM-PCR and cell-cycle/mutation assays","pmids":["19010771"],"confidence":"Medium","gaps":["Overlaps with prior CSR study","APE2-specific versus APE1 contribution not separated"]},{"year":2009,"claim":"Identified APE2 as the dominant APE driving somatic hypermutation via its 3'-5' exonuclease activity, while challenging a strict requirement for either APE in CSR.","evidence":"Apex2-deficient mouse with sequencing-based SHM, plus Apex1 knockdown in CH12F3-2 cells","pmids":["19556307"],"confidence":"Medium","gaps":["Contradicts earlier CSR findings","System-dependence of CSR result unresolved"]},{"year":2013,"claim":"Established APE2 as a direct mediator of ATR-Chk1 checkpoint signaling, defining the PIP-box-dependent recruitment, resection-driven RPA-ssDNA generation, and a Claspin-like Chk1-binding role requiring Ser86.","evidence":"Xenopus egg extract immunodepletion, Chk1 phosphorylation and RPA-ssDNA assays, Co-IP, PIP-box and S86A mutants","pmids":["23754435"],"confidence":"High","gaps":["Human cell validation pending at this stage","Structural basis of Chk1 binding undefined"]},{"year":2014,"claim":"Refined the SHM mechanism, showing germinal-center-enriched APE2 produces SSB entry points for MSH2-MSH6-recruited exonuclease and translesion polymerases generating A:T mutations.","evidence":"Multiple mouse KO combinations (APE2, UNG/APE2 double) with sequencing-based SHM analysis and immunofluorescence","pmids":["24927551"],"confidence":"High","gaps":["Direct biochemical handoff to MMR not reconstituted","Role of mismatch versus abasic substrate not separated"]},{"year":2016,"claim":"Provided the structural basis for ssDNA recognition, showing the Zf-GRF domain is a crescent-shaped ssDNA-binding claw that regulates 3'-5' resection and is required for RPA recruitment and ATR-Chk1 activation.","evidence":"X-ray crystallography, NMR, SAXS, Zf-GRF mutagenesis, in vitro exonuclease assays, Xenopus checkpoint assays","pmids":["28028224"],"confidence":"High","gaps":["Structure of full-length APE2 not solved","Coordination between catalytic core and Zf-GRF not visualized"]},{"year":2018,"claim":"Extended checkpoint signaling to a defined single-strand break and revealed a second Zf-GRF-PCNA interaction that organizes resection and checkpoint complex assembly.","evidence":"Xenopus HSS site-specific SSB system, immunodepletion, Co-IP, Zf-GRF/PCNA pull-downs, Chk1 phosphorylation","pmids":["29361157"],"confidence":"High","gaps":["Stoichiometry of dual PCNA contacts unresolved","Human cellular validation limited"]},{"year":2019,"claim":"Identified APE2 as a synthetic-lethal dependency of BRCA2-deficient cells, requiring its endonuclease activity and PCNA-binding domain, defining a distinct repair pathway needed when HR fails.","evidence":"shRNA/CRISPR genetic screens in BRCA2-isogenic cells with domain-mutant rescue and viability assays","pmids":["30686591"],"confidence":"High","gaps":["Nature of the lethal lesion not yet identified","Mechanism distinguishing APE2 from APE1 unclear at this stage"]},{"year":2020,"claim":"Defined the lesion driving BRCA synthetic lethality, showing APE2's core role is reversing 3'-blocking lesions from TOP1 processing of genomic ribonucleotides.","evidence":"Genome-wide genetic interaction profiling, biochemistry on 3'-blocked substrates, structural analysis, epistasis with TDP1/TOP1/RNaseH2","pmids":["32516598"],"confidence":"High","gaps":["Full-length human APE2-DNA structure not resolved","Other sources of 3'-blocking lesions not enumerated"]},{"year":2022,"claim":"Visualized the catalytic mechanism using the yeast ortholog Apn2, showing a DNA-fraying wedge-and-cut mechanism for processing Top1-generated ribonucleotide lesions.","evidence":"X-ray Apn2-DNA complex structures, DNA-wedge mutants, biochemical assays, yeast genetics in RER-defective backgrounds","pmids":["36198268"],"confidence":"High","gaps":["Direct human APE2-DNA complex not solved","Conservation of wedge mechanism in human enzyme assumed from orthology"]},{"year":2022,"claim":"Identified post-translational control of APE2 abundance through the Hsp70-Hsp90 chaperone axis, conserved from yeast to human.","evidence":"Co-IP (Apn2-Ssa1/Hsp82, APE2-Hsp70/Hsp90), chaperone inhibitor treatment with protein-stability Westerns","pmids":["35883419"],"confidence":"Medium","gaps":["Co-chaperone specificity in human cells not fully mapped","Effect of chaperone loss on repair function not measured"]},{"year":2023,"claim":"Established APE2 as an effector of microhomology-mediated end joining via intrinsic flap-cleaving activity epistatic with Pol Theta.","evidence":"MMEJ reporter and telomere deprotection assays, APE2-PolQ epistasis, in vitro flap cleavage, nuclease-dead and domain-deletion mutants","pmids":["37044098"],"confidence":"High","gaps":["The domain required for DSB recruitment is uncharacterized","Order of APE2 versus PolQ action not resolved"]},{"year":2023,"claim":"Dissected domain requirements in SHM, showing the PCNA-binding C-terminus promotes SHM and CSR while the Zf-GRF (checkpoint) domain is dispensable, and that APE1 downregulation permissively enables SHM.","evidence":"Primary murine B cells with apex1+/- and APE2-overexpression genetics, domain mutants, sequencing-based SHM","pmids":["37074207"],"confidence":"Medium","gaps":["Engineered expression context may not reflect native germinal center","Mechanistic basis of APE1 suppression unclear"]},{"year":2024,"claim":"Defined how APE2 levels are negatively regulated, identifying MKRN3 as the E3 ligase mediating K48-linked polyubiquitination at K371 targeting APE2 for proteasomal degradation.","evidence":"K48-linkage ubiquitination assays, K371R mutant, in vitro reconstitution with MKRN3, Co-IP, stability Westerns","pmids":["38705397"],"confidence":"High","gaps":["Physiological signal triggering MKRN3-APE2 turnover unknown","Impact on repair/checkpoint outcomes not measured"]},{"year":2020,"claim":"Revealed a non-canonical mitochondrial pathology role, showing cisplatin-upregulated APE2 binds MYH9 to drive mitochondrial fragmentation and acute kidney injury.","evidence":"APE2 transgenic and KO mice, cisplatin treatment, pulldown-MS identifying MYH9, Co-IP, histopathology","pmids":["33288657"],"confidence":"Medium","gaps":["Link between APE2-MYH9 binding and mitochondrial fragmentation not reconstituted in vitro","Relationship to APE2's nuclease function unclear"]},{"year":2025,"claim":"Extended the APE2-MYH9 toxicity axis to cochlear hair cells, mapping MYH9-binding domains and linking APE2 to cisplatin-induced hearing loss and an ATR-p53 apoptotic response.","evidence":"Inducible hair-cell APE2 transgenic mice, EM, Co-IP with domain mapping, APE2 knockdown, mitochondrial/apoptosis/p53 assays","pmids":["40464565"],"confidence":"Medium","gaps":["Mechanistic link between MYH9 binding and mitochondrial/apoptotic effects not reconstituted","Single lab, in vivo overexpression model"]},{"year":2025,"claim":"Expanded the substrate repertoire, showing human APE2 (with TREX2) can cleave 3'-DNA-peptide cross-links derived from abasic sites.","evidence":"In vitro cleavage assays with chemically synthesized 3'-histone-DPC substrates and purified APE2/TREX2","pmids":["41257340"],"confidence":"Medium","gaps":["No mutagenesis or in vivo validation","Cellular relevance of 3'-DPC repair by APE2 not established"]},{"year":null,"claim":"The identity of the domain mediating APE2 recruitment to double-strand breaks, and how its multiple roles (checkpoint, MMEJ, ribonucleotide lesion repair, immunoglobulin diversification) are coordinated and toggled in vivo, remain unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["DSB-recruitment domain uncharacterized [#18]","No full-length human APE2-DNA structure","Integration/regulation of competing functions not defined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140097","term_label":"catalytic activity, acting on DNA","supporting_discovery_ids":[2,3,16,18,22]},{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[3,5,13]},{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[10]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[8,11]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[0,5]},{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[0,14]},{"term_id":"GO:0000228","term_label":"nuclear chromosome","supporting_discovery_ids":[8,11]}],"pathway":[{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[3,5,13,18]},{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[8,11,15]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[4,9,19]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[12,13,14,21]}],"complexes":[],"partners":["PCNA","CHEK1","MYH9","MKRN3","HSPA1A","HSP90AA1","TREX2"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9UBZ4","full_name":"DNA-(apurinic or apyrimidinic site) endonuclease 2","aliases":["AP endonuclease XTH2","APEX nuclease 2","APEX nuclease-like 2","Apurinic-apyrimidinic endonuclease 2","AP endonuclease 2"],"length_aa":518,"mass_kda":57.4,"function":"Functions as a weak apurinic/apyrimidinic (AP) endodeoxyribonuclease in the DNA base excision repair (BER) pathway of DNA lesions induced by oxidative and alkylating agents (PubMed:16687656). Initiates repair of AP sites in DNA by catalyzing hydrolytic incision of the phosphodiester backbone immediately adjacent to the damage, generating a single-strand break with 5'-deoxyribose phosphate and 3'-hydroxyl ends. Also displays double-stranded DNA 3'-5' exonuclease, 3'-phosphodiesterase activities (PubMed:16687656, PubMed:19443450, PubMed:32516598). Shows robust 3'-5' exonuclease activity on 3'-recessed heteroduplex DNA and is able to remove mismatched nucleotides preferentially (PubMed:16687656, PubMed:19443450). Also exhibits 3'-5' exonuclease activity on a single nucleotide gap containing heteroduplex DNA and on blunt-ended substrates (PubMed:16687656). Shows fairly strong 3'-phosphodiesterase activity involved in the removal of 3'-damaged termini formed in DNA by oxidative agents (PubMed:16687656, PubMed:19443450). In