{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,2]],"date-time":"2026-10-02T23:52:03Z","timestamp":1790985123981,"version":"4.1.0"},"reference-count":52,"publisher":"MDPI AG","issue":"2","license":[{"start":{"date-parts":[[2026,1,16]],"date-time":"2026-01-16T00:00:00Z","timestamp":1768521600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Associazione Italiana per la Ricerca sul Cancro","award":["IG-21535"],"award-info":[{"award-number":["IG-21535"]}]},{"name":"Italian National Research Council"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Cells"],"abstract":"<jats:p>DNA double-strand breaks (DSBs) can be induced by cellular byproducts or genotoxic agents. Improper processing of these lesions leads to increased genome instability, which constitutes a hallmark of pathological conditions and fuels carcinogenesis. DSBs are primarily repaired by homologous recombination (HR) and non-homologous end joining (NHEJ) and the proper balance between these two pathways is finely modulated by specific molecular events. Here, we report that the histone chaperone DAXX plays a fundamental role in the response to DSBs. Indeed, in human cells, DSBs induce ATM\/ATR-dependent phosphorylation of DAXX on serine 424 and 712 and promote its binding to chromatin and the deposition of the histone variant H3.3 in proximity to DNA breaks. Enrichment of H3.3 at DSBs promotes 53BP1 recruitment to these lesions and the repair of DNA breaks by HR pathways. Moreover, H3.3-specific post translational modifications, particularly K36 tri-methylation, play a key role in these processes. Altogether, these findings indicate that DAXX and H3.3 mutations may contribute to tumorigenesis-enhancing genome instability.<\/jats:p>","DOI":"10.3390\/cells15020162","type":"journal-article","created":{"date-parts":[[2026,1,16]],"date-time":"2026-01-16T08:08:21Z","timestamp":1768550901000},"page":"162","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Daxx-Dependent H3.3 Deposition Promotes Double-Strand Breaks Repair by Homologous Recombination"],"prefix":"10.3390","volume":"15","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-8731-6507","authenticated-orcid":false,"given":"Laura","family":"Zannini","sequence":"first","affiliation":[{"name":"Istituto di Genetica Molecolare Luigi Luca Cavalli-Sforza, Consiglio Nazionale delle Ricerche (IGM-CNR), Via Abbiategrasso 207, 27100 Pavia, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Simona","family":"Aliprandi","sequence":"additional","affiliation":[{"name":"Dipartimento di Bioscienze, Universit\u00e0 degli Studi di Milano, Via Celoria 26, 20133 Milan, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Domenico","family":"Delia","sequence":"additional","affiliation":[{"name":"IFOM (Istituto FIRC di Oncologia Molecolare), Via Adamello 16, 20139 Milan, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Giacomo","family":"Buscemi","sequence":"additional","affiliation":[{"name":"Istituto di Genetica Molecolare Luigi Luca Cavalli-Sforza, Consiglio Nazionale delle Ricerche (IGM-CNR), Via Abbiategrasso 207, 27100 Pavia, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,1,16]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"179","DOI":"10.1016\/j.molcel.2010.09.019","article-title":"The DNA Damage Response: Making It Safe to Play with Knives","volume":"40","author":"Ciccia","year":"2010","journal-title":"Mol. Cell"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"251","DOI":"10.1