the nucleus functions in the PCNA-dependent BER pathway (PubMed:11376153). Plays a role in reversing blocked 3' DNA ends, problematic lesions that preclude DNA synthesis (PubMed:32516598). Required for somatic hypermutation (SHM) and DNA cleavage step of class switch recombination (CSR) of immunoglobulin genes (By similarity). Required for proper cell cycle progression during proliferation of peripheral lymphocytes (By similarity)","subcellular_location":"Nucleus; Cytoplasm; Mitochondrion","url":"https://www.uniprot.org/uniprotkb/Q9UBZ4/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/APEX2","classification":"Not Classified","n_dependent_lines":186,"n_total_lines":1208,"dependency_fraction":0.15397350993377484},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"COMMD4","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/APEX2","total_profiled":1310},"omim":[{"mim_id":"619684","title":"MITOCHONDRIA-LOCALIZED GLUTAMIC ACID-RICH PROTEIN; MGARP","url":"https://www.omim.org/entry/619684"},{"mim_id":"300773","title":"APEX NUCLEASE (APURINIC/APYRIMIDINIC ENDONUCLEASE) 2; APEX2","url":"https://www.omim.org/entry/300773"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"},{"location":"Nucleoli fibrillar center","reliability":"Supported"},{"location":"Vesicles","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/APEX2"},"hgnc":{"alias_symbol":["APEXL2","APE2","XTH2","ZGRF2"],"prev_symbol":[]},"alphafold":{"accession":"Q9UBZ4","domains":[{"cath_id":"3.60.10.10","chopping":"1-327","consensus_level":"high","plddt":91.743,"start":1,"end":327},{"cath_id":"-","chopping":"464-518","consensus_level":"medium","plddt":86.6264,"start":464,"end":518}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9UBZ4","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9UBZ4-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9UBZ4-F1-predicted_aligned_error_v6.png","plddt_mean":78.38},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=APEX2","jax_strain_url":"https://www.jax.org/strain/search?query=APEX2"},"sequence":{"accession":"Q9UBZ4","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9UBZ4.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9UBZ4/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9UBZ4"}},"corpus_meta":[{"pmid":"25419960","id":"PMC_25419960","title":"Directed evolution of APEX2 for electron microscopy and proximity labeling.","date":"2014","source":"Nature methods","url":"https://pubmed.ncbi.nlm.nih.gov/25419960","citation_count":1101,"is_preprint":false},{"pmid":"26866790","id":"PMC_26866790","title":"Spatially resolved proteomic mapping in living cells with the engineered peroxidase APEX2.","date":"2016","source":"Nature protocols","url":"https://pubmed.ncbi.nlm.nih.gov/26866790","citation_count":443,"is_preprint":false},{"pmid":"30686591","id":"PMC_30686591","title":"Genetic Screens Reveal FEN1 and APEX2 as BRCA2 Synthetic Lethal Targets.","date":"2019","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/30686591","citation_count":170,"is_preprint":false},{"pmid":"28796234","id":"PMC_28796234","title":"Electron microscopy using the genetically encoded APEX2 tag in cultured mammalian cells.","date":"2017","source":"Nature protocols","url":"https://pubmed.ncbi.nlm.nih.gov/28796234","citation_count":153,"is_preprint":false},{"pmid":"30848125","id":"PMC_30848125","title":"Directed Evolution of Split APEX2 Peroxidase.","date":"2019","source":"ACS chemical biology","url":"https://pubmed.ncbi.nlm.nih.gov/30848125","citation_count":134,"is_preprint":false},{"pmid":"11376153","id":"PMC_11376153","title":"Human APE2 protein is mostly localized in the nuclei and to some extent in the mitochondria, while nuclear APE2 is partly associated with proliferating cell nuclear antigen.","date":"2001","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/11376153","citation_count":131,"is_preprint":false},{"pmid":"18025127","id":"PMC_18025127","title":"APE1- and APE2-dependent DNA breaks in immunoglobulin class switch recombination.","date":"2007","source":"The Journal of experimental medicine","url":"https://pubmed.ncbi.nlm.nih.gov/18025127","citation_count":129,"is_preprint":false},{"pmid":"11866537","id":"PMC_11866537","title":"Determinants in nuclease specificity of Ape1 and Ape2, human homologues of Escherichia coli exonuclease III.","date":"2002","source":"Journal of molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/11866537","citation_count":110,"is_preprint":false},{"pmid":"16687656","id":"PMC_16687656","title":"Human Ape2 protein has a 3'-5' exonuclease activity that acts preferentially on mismatched base pairs.","date":"2006","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/16687656","citation_count":106,"is_preprint":false},{"pmid":"23754435","id":"PMC_23754435","title":"APE2 is required for ATR-Chk1 checkpoint activation in response to oxidative stress.","date":"2013","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/23754435","citation_count":93,"is_preprint":false},{"pmid":"32516598","id":"PMC_32516598","title":"Endogenous DNA 3' Blocks Are Vulnerabilities for BRCA1 and BRCA2 Deficiency and Are Reversed by the APE2 Nuclease.","date":"2020","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/32516598","citation_count":90,"is_preprint":false},{"pmid":"31240809","id":"PMC_31240809","title":"Expanding APEX2 Substrates for Proximity-Dependent Labeling of Nucleic Acids and Proteins in Living Cells.","date":"2019","source":"Angewandte Chemie (International ed. in English)","url":"https://pubmed.ncbi.nlm.nih.gov/31240809","citation_count":84,"is_preprint":false},{"pmid":"34381044","id":"PMC_34381044","title":"Cell-type and subcellular compartment-specific APEX2 proximity labeling reveals activity-dependent nuclear proteome dynamics in the striatum.","date":"2021","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/34381044","citation_count":69,"is_preprint":false},{"pmid":"19443450","id":"PMC_19443450","title":"Role of PCNA-dependent stimulation of 3'-phosphodiesterase and 3'-5' exonuclease activities of human Ape2 in repair of oxidative DNA damage.","date":"2009","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/19443450","citation_count":66,"is_preprint":false},{"pmid":"34550632","id":"PMC_34550632","title":"APEX2-based Proximity Labeling of Atox1 Identifies CRIP2 as a Nuclear Copper-binding Protein that Regulates Autophagy Activation.","date":"2021","source":"Angewandte Chemie (International ed. in English)","url":"https://pubmed.ncbi.nlm.nih.gov/34550632","citation_count":66,"is_preprint":false},{"pmid":"30110897","id":"PMC_30110897","title":"Single-Strand Break End Resection in Genome Integrity: Mechanism and Regulation by APE2.","date":"2018","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/30110897","citation_count":59,"is_preprint":false},{"pmid":"28028224","id":"PMC_28028224","title":"APE2 Zf-GRF facilitates 3'-5' resection of DNA damage following oxidative stress.","date":"2016","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/28028224","citation_count":54,"is_preprint":false},{"pmid":"27274088","id":"PMC_27274088","title":"Proximity-dependent biotin labelling in yeast using the engineered ascorbate peroxidase APEX2.","date":"2016","source":"The Biochemical journal","url":"https://pubmed.ncbi.nlm.nih.gov/27274088","citation_count":51,"is_preprint":false},{"pmid":"29361157","id":"PMC_29361157","title":"APE2 promotes DNA damage response pathway from a single-strand break.","date":"2018","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/29361157","citation_count":47,"is_preprint":false},{"pmid":"24927551","id":"PMC_24927551","title":"Differential expression of APE1 and APE2 in germinal centers promotes error-prone repair and A:T mutations during somatic hypermutation.","date":"2014","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/24927551","citation_count":46,"is_preprint":false},{"pmid":"30805613","id":"PMC_30805613","title":"Determination of local chromatin interactions using a combined CRISPR and peroxidase APEX2 system.","date":"2019","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/30805613","citation_count":46,"is_preprint":false},{"pmid":"28881650","id":"PMC_28881650","title":"APEX2-enhanced electron microscopy distinguishes sigma-1 receptor localization in the nucleoplasmic reticulum.","date":"2017","source":"Oncotarget","url":"https://pubmed.ncbi.nlm.nih.gov/28881650","citation_count":44,"is_preprint":false},{"pmid":"19010771","id":"PMC_19010771","title":"The roles of APE1, APE2, DNA polymerase beta and mismatch repair in creating S region DNA breaks during antibody class switch.","date":"2009","source":"Philosophical transactions of the Royal Society of London. Series B, Biological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/19010771","citation_count":42,"is_preprint":false},{"pmid":"15319281","id":"PMC_15319281","title":"Growth retardation and dyslymphopoiesis accompanied by G2/M arrest in APEX2-null mice.","date":"2004","source":"Blood","url":"https://pubmed.ncbi.nlm.nih.gov/15319281","citation_count":40,"is_preprint":false},{"pmid":"31519755","id":"PMC_31519755","title":"Proteomic mapping by rapamycin-dependent targeting of APEX2 identifies binding partners of VAPB at the inner nuclear membrane.","date":"2019","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/31519755","citation_count":38,"is_preprint":false},{"pmid":"33288657","id":"PMC_33288657","title":"Cisplatin-Mediated Upregulation of APE2 Binding to MYH9 Provokes Mitochondrial Fragmentation and Acute Kidney Injury.","date":"2020","source":"Cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/33288657","citation_count":35,"is_preprint":false},{"pmid":"28955036","id":"PMC_28955036","title":"Optimizing the fragment complementation of APEX2 for detection of specific protein-protein interactions in live cells.","date":"2017","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/28955036","citation_count":35,"is_preprint":false},{"pmid":"37044098","id":"PMC_37044098","title":"The APE2 nuclease is essential for DNA double-strand break repair by microhomology-mediated end joining.","date":"2023","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/37044098","citation_count":34,"is_preprint":false},{"pmid":"19556307","id":"PMC_19556307","title":"Apex2 is required for efficient somatic hypermutation but not for class switch recombination of immunoglobulin genes.","date":"2009","source":"International immunology","url":"https://pubmed.ncbi.nlm.nih.gov/19556307","citation_count":34,"is_preprint":false},{"pmid":"33111110","id":"PMC_33111110","title":"An Optimized Protocol for Proximity Biotinylation in Confluent Epithelial Cell Cultures Using the Peroxidase APEX2.","date":"2020","source":"STAR protocols","url":"https://pubmed.ncbi.nlm.nih.gov/33111110","citation_count":32,"is_preprint":false},{"pmid":"33299006","id":"PMC_33299006","title":"Wnt-inducible Lrp6-APEX2 interacting proteins identify ESCRT machinery and Trk-fused gene as components of the Wnt signaling pathway.","date":"2020","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/33299006","citation_count":30,"is_preprint":false},{"pmid":"33306092","id":"PMC_33306092","title":"An APEX2 proximity ligation method for mapping interactions with the nuclear lamina.","date":"2021","source":"The Journal of cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/33306092","citation_count":27,"is_preprint":false},{"pmid":"32111912","id":"PMC_32111912","title":"Genomic alterations and abnormal expression of APE2 in multiple cancers.","date":"2020","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/32111912","citation_count":25,"is_preprint":false},{"pmid":"12573260","id":"PMC_12573260","title":"Characterization of the genomic structure and expression of the mouse Apex2 gene.","date":"2003","source":"Genomics","url":"https://pubmed.ncbi.nlm.nih.gov/12573260","citation_count":25,"is_preprint":false},{"pmid":"31323700","id":"PMC_31323700","title":"APEX2-mediated proximity labeling resolves protein networks in Saccharomyces cerevisiae cells.","date":"2019","source":"The FEBS journal","url":"https://pubmed.ncbi.nlm.nih.gov/31323700","citation_count":21,"is_preprint":false},{"pmid":"38122865","id":"PMC_38122865","title":"Once-Daily Oral Berotralstat for Long-Term Prophylaxis of Hereditary Angioedema: The Open-Label Extension of the APeX-2 Randomized Trial.","date":"2023","source":"The journal of allergy and clinical immunology. In practice","url":"https://pubmed.ncbi.nlm.nih.gov/38122865","citation_count":21,"is_preprint":false},{"pmid":"33568455","id":"PMC_33568455","title":"APEX2 Proximity Proteomics Resolves Flagellum Subdomains and Identifies Flagellum Tip-Specific Proteins in Trypanosoma brucei.","date":"2021","source":"mSphere","url":"https://pubmed.ncbi.nlm.nih.gov/33568455","citation_count":21,"is_preprint":false},{"pmid":"29812912","id":"PMC_29812912","title":"Proximity Labeling by a Recombinant APEX2-FGF1 Fusion Protein Reveals Interaction of FGF1 with the Proteoglycans CD44 and CSPG4.","date":"2018","source":"Biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/29812912","citation_count":20,"is_preprint":false},{"pmid":"34083046","id":"PMC_34083046","title":"Function and molecular mechanisms of APE2 in genome and epigenome integrity.","date":"2020","source":"Mutation research. Reviews in mutation research","url":"https://pubmed.ncbi.nlm.nih.gov/34083046","citation_count":19,"is_preprint":false},{"pmid":"34555496","id":"PMC_34555496","title":"Defining Proximity Proteome of Histone Modifications by Antibody-mediated Protein A-APEX2 Labeling.","date":"2021","source":"Genomics, proteomics & bioinformatics","url":"https://pubmed.ncbi.nlm.nih.gov/34555496","citation_count":18,"is_preprint":false},{"pmid":"34796173","id":"PMC_34796173","title":"APE2 Is a General Regulator of the ATR-Chk1 DNA Damage Response Pathway to Maintain Genome Integrity in Pancreatic Cancer Cells.","date":"2021","source":"Frontiers in cell and developmental biology","url":"https://pubmed.ncbi.nlm.nih.gov/34796173","citation_count":16,"is_preprint":false},{"pmid":"31306516","id":"PMC_31306516","title":"The cysteine-free single mutant C32S of APEX2 is a highly expressed and active fusion tag for proximity labeling applications.","date":"2019","source":"Protein science : a publication of the Protein Society","url":"https://pubmed.ncbi.nlm.nih.gov/31306516","citation_count":14,"is_preprint":false},{"pmid":"37967559","id":"PMC_37967559","title":"Proteomics from compartment-specific APEX2 labeling in Mycobacterium tuberculosis reveals Type VII secretion substrates in the cell wall.","date":"2023","source":"Cell chemical biology","url":"https://pubmed.ncbi.nlm.nih.gov/37967559","citation_count":14,"is_preprint":false},{"pmid":"31010013","id":"PMC_31010013","title":"Periplasmic Nanobody-APEX2 Fusions Enable Facile Visualization of Ebola, Marburg, and Mĕnglà virus Nucleoproteins, Alluding to Similar Antigenic Landscapes among Marburgvirus and Dianlovirus.","date":"2019","source":"Viruses","url":"https://pubmed.ncbi.nlm.nih.gov/31010013","citation_count":14,"is_preprint":false},{"pmid":"26063640","id":"PMC_26063640","title":"An enhanced ascorbate peroxidase 2/antibody-binding domain fusion protein (APEX2-ABD) as a recombinant target-specific signal amplifier.","date":"2015","source":"Chemical communications (Cambridge, England)","url":"https://pubmed.ncbi.nlm.nih.gov/26063640","citation_count":14,"is_preprint":false},{"pmid":"32775374","id":"PMC_32775374","title":"Identification of APEX2 as an oncogene in liver cancer.","date":"2020","source":"World journal of clinical cases","url":"https://pubmed.ncbi.nlm.nih.gov/32775374","citation_count":12,"is_preprint":false},{"pmid":"34302888","id":"PMC_34302888","title":"Apurinic/Apyrimidinic Endonuclease 2 (APE2): An ancillary enzyme for contextual base excision repair mechanisms to preserve genome stability.","date":"2021","source":"Biochimie","url":"https://pubmed.ncbi.nlm.nih.gov/34302888","citation_count":11,"is_preprint":false},{"pmid":"33757879","id":"PMC_33757879","title":"Identification of cellular proteins interacting with PEDV M protein through APEX2 labeling.","date":"2021","source":"Journal of proteomics","url":"https://pubmed.ncbi.nlm.nih.gov/33757879","citation_count":11,"is_preprint":false},{"pmid":"29238746","id":"PMC_29238746","title":"Determining the target protein localization in 3D using the combination of FIB-SEM and APEX2.","date":"2017","source":"Biophysics reports","url":"https://pubmed.ncbi.nlm.nih.gov/29238746","citation_count":11,"is_preprint":false},{"pmid":"3366666","id":"PMC_3366666","title":"Scanning microfluorometric measurement of cell constituents. Principles of the method and its application to the determination of NAD content and redox state of XTH-2 cells in culture.","date":"1988","source":"Histochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/3366666","citation_count":11,"is_preprint":false},{"pmid":"9270873","id":"PMC_9270873","title":"Ortho-vanadate affects both the tyrosination/detyrosination state of spindle microtubules and the organization of XTH-2 spindles.","date":"1997","source":"European journal of cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/9270873","citation_count":11,"is_preprint":false},{"pmid":"35331378","id":"PMC_35331378","title":"Optimized APEX2 peroxidase-mediated proximity labeling in fast- and slow-growing mycobacteria.","date":"2021","source":"Methods in enzymology","url":"https://pubmed.ncbi.nlm.nih.gov/35331378","citation_count":10,"is_preprint":false},{"pmid":"32138363","id":"PMC_32138363","title":"Probing the Environment of Emerin by Enhanced Ascorbate Peroxidase 2 (APEX2)-Mediated Proximity Labeling.","date":"2020","source":"Cells","url":"https://pubmed.ncbi.nlm.nih.gov/32138363","citation_count":10,"is_preprint":false},{"pmid":"31124088","id":"PMC_31124088","title":"Identification of Lipid Droplet Proteomes by Proximity Labeling Proteomics Using APEX2.","date":"2019","source":"Methods in molecular biology (Clifton, N.J.)","url":"https://pubmed.ncbi.nlm.nih.gov/31124088","citation_count":10,"is_preprint":false},{"pmid":"38336829","id":"PMC_38336829","title":"Using the heme peroxidase APEX2 to probe intracellular H2O2 flux and diffusion.","date":"2024","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/38336829","citation_count":9,"is_preprint":false},{"pmid":"36755963","id":"PMC_36755963","title":"APE2: catalytic function and synthetic lethality draw attention as a cancer therapy target.","date":"2023","source":"NAR cancer","url":"https://pubmed.ncbi.nlm.nih.gov/36755963","citation_count":8,"is_preprint":false},{"pmid":"32560054","id":"PMC_32560054","title":"The GABARAP Co-Secretome Identified by APEX2-GABARAP Proximity Labelling of Extracellular Vesicles.","date":"2020","source":"Cells","url":"https://pubmed.ncbi.nlm.nih.gov/32560054","citation_count":8,"is_preprint":false},{"pmid":"39718986","id":"PMC_39718986","title":"A subcellular selective APEX2-based proximity labeling used for identifying mitochondrial G-quadruplex DNA binding proteins.","date":"2025","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/39718986","citation_count":7,"is_preprint":false},{"pmid":"38755230","id":"PMC_38755230","title":"APEX2-based proximity proteomic analysis identifies candidate interactors for Plasmodium falciparum knob-associated histidine-rich protein in infected erythrocytes.","date":"2024","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/38755230","citation_count":7,"is_preprint":false},{"pmid":"36198268","id":"PMC_36198268","title":"Molecular basis for processing of topoisomerase 1-triggered DNA damage by Apn2/APE2.","date":"2022","source":"Cell reports","url":"https://pubmed.ncbi.nlm.nih.gov/36198268","citation_count":6,"is_preprint":false},{"pmid":"32024705","id":"PMC_32024705","title":"Three-Dimensional Visualization of APEX2-Tagged Erg11 in Saccharomyces cerevisiae Using Focused Ion Beam Scanning Electron Microscopy.","date":"2020","source":"mSphere","url":"https://pubmed.ncbi.nlm.nih.gov/32024705","citation_count":6,"is_preprint":false},{"pmid":"32548814","id":"PMC_32548814","title":"Selective Visualization of Caveolae by TEM Using APEX2.","date":"2020","source":"Methods in molecular biology (Clifton, N.J.)","url":"https://pubmed.ncbi.nlm.nih.gov/32548814","citation_count":6,"is_preprint":false},{"pmid":"30333092","id":"PMC_30333092","title":"Nano-structural analysis of engrafted human induced pluripotent stem cell-derived cardiomyocytes in mouse hearts using a genetic-probe APEX2.","date":"2018","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/30333092","citation_count":6,"is_preprint":false},{"pmid":"35563580","id":"PMC_35563580","title":"APEX2-Mediated Proximity Labeling Resolves the DDIT4-Interacting Proteome.","date":"2022","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/35563580","citation_count":5,"is_preprint":false},{"pmid":"35157287","id":"PMC_35157287","title":"Visualizing Filoviral Nucleoproteins Using Nanobodies Fused to the Ascorbate Peroxidase Derivatives APEX2 and dEAPX.","date":"2022","source":"Methods in molecular biology (Clifton, N.J.)","url":"https://pubmed.ncbi.nlm.nih.gov/35157287","citation_count":5,"is_preprint":false},{"pmid":"39512889","id":"PMC_39512889","title":"Detecting Native Protein-Protein Interactions by APEX2 Proximity Labeling in Drosophila Tissues.","date":"2024","source":"Bio-protocol","url":"https://pubmed.ncbi.nlm.nih.gov/39512889","citation_count":4,"is_preprint":false},{"pmid":"38099807","id":"PMC_38099807","title":"Clickable APEX2 Probes for Enhanced RNA Proximity Labeling in Live Cells.","date":"2023","source":"Analytical chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/38099807","citation_count":4,"is_preprint":false},{"pmid":"35883419","id":"PMC_35883419","title":"The APE2 Exonuclease Is a Client of the Hsp70-Hsp90 Axis in Yeast and Mammalian Cells.","date":"2022","source":"Biomolecules","url":"https://pubmed.ncbi.nlm.nih.gov/35883419","citation_count":4,"is_preprint":false},{"pmid":"30691651","id":"PMC_30691651","title":"Adapting dCas9-APEX2 for subnuclear proteomic profiling.","date":"2018","source":"Methods in enzymology","url":"https://pubmed.ncbi.nlm.nih.gov/30691651","citation_count":4,"is_preprint":false},{"pmid":"35733943","id":"PMC_35733943","title":"In vivo Proximity Labeling of Nuclear and Nucleolar Proteins by a Stably Expressed, DNA Damage-Responsive NONO-APEX2 Fusion Protein.","date":"2022","source":"Frontiers in molecular biosciences","url":"https://pubmed.ncbi.nlm.nih.gov/35733943","citation_count":4,"is_preprint":false},{"pmid":"37074207","id":"PMC_37074207","title":"APE2 Promotes AID-Dependent Somatic Hypermutation in Primary B Cell Cultures That Is Suppressed by APE1.","date":"2023","source":"Journal of immunology (Baltimore, Md. : 1950)","url":"https://pubmed.ncbi.nlm.nih.gov/37074207","citation_count":3,"is_preprint":false},{"pmid":"37260921","id":"PMC_37260921","title":"APEX2-Mediated Proximity Labeling of Wnt Receptor Interactors Upon Pathway Activation.","date":"2023","source":"microPublication biology","url":"https://pubmed.ncbi.nlm.nih.gov/37260921","citation_count":3,"is_preprint":false},{"pmid":"38958087","id":"PMC_38958087","title":"An APEX2-based proximity-dependent biotinylation assay with temporal specificity to study protein interactions during autophagy in the yeast Saccharomyces cerevisiae.","date":"2024","source":"Autophagy","url":"https://pubmed.ncbi.nlm.nih.gov/38958087","citation_count":3,"is_preprint":false},{"pmid":"37258967","id":"PMC_37258967","title":"Identification of Substrates of Secreted Bacterial Protease by APEX2-Based Proximity Labeling.","date":"2023","source":"Methods in molecular biology (Clifton, N.J.)","url":"https://pubmed.ncbi.nlm.nih.gov/37258967","citation_count":3,"is_preprint":false},{"pmid":"37961097","id":"PMC_37961097","title":"Ligand-Dependent Mechanisms of CC Chemokine Receptor 5 (CCR5) Trafficking Revealed by APEX2 Proximity Labeling Proteomics.","date":"2024","source":"bioRxiv : the preprint server for biology","url":"https://pubmed.ncbi.nlm.nih.gov/37961097","citation_count":3,"is_preprint":false},{"pmid":"39460941","id":"PMC_39460941","title":"Protocol to profile spatially resolved NLRP3 inflammasome complexes using APEX2-based proximity labeling.","date":"2024","source":"STAR protocols","url":"https://pubmed.ncbi.nlm.nih.gov/39460941","citation_count":3,"is_preprint":false},{"pmid":"38298178","id":"PMC_38298178","title":"Comparison of two peroxidases with high potential for biotechnology applications - HRP vs. APEX2.","date":"2024","source":"Computational and structural biotechnology journal","url":"https://pubmed.ncbi.nlm.nih.gov/38298178","citation_count":3,"is_preprint":false},{"pmid":"38705397","id":"PMC_38705397","title":"Ubiquitin-mediated regulation of APE2 protein abundance.","date":"2024","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/38705397","citation_count":2,"is_preprint":false},{"pmid":"40464565","id":"PMC_40464565","title":"Cisplatin-Induced APE2 Overexpression Disrupts MYH9 Function and Causes Hearing Loss.","date":"2025","source":"Cancer research communications","url":"https://pubmed.ncbi.nlm.nih.gov/40464565","citation_count":2,"is_preprint":false},{"pmid":"41118767","id":"PMC_41118767","title":"APEX2 proximity labeling of RNA in bacteria.","date":"2025","source":"Cell reports methods","url":"https://pubmed.ncbi.nlm.nih.gov/41118767","citation_count":2,"is_preprint":false},{"pmid":"37930538","id":"PMC_37930538","title":"Identification of Protein Partners by APEX2 Proximity Labeling.","date":"2024","source":"Methods in molecular biology (Clifton, N.J.)","url":"https://pubmed.ncbi.nlm.nih.gov/37930538","citation_count":2,"is_preprint":false},{"pmid":"36807792","id":"PMC_36807792","title":"Localization of Mitochondrial Nucleoids by Transmission Electron Microscopy Using the Transgenic Expression of the Mitochondrial Helicase Twinkle and APEX2.","date":"2023","source":"Methods in molecular biology (Clifton, N.J.)","url":"https://pubmed.ncbi.nlm.nih.gov/36807792","citation_count":2,"is_preprint":false},{"pmid":"37792491","id":"PMC_37792491","title":"Targeting APEX2 to the mRNA encoding fatty acid synthase β in yeast identifies interacting proteins that control its abundance in the cell cycle.","date":"2023","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/37792491","citation_count":2,"is_preprint":false},{"pmid":"35524122","id":"PMC_35524122","title":"Proteomic Mapping by APEX2-Catalyzed Proximity Labeling in Saccharomyces cerevisiae Semipermeabilized Cells.","date":"2022","source":"Methods in molecular biology (Clifton, N.J.)","url":"https://pubmed.ncbi.nlm.nih.gov/35524122","citation_count":2,"is_preprint":false},{"pmid":"41446759","id":"PMC_41446759","title":"Contrasting roles of APE1 and APE2 in genome maintenance, cancer development, and therapeutic targeting.","date":"2025","source":"NAR cancer","url":"https://pubmed.ncbi.nlm.nih.gov/41446759","citation_count":1,"is_preprint":false},{"pmid":"40694042","id":"PMC_40694042","title":"APEX2-based quantitative proteomics of LAT and CD3ζ interactomes in living human Jurkat T cells unveils new interactors.","date":"2025","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/40694042","citation_count":1,"is_preprint":false},{"pmid":"40638056","id":"PMC_40638056","title":"Antibody-Mediated Protein A-APEX2 Labeling (AMAPEX) for Proximity Proteome Exploration.","date":"2025","source":"Methods in molecular biology (Clifton, N.J.)","url":"https://pubmed.ncbi.nlm.nih.gov/40638056","citation_count":1,"is_preprint":false},{"pmid":"39392747","id":"PMC_39392747","title":"Protocols for identifying endogenous interactors of RNA-binding proteins in mammalian cells using the peroxidase APEX2 biotin-labeling method.","date":"2024","source":"STAR protocols","url":"https://pubmed.ncbi.nlm.nih.gov/39392747","citation_count":1,"is_preprint":false},{"pmid":"20949644","id":"PMC_20949644","title":"Molecular diagnosis of Down syndrome using quantitative APEX-2 microarrays.","date":"2010","source":"Prenatal diagnosis","url":"https://pubmed.ncbi.nlm.nih.gov/20949644","citation_count":1,"is_preprint":false},{"pmid":"10527549","id":"PMC_10527549","title":"Protein kinase C affects reformation of endothelial junctions in xenopus XTH-2 cells.","date":"1999","source":"Cell biology international","url":"https://pubmed.ncbi.nlm.nih.gov/10527549","citation_count":1,"is_preprint":false},{"pmid":"41372295","id":"PMC_41372295","title":"APEX2 and TurboID define unique subcellular proteomes.","date":"2025","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/41372295","citation_count":0,"is_preprint":false},{"pmid":"38954202","id":"PMC_38954202","title":"APEX2-Mediated Proximity Protein Labeling in Dictyostelium.","date":"2024","source":"Methods in molecular biology (Clifton, N.J.)","url":"https://pubmed.ncbi.nlm.nih.gov/38954202","citation_count":0,"is_preprint":false},{"pmid":"40655423","id":"PMC_40655423","title":"APEX2 RNA Proximity Labeling in Mammalian Cell Lines With Low Biotin Permeability.","date":"2025","source":"Bio-protocol","url":"https://pubmed.ncbi.nlm.nih.gov/40655423","citation_count":0,"is_preprint":false},{"pmid":"41257340","id":"PMC_41257340","title":"Human APE2 and TREX2 Repair 3'-DNA-Peptide Cross-links Derived from Abasic Sites.","date":"2025","source":"Chemical research in toxicology","url":"https://pubmed.ncbi.nlm.nih.gov/41257340","citation_count":0,"is_preprint":false},{"pmid":"39345536","id":"PMC_39345536","title":"APEX2 proximity labeling of RNA in bacteria.","date":"2025","source":"bioRxiv : the preprint server for biology","url":"https://pubmed.ncbi.nlm.nih.gov/39345536","citation_count":0,"is_preprint":false},{"pmid":"41131709","id":"PMC_41131709","title":"Comprehensive analysis of the role of ASC1 and APE2 introns on cellular fitness, transcription, and post-transcriptional dynamics.","date":"2025","source":"The FEBS journal","url":"https://pubmed.ncbi.nlm.nih.gov/41131709","citation_count":0,"is_preprint":false},{"pmid":"39940790","id":"PMC_39940790","title":"Identification of MCM2-Interacting Proteins Associated with Replication Initiation Using APEX2-Based Proximity Labeling Technology.","date":"2025","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/39940790","citation_count":0,"is_preprint":false},{"pmid":"41903118","id":"PMC_41903118","title":"Directed Evolution Improves the Catalytic Efficiency of APEX2-Mediated Proximity-Dependent RNA Labeling.","date":"2026","source":"Advanced science (Weinheim, Baden-Wurttemberg, Germany)","url":"https://pubmed.ncbi.nlm.nih.gov/41903118","citation_count":0,"is_preprint":false},{"pmid":"41769260","id":"PMC_41769260","title":"Identification of the Subcompartment-Specific Mitochondrial Proteome by APEX2 Proximity Labeling in Saccharomyces cerevisiae.","date":"2026","source":"Bio-protocol","url":"https://pubmed.ncbi.nlm.nih.gov/41769260","citation_count":0,"is_preprint":false},{"pmid":"29892021","id":"PMC_29892021","title":"Publisher Correction: Optimizing the fragment complementation of APEX2 for detection of specific protein-protein interactions in live cells.","date":"2018","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/29892021","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":43933,"output_tokens":7590,"usd":0.122825,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":16782,"output_tokens":6507,"usd":0.123292,"stage2_stop_reason":"end_turn"},"total_usd":0.246117,"stage1_batch_id":"msgbatch_0179Z2pxRYBsXi2xADneafRw","stage2_batch_id":"msgbatch_01LaY9ZaVdgjWArcZo9eqQE5","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2001,\n      \"finding\": \"Human APE2 protein localizes predominantly to the nucleus and partially to mitochondria; its N-terminal 15 amino acids function as a mitochondrial targeting sequence (MTS). APE2 contains a functional PCNA-binding motif in its C-terminal region, demonstrated by immunoprecipitation and in vitro pull-down assays, and co-localizes with PCNA in nuclear foci. Treatment with HAT medium containing deoxyuridine increased APE2-PCNA co-localization, suggesting a role in PCNA-dependent base excision repair.\",\n      \"method\": \"Subcellular fractionation, Western blot, electron microscopic immunocytochemistry, immunoprecipitation, in vitro pull-down, laser scanning immunofluorescence microscopy, GFP-fusion construct expression in HeLa cells\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP and in vitro pulldown for PCNA interaction, direct EM immunocytochemistry and fractionation for localization, multiple orthogonal methods in a single focused study\",\n      \"pmids\": [\"11376153\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Ape2 (human) shares homology with E. coli ExoIII but exhibits comparatively weak AP site-specific endonuclease and 3'-nuclease activities compared to Ape1. Substitutions in the hydrophobic active-site pocket of Ape1 (F266, W280, L282) dramatically reduce abasic incision potency, and introduction of an ExoIII-like pocket into Ape2 enhances its AP endonuclease function. Mutations at F266 and W280 of Ape1 increase 3'-5' exonuclease activity, indicating this pocket governs substrate specificity across the enzyme family.\",\n      \"method\": \"Homology modeling, site-directed mutagenesis, in vitro endonuclease and exonuclease activity assays\",\n      \"journal\": \"Journal of molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro enzymatic assays with active-site mutagenesis, multiple mutants tested, single lab\",\n      \"pmids\": [\"11866537\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"APEX2-null mice display growth retardation (~80% body size), dyshematopoiesis, and severe lymphopoiesis defects. Both thymocytes and mitogen-stimulated splenocytes from APEX2-null mice accumulate in G2/M phase, demonstrating that APEX2 is required for proper cell cycle progression of proliferating lymphocytes. APEX2 associates with PCNA and its expression peaks in late S phase.\",\n      \"method\": \"Gene knockout in mice (homologous recombination in ES cells), flow cytometry (cell cycle analysis), immunoprecipitation, Western blot, mRNA expression analysis\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean KO mouse with defined cellular phenotype (G2/M arrest), multiple cell types analyzed, replicated in lymphocyte populations\",\n      \"pmids\": [\"15319281\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Human Ape2 possesses strong 3'-5' exonuclease and 3'-phosphodiesterase activities and only weak AP endonuclease activity. Mutation of the active-site residue Asp277 to Ala inactivates all these activities. Ape2 preferentially acts on mismatched deoxyribonucleotides at the recessed 3'-termini of partial DNA duplexes, suggesting a role as a 3'-5' exonuclease involved in mismatch processing.\",\n      \"method\": \"In vitro biochemical assays (3'-5' exonuclease, 3'-phosphodiesterase, AP endonuclease), active-site mutagenesis (D277A)\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — reconstituted in vitro enzymatic assays with catalytic-residue mutagenesis, multiple substrate types tested, single lab\",\n      \"pmids\": [\"16687656\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Both APE1 and APE2 function in immunoglobulin class switch recombination (CSR) to generate the double-strand breaks (DSBs) necessary for CSR in vivo. APE2-deficient mice haploinsufficient for APE1 show reduced CSR and DSBs in switch-region DNA, demonstrating that abasic sites generated by UNG are converted to single-strand breaks by APEs as a step in DSB formation during CSR.\",\n      \"method\": \"Genetic epistasis using APE2-knockout and APE1-haploinsufficient mice, CSR assay (flow cytometry for Ig isotype switching), DSB measurement (ligation-mediated PCR)\",\n      \"journal\": \"The Journal of experimental medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo genetic epistasis with isogenic mouse models, CSR and DSB assays providing mechanistic pathway placement\",\n      \"pmids\": [\"18025127\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"PCNA strongly stimulates the 3'-5' exonuclease and 3'-phosphodiesterase activities of Ape2 but has no effect on its AP endonuclease activity. Upon hydrogen-peroxide treatment, Ape2 redistributes to nuclear foci and co-localizes with PCNA. Biochemically, Ape2 can remove 3'-adenine opposite 8-oxoG, suggesting PCNA-dependent participation in oxidative DNA damage repair.\",\n      \"method\": \"In vitro enzymatic assays with and without PCNA, fluorescence microscopy of H2O2-treated cells, co-localization analysis\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with PCNA stimulation of specific activities, paired with direct localization experiments in cells after oxidative stress, single lab with two orthogonal methods\",\n      \"pmids\": [\"19443450\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"APE1 and APE2 convert abasic sites generated by UNG into single-strand breaks (SSBs) in immunoglobulin switch-region DNA during CSR. Mismatch repair is additionally required to convert distal SSBs into DSBs. DNA polymerase beta attempts to correctly repair APE-generated SSBs in switching B cells, but the high frequency of AID-instigated breaks results in net DSB and mutation generation. S region DSBs are introduced and resolved during G1 phase.\",\n      \"method\": \"Genetic epistasis (APE1/APE2-deficient B cells), ligation-mediated PCR for DSB detection, cell-cycle analysis, mutation frequency assays\",\n      \"journal\": \"Philosophical transactions of the Royal Society of London. Series B, Biological sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — genetic epistasis in primary B cells with DSB assays, single lab, partially overlapping with PMID 18025127\",\n      \"pmids\": [\"19010771\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Apex2 deficiency in mice causes a drastic reduction in somatic hypermutation (SHM) frequency and mutations per clone without affecting the pattern of base substitution, suggesting Apex2 promotes SHM through its 3'-5' exonuclease activity. Unexpectedly, CSR efficiency was not reduced in Apex2-deficient B cells, and Apex1 knockdown in CH12F3-2 cells also did not reduce CSR, indicating neither APE alone is required for CSR in this system.\",\n      \"method\": \"Apex2-deficient mouse model, SHM frequency quantification by sequencing, CSR assay, Apex1 shRNA knockdown in B lymphoma cells\",\n      \"journal\": \"International immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean KO mouse with sequencing-based SHM readout, plus complementary knockdown experiment; contradicts PMID 18025127 on CSR role\",\n      \"pmids\": [\"19556307\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"APE2 is required for ATR-Chk1 checkpoint activation in response to oxidative stress (H2O2) in Xenopus egg extracts. APE2 is necessary for generation of RPA-bound single-stranded DNA, recruitment of the ATR-ATRIP-Rad9 checkpoint complex to damage sites, and Chk1 phosphorylation. The PCNA-interaction protein (PIP) box of APE2 is essential for its recruitment to H2O2-damaged chromatin. APE2's 3'-phosphodiesterase and 3'-5' exonuclease activities drive 3'-5' SSB end resection to generate ssDNA. APE2 directly associates with Chk1, and Ser86 in its Chk1-binding motif is essential for Chk1 phosphorylation, indicating a Claspin-like mediator role.\",\n      \"method\": \"Xenopus egg extract system, immunodepletion of APE2, Chk1 phosphorylation assay, RPA-ssDNA formation assay, checkpoint protein recruitment assay, co-immunoprecipitation (APE2-Chk1), domain/motif mutagenesis (PIP box mutant, S86A mutant)\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — biochemical reconstitution in Xenopus extracts with immunodepletion, co-IP, and structure-function mutagenesis; multiple orthogonal methods in a single rigorous study\",\n      \"pmids\": [\"23754435\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"APE2 (but not APE1) is highly expressed in germinal center B cells and contributes to somatic hypermutation (SHM) frequency, A:T mutations, insertions, and deletions. In the absence of both UNG and APE2, A:T mutations are dramatically reduced. APE2-generated SSBs serve as entry points for exonuclease recruited by MSH2-MSH6 mismatch repair proteins, which can recruit translesion polymerases to create mutations. APE1 is expressed at low levels in germinal center B cells and has little effect on SHM.\",\n      \"method\": \"APE2-deficient mice, APE1-haploinsufficient mice, SHM frequency and pattern analysis by sequencing, UNG/APE2 double-deficient mice, immunofluorescence for protein expression\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic epistasis with multiple mouse KO combinations, sequencing-based SHM analysis, mechanistic pathway placement via genetic interaction with UNG and MMR\",\n      \"pmids\": [\"24927551\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"APE2's C-terminal Zf-GRF domain is a nucleic acid (particularly ssDNA)-binding module that regulates APE2's 3'-5' resection activity after oxidative DNA damage. X-ray crystallography of the Zf-GRF domain revealed a crescent-shaped ssDNA-binding claw flexibly appended to the EEP catalytic core. Structure-guided Zf-GRF mutations impair APE2 DNA binding and 3'-5' exonuclease processing, and prevent efficient APE2-dependent RPA recruitment to damaged chromatin and ATR-Chk1 DDR activation in Xenopus egg extracts.\",\n      \"method\": \"X-ray crystallography (Zf-GRF domain structure), NMR (nucleic acid binding), SAXS, site-directed mutagenesis of Zf-GRF, in vitro exonuclease activity assays, Xenopus egg extract checkpoint activation assays, RPA chromatin recruitment 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 — X-ray crystal structure combined with NMR, SAXS, mutagenesis, in vitro activity assays, and in vivo functional validation in Xenopus extracts; multiple orthogonal methods\",\n      \"pmids\": [\"28028224\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"APE2 promotes ATR-Chk1 DDR signaling from a site-specific single-strand break (SSB). APE2 interacts with PCNA via its PIP box, and a novel mode of APE2-PCNA interaction was identified via the APE2 Zf-GRF domain and PCNA C-terminus. The Zf-GRF-PCNA interaction facilitates 3'-5' SSB end resection, checkpoint protein complex assembly (ATR, ATRIP, TopBP1, Rad9, Claspin), and SSB-induced ATR-Chk1 signaling. SSB-induced ATR DDR is also essential for SSB repair.\",\n      \"method\": \"Xenopus HSS (high-speed supernatant) system with plasmid-based site-specific SSB, immunodepletion, co-immunoprecipitation, in vitro pull-down assays (Zf-GRF/PCNA), checkpoint activation assay (Chk1 phosphorylation)\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — biochemical reconstitution in Xenopus extracts, co-IP and pull-down for protein interactions, multiple domain mutants tested, single lab\",\n      \"pmids\": [\"29361157\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"BRCA2-deficient cells are synthetically lethal with APEX2 (APE2) loss. BRCA2-deficient cells specifically require the apurinic endonuclease activity and the PCNA-binding domain of APE2 (but not APE1) for viability, placing APE2 in a distinct repair pathway essential when HR is compromised.\",\n      \"method\": \"shRNA and CRISPR-based genetic screen in BRCA2-isogenic cell lines, domain function analysis (endonuclease-dead mutants, PCNA-binding mutant), cell viability assays\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome-wide genetic screen validated with isogenic pairs, specific domain mutants confirm mechanistic requirement, independently replicated with two library types\",\n      \"pmids\": [\"30686591\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"The primary role of APE2 is to reverse blocked 3'-DNA ends (3'-blocking lesions) that preclude DNA synthesis. APE2 deficiency is synthetically lethal with BRCA1/BRCA2 loss because BRCA-deficient cells are exquisitely sensitive to 3'-blocking lesions. TOP1 processing of genomic ribonucleotides is identified as the main source of 3'-blocking lesions relevant to the APEX2-BRCA1/2 synthetic lethality. Structural and biochemical dissection defines APE2's activity on blocked 3' termini.\",\n      \"method\": \"Genetic interaction profiling (genome-wide screens), biochemical assays on 3'-blocked DNA substrates, structural analysis of APE2, genetic epistasis with TDP1, TOP1, and RNaseH2\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — structure-guided biochemical dissection combined with genome-wide genetic interaction profiling and epistasis analysis, multiple orthogonal methods, independent replication of synthetic lethality\",\n      \"pmids\": [\"32516598\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Cisplatin treatment upregulates APE2 in proximal tubule cells; APE2 binds to myosin heavy-chain 9 (MYH9) in mitochondria, leading to MYH9 dysfunction and mitochondrial fragmentation contributing to cisplatin-induced acute kidney injury (AKI). APE2-knockout mice are protected from cisplatin-induced AKI. APE2 transgenic mice recapitulate AKI pathophysiology in the absence of cisplatin.\",\n      \"method\": \"APE2 transgenic and knockout mouse models, cisplatin treatment, APE2 pulldown-mass spectrometry (identification of MYH9 as binding partner), co-immunoprecipitation (APE2-MYH9), histopathology, kidney function assays\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pulldown-MS and co-IP for APE2-MYH9 interaction, transgenic and KO mouse phenotypes, but mechanistic link between APE2-MYH9 binding and mitochondrial fragmentation is not fully reconstituted in vitro\",\n      \"pmids\": [\"33288657\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"APE2 is a general regulator of the ATR-Chk1 DDR pathway in human pancreatic cancer cells in response to oxidative stress, DNA replication stress, and DNA double-strand breaks. APE2 knockdown enhances γH2AX foci and micronuclei formation. Celastrol was identified as an APE2 inhibitor that specifically blocks APE2 (but not APE1) binding to ssDNA and inhibits APE2 3'-5' exonuclease activity. Celastrol impairs ATR-Chk1 DDR in both Xenopus egg extracts and human pancreatic cancer cells.\",\n      \"method\": \"APE2 siRNA knockdown in pancreatic cancer cells, γH2AX foci and micronuclei assay (fluorescence microscopy), Xenopus egg extract DDR assay, in vitro ssDNA-binding and exonuclease activity assays with Celastrol, cell viability assays\",\n      \"journal\": \"Frontiers in cell and developmental biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KD with defined cellular phenotype, in vitro biochemical validation of inhibitor mechanism, two orthogonal systems (human cells + Xenopus), single lab\",\n      \"pmids\": [\"34796173\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"X-ray crystal structures of yeast Apn2 (ortholog of human APE2) in complex with DNA reveal that Apn2 frays and cleaves 3'-DNA termini via a wedging mechanism that facilitates 1-6 nucleotide endonucleolytic cleavages. APN2 deletion or DNA-wedge mutant strains display mutator phenotypes, cell growth defects, and genotoxic stress sensitivity in a ribonucleotide excision repair (RER)-defective background, demonstrating that Apn2 processes Top1-generated complex DNA lesions at ribonucleotides via a wedge-and-cut mechanism.\",\n      \"method\": \"X-ray crystallography (Apn2-DNA complex structures), biochemical endonuclease/exonuclease assays, site-directed mutagenesis (DNA-wedge mutants), yeast genetic assays (mutation rate, growth, genotoxin sensitivity in RER-defective backgrounds)\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — X-ray crystal structures of enzyme-DNA complexes combined with mutagenesis, biochemical assays, and in vivo genetic validation; multiple orthogonal methods in a single study\",\n      \"pmids\": [\"36198268\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"APE2 (and its yeast ortholog Apn2) are clients of the Hsp70-Hsp90 chaperone axis. Apn2 physically interacts with Ssa1 (Hsp70) and Hsp82 (Hsp90) and the co-chaperone Ydj1 in yeast. Human APE2 also binds to Hsp70 and Hsp90 in mammalian cells. Pharmacological inhibition of Hsp70/Hsp90 leads to rapid loss of APE2 protein in cancer cell lines, demonstrating chaperone-dependent APE2 stability.\",\n      \"method\": \"Co-immunoprecipitation (Apn2-Ssa1, Apn2-Hsp82, APE2-Hsp70/Hsp90), small molecule chaperone inhibitor treatment, Western blot for protein stability across cancer cell lines\",\n      \"journal\": \"Biomolecules\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP in yeast and mammalian cells, pharmacological perturbation of chaperones with protein stability readout, conserved interaction validated across species; single lab\",\n      \"pmids\": [\"35883419\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"APE2 is an effector of microhomology-mediated end joining (MMEJ). Loss of APE2 inhibits MMEJ at deprotected telomeres and at intra-chromosomal DSBs and is epistatic with Pol Theta (PolQ) for MMEJ activity. APE2 possesses intrinsic flap-cleaving nuclease activity, and its MMEJ function in cells depends on nuclease activity. An uncharacterized domain is required for APE2 recruitment to DSBs.\",\n      \"method\": \"MMEJ reporter assay, telomere deprotection assay, APE2 loss-of-function (knockout/knockdown), epistasis with Pol Theta, in vitro flap-cleavage assays, domain-deletion and nuclease-dead mutants, DSB recruitment assay\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro flap-cleavage assay combined with genetic epistasis (APE2-PolQ), multiple loss-of-function approaches, specific domain/activity mutants; rigorous mechanistic dissection\",\n      \"pmids\": [\"37044098\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In primary murine B cell cultures, APE2 promotes AID-dependent somatic hypermutation (SHM), while APE1 suppresses SHM. When GC-level APE1/APE2 expression is engineered (reduced APE1 genetically + overexpressed APE2), AID-dependent VDJH4 intron SHM becomes detectable. The C-terminus of APE2 that interacts with PCNA promotes both SHM and CSR, but the Zf-GRF domain (required for ATR-Chk1 interaction) is not required for SHM. APE1 downregulation in GC is required permissively for SHM.\",\n      \"method\": \"Primary murine B cell cultures, genetic manipulation (apex1+/- mice, APE2 overexpression), SHM frequency and pattern analysis by sequencing, domain/motif mutants (PCNA-binding mutant, Zf-GRF mutant)\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic engineering in primary B cells with domain mutants and sequencing-based SHM readout; dissects domain requirements mechanistically, single lab\",\n      \"pmids\": [\"37074207\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"APE2 protein abundance is regulated by ubiquitin-mediated proteasomal degradation. APE2 is poly-ubiquitinated via K48-linked chains, with K371 identified as the key ubiquitination site. MKRN3 was identified and validated as the E3 ubiquitin ligase responsible for APE2 ubiquitination both in cells and in vitro.\",\n      \"method\": \"Ubiquitination assay (K48-linkage-specific antibodies, proteasome inhibitor treatment), site-directed mutagenesis (K371R mutant), in vitro ubiquitination reconstitution assay with MKRN3, co-immunoprecipitation, Western blot for protein stability\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution of ubiquitination with MKRN3 as E3 ligase, plus mutagenesis (K371R) and in-cell ubiquitination assays; multiple orthogonal methods in a single study\",\n      \"pmids\": [\"38705397\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"APE2 directly interacts with MYH9 in cochlear outer hair cells following cisplatin treatment. APE2 overexpression alone (using an inducible transgenic mouse model) is sufficient to cause high-frequency hearing loss with hair cell loss and stereocilia disorganization. Critical MYH9-binding domains of APE2 were mapped. APE2 depletion preserved mitochondrial metabolism and protected cochlear cells from cisplatin-induced apoptosis, and activated an ATR-p53 signaling axis promoting nuclear p53 localization.\",\n      \"method\": \"Inducible outer hair cell-specific APE2 transgenic mouse model, electron microscopy (stereocilia ultrastructure), co-immunoprecipitation (APE2-MYH9), domain mapping, APE2 knockdown in cochlear cells, mitochondrial metabolism assay, apoptosis assay, p53 localization by immunofluorescence\",\n      \"journal\": \"Cancer research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — transgenic mouse and co-IP with domain mapping, but mechanistic link between APE2-MYH9 interaction and mitochondrial/apoptotic effects not fully reconstituted in vitro; single lab\",\n      \"pmids\": [\"40464565\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Human APE2 (along with TREX2) can repair 3'-DNA-peptide cross-links (3'-histone-DPCs) derived from abasic (AP) sites. APE2 cleaves chemically synthesized adducts resembling proteolyzed Schiff base 3'-histone-DPCs, extending the known substrate repertoire of APE2 to include these complex 3'-blocking lesions.\",\n      \"method\": \"In vitro cleavage assays using chemically synthesized 3'-DPC substrates with purified human APE2 and TREX2\",\n      \"journal\": \"Chemical research in toxicology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro reconstitution with chemically defined substrates, single lab, no mutagenesis or in vivo validation reported\",\n      \"pmids\": [\"41257340\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"APE2 (APEX2) is a multifunctional DNA repair nuclease with PCNA-dependent 3'-5' exonuclease, 3'-phosphodiesterase, and weak AP endonuclease activities; it localizes to both nucleus and mitochondria, is recruited to oxidative DNA damage sites via its PIP box, and uses its C-terminal Zf-GRF domain to bind ssDNA and drive 3'-5' SSB end resection that generates RPA-coated ssDNA, recruits the ATR-ATRIP-Rad9 complex, and directly mediates Chk1 phosphorylation (acting as a Claspin-like mediator); APE2 also participates in immunoglobulin diversification (CSR and SHM) by generating SSBs at AID-induced abasic sites, acts as an effector of microhomology-mediated end joining (MMEJ) via intrinsic flap-cleaving activity epistatic with Pol Theta, reverses 3'-blocking DNA lesions arising from TOP1 processing of genomic ribonucleotides (making it synthetically lethal with BRCA1/BRCA2 loss), interacts with MYH9 in mitochondria to contribute to cisplatin-induced organ toxicity, and is itself regulated by MKRN3-mediated K48-linked ubiquitination at K371 and by Hsp70-Hsp90 chaperone-dependent stabilization.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"APE2 (APEX2) is a multifunctional DNA repair nuclease that processes 3'-blocking DNA lesions and links damage processing to checkpoint signaling [#13, #8]. Biochemically it carries strong 3'-5' exonuclease and 3'-phosphodiesterase activities together with only weak AP endonuclease activity, all dependent on the active-site residue Asp277, and it preferentially acts on mismatched or blocked recessed 3'-termini [#3]; these activities are governed by an ExoIII-like active-site pocket that distinguishes APE2 from the more potent AP endonuclease APE1 [#2]. APE2 docks onto PCNA through a C-terminal PIP box, which strongly stimulates its 3'-5' exonuclease and 3'-phosphodiesterase activities and recruits it to oxidative damage foci [#0, #5], while a second PCNA contact and a C-terminal Zf-GRF zinc-finger domain — a crescent-shaped ssDNA-binding claw appended to the catalytic core — drives 3'-5' single-strand-break end resection to generate RPA-coated ssDNA [#10, #11]. This resected ssDNA recruits the ATR-ATRIP-Rad9-TopBP1-Claspin complex, and APE2 directly binds Chk1 through a motif requiring Ser86, acting as a Claspin-like mediator of ATR-Chk1 checkpoint activation following oxidative stress, replication stress, and double-strand breaks [#8, #11, #15]. The same end-processing biochemistry underlies a defined role in reversing 3'-blocking lesions that arise from TOP1 processing of genomic ribonucleotides, rendering APE2 synthetically lethal with BRCA1/BRCA2 loss [#13, #12], and APE2 additionally functions as an effector of microhomology-mediated end joining via intrinsic flap-cleaving activity epistatic with Pol Theta [#18]. In adaptive immunity, APE2 converts AID/UNG-generated abasic sites into single-strand breaks to promote somatic hypermutation and immunoglobulin diversification, with its PCNA-binding C-terminus required for this activity [#9, #19, #4]. APE2 is itself regulated by MKRN3-mediated K48-linked polyubiquitination at K371 and by Hsp70-Hsp90 chaperone-dependent protein stabilization [#20, #17], and a mitochondrial pool that binds MYH9 contributes to cisplatin-induced kidney and cochlear toxicity [#14, #21].\",\n  \"teleology\": [\n    {\n      \"year\": 2001,\n      \"claim\": \"Established where APE2 acts and how it is anchored, showing it is a dual nucleus/mitochondrion protein that physically engages PCNA in repair foci.\",\n      \"evidence\": \"Subcellular fractionation, EM immunocytochemistry, reciprocal Co-IP and in vitro pull-down with PCNA in HeLa cells\",\n      \"pmids\": [\"11376153\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional consequence of PCNA binding not yet defined\", \"Mitochondrial role not characterized\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Defined why APE2 is a weak AP endonuclease relative to APE1, attributing substrate specificity across the family to an active-site hydrophobic pocket.\",\n      \"evidence\": \"Homology modeling and active-site mutagenesis with in vitro endonuclease/exonuclease assays\",\n      \"pmids\": [\"11866537\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not establish APE2's preferred cellular substrate\", \"No structure of APE2 itself\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Reframed APE2 as primarily a 3'-5' exonuclease/3'-phosphodiesterase acting on recessed mismatched termini, identifying Asp277 as the catalytic residue for all activities.\",\n      \"evidence\": \"Reconstituted in vitro biochemical assays with D277A catalytic mutant and multiple substrates\",\n      \"pmids\": [\"16687656\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo mismatch-processing role not demonstrated\", \"Regulation of activity not addressed\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"Connected APE2 to organismal proliferation, showing it is required for lymphocyte cell-cycle progression and PCNA-associated late-S-phase function.\",\n      \"evidence\": \"APEX2-null mouse with flow-cytometric cell-cycle analysis and Co-IP\",\n      \"pmids\": [\"15319281\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular cause of G2/M accumulation not resolved\", \"Link between repair activity and proliferation defect unclear\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Placed APE2 in immunoglobulin class switch recombination as an enzyme converting UNG-generated abasic sites to single-strand breaks during DSB formation.\",\n      \"evidence\": \"Genetic epistasis with APE1-haploinsufficient mice, CSR flow cytometry, LM-PCR DSB detection\",\n      \"pmids\": [\"18025127\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Relative APE1/APE2 contribution debated in later work\", \"Which catalytic activity is used in vivo not isolated\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Showed PCNA selectively stimulates APE2's 3'-5' exonuclease/phosphodiesterase activities (not AP endonuclease) and that APE2 redistributes to foci on oxidative stress, tying its enzymology to oxidative damage repair.\",\n      \"evidence\": \"In vitro assays +/- PCNA, fluorescence microscopy of H2O2-treated cells, 8-oxoG substrate processing\",\n      \"pmids\": [\"19443450\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Downstream pathway from foci recruitment not defined\", \"Specificity of stimulation mechanism not structurally resolved\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Clarified the CSR mechanism, showing APE-generated SSBs require MMR to become DSBs and that Pol beta competes to repair them in G1.\",\n      \"evidence\": \"Genetic epistasis in primary B cells with LM-PCR and cell-cycle/mutation assays\",\n      \"pmids\": [\"19010771\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Overlaps with prior CSR study\", \"APE2-specific versus APE1 contribution not separated\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Identified APE2 as the dominant APE driving somatic hypermutation via its 3'-5' exonuclease activity, while challenging a strict requirement for either APE in CSR.\",\n      \"evidence\": \"Apex2-deficient mouse with sequencing-based SHM, plus Apex1 knockdown in CH12F3-2 cells\",\n      \"pmids\": [\"19556307\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Contradicts earlier CSR findings\", \"System-dependence of CSR result unresolved\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Established APE2 as a direct mediator of ATR-Chk1 checkpoint signaling, defining the PIP-box-dependent recruitment, resection-driven RPA-ssDNA generation, and a Claspin-like Chk1-binding role requiring Ser86.\",\n      \"evidence\": \"Xenopus egg extract immunodepletion, Chk1 phosphorylation and RPA-ssDNA assays, Co-IP, PIP-box and S86A mutants\",\n      \"pmids\": [\"23754435\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Human cell validation pending at this stage\", \"Structural basis of Chk1 binding undefined\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Refined the SHM mechanism, showing germinal-center-enriched APE2 produces SSB entry points for MSH2-MSH6-recruited exonuclease and translesion polymerases generating A:T mutations.\",\n      \"evidence\": \"Multiple mouse KO combinations (APE2, UNG/APE2 double) with sequencing-based SHM analysis and immunofluorescence\",\n      \"pmids\": [\"24927551\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct biochemical handoff to MMR not reconstituted\", \"Role of mismatch versus abasic substrate not separated\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Provided the structural basis for ssDNA recognition, showing the Zf-GRF domain is a crescent-shaped ssDNA-binding claw that regulates 3'-5' resection and is required for RPA recruitment and ATR-Chk1 activation.\",\n      \"evidence\": \"X-ray crystallography, NMR, SAXS, Zf-GRF mutagenesis, in vitro exonuclease assays, Xenopus checkpoint assays\",\n      \"pmids\": [\"28028224\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structure of full-length APE2 not solved\", \"Coordination between catalytic core and Zf-GRF not visualized\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Extended checkpoint signaling to a defined single-strand break and revealed a second Zf-GRF-PCNA interaction that organizes resection and checkpoint complex assembly.\",\n      \"evidence\": \"Xenopus HSS site-specific SSB system, immunodepletion, Co-IP, Zf-GRF/PCNA pull-downs, Chk1 phosphorylation\",\n      \"pmids\": [\"29361157\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Stoichiometry of dual PCNA contacts unresolved\", \"Human cellular validation limited\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Identified APE2 as a synthetic-lethal dependency of BRCA2-deficient cells, requiring its endonuclease activity and PCNA-binding domain, defining a distinct repair pathway needed when HR fails.\",\n      \"evidence\": \"shRNA/CRISPR genetic screens in BRCA2-isogenic cells with domain-mutant rescue and viability assays\",\n      \"pmids\": [\"30686591\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Nature of the lethal lesion not yet identified\", \"Mechanism distinguishing APE2 from APE1 unclear at this stage\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Defined the lesion driving BRCA synthetic lethality, showing APE2's core role is reversing 3'-blocking lesions from TOP1 processing of genomic ribonucleotides.\",\n      \"evidence\": \"Genome-wide genetic interaction profiling, biochemistry on 3'-blocked substrates, structural analysis, epistasis with TDP1/TOP1/RNaseH2\",\n      \"pmids\": [\"32516598\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full-length human APE2-DNA structure not resolved\", \"Other sources of 3'-blocking lesions not enumerated\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Visualized the catalytic mechanism using the yeast ortholog Apn2, showing a DNA-fraying wedge-and-cut mechanism for processing Top1-generated ribonucleotide lesions.\",\n      \"evidence\": \"X-ray Apn2-DNA complex structures, DNA-wedge mutants, biochemical assays, yeast genetics in RER-defective backgrounds\",\n      \"pmids\": [\"36198268\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct human APE2-DNA complex not solved\", \"Conservation of wedge mechanism in human enzyme assumed from orthology\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Identified post-translational control of APE2 abundance through the Hsp70-Hsp90 chaperone axis, conserved from yeast to human.\",\n      \"evidence\": \"Co-IP (Apn2-Ssa1/Hsp82, APE2-Hsp70/Hsp90), chaperone inhibitor treatment with protein-stability Westerns\",\n      \"pmids\": [\"35883419\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Co-chaperone specificity in human cells not fully mapped\", \"Effect of chaperone loss on repair function not measured\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Established APE2 as an effector of microhomology-mediated end joining via intrinsic flap-cleaving activity epistatic with Pol Theta.\",\n      \"evidence\": \"MMEJ reporter and telomere deprotection assays, APE2-PolQ epistasis, in vitro flap cleavage, nuclease-dead and domain-deletion mutants\",\n      \"pmids\": [\"37044098\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"The domain required for DSB recruitment is uncharacterized\", \"Order of APE2 versus PolQ action not resolved\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Dissected domain requirements in SHM, showing the PCNA-binding C-terminus promotes SHM and CSR while the Zf-GRF (checkpoint) domain is dispensable, and that APE1 downregulation permissively enables SHM.\",\n      \"evidence\": \"Primary murine B cells with apex1+/- and APE2-overexpression genetics, domain mutants, sequencing-based SHM\",\n      \"pmids\": [\"37074207\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Engineered expression context may not reflect native germinal center\", \"Mechanistic basis of APE1 suppression unclear\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defined how APE2 levels are negatively regulated, identifying MKRN3 as the E3 ligase mediating K48-linked polyubiquitination at K371 targeting APE2 for proteasomal degradation.\",\n      \"evidence\": \"K48-linkage ubiquitination assays, K371R mutant, in vitro reconstitution with MKRN3, Co-IP, stability Westerns\",\n      \"pmids\": [\"38705397\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological signal triggering MKRN3-APE2 turnover unknown\", \"Impact on repair/checkpoint outcomes not measured\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Revealed a non-canonical mitochondrial pathology role, showing cisplatin-upregulated APE2 binds MYH9 to drive mitochondrial fragmentation and acute kidney injury.\",\n      \"evidence\": \"APE2 transgenic and KO mice, cisplatin treatment, pulldown-MS identifying MYH9, Co-IP, histopathology\",\n      \"pmids\": [\"33288657\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Link between APE2-MYH9 binding and mitochondrial fragmentation not reconstituted in vitro\", \"Relationship to APE2's nuclease function unclear\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Extended the APE2-MYH9 toxicity axis to cochlear hair cells, mapping MYH9-binding domains and linking APE2 to cisplatin-induced hearing loss and an ATR-p53 apoptotic response.\",\n      \"evidence\": \"Inducible hair-cell APE2 transgenic mice, EM, Co-IP with domain mapping, APE2 knockdown, mitochondrial/apoptosis/p53 assays\",\n      \"pmids\": [\"40464565\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanistic link between MYH9 binding and mitochondrial/apoptotic effects not reconstituted\", \"Single lab, in vivo overexpression model\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Expanded the substrate repertoire, showing human APE2 (with TREX2) can cleave 3'-DNA-peptide cross-links derived from abasic sites.\",\n      \"evidence\": \"In vitro cleavage assays with chemically synthesized 3'-histone-DPC substrates and purified APE2/TREX2\",\n      \"pmids\": [\"41257340\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No mutagenesis or in vivo validation\", \"Cellular relevance of 3'-DPC repair by APE2 not established\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The identity of the domain mediating APE2 recruitment to double-strand breaks, and how its multiple roles (checkpoint, MMEJ, ribonucleotide lesion repair, immunoglobulin diversification) are coordinated and toggled in vivo, remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"DSB-recruitment domain uncharacterized [#18]\", \"No full-length human APE2-DNA structure\", \"Integration/regulation of competing functions not defined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140097\", \"supporting_discovery_ids\": [2, 3, 16, 18, 22]},\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [3, 5, 13]},\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [10]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [8, 11]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [0, 5]},\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [0, 14]},\n      {\"term_id\": \"GO:0000228\", \"supporting_discovery_ids\": [8, 11]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [3, 5, 13, 18]},\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [8, 11, 15]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [4, 9, 19]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [12, 13, 14, 21]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"PCNA\", \"CHEK1\", \"MYH9\", \"MKRN3\", \"HSPA1A\", \"HSP90AA1\", \"TREX2\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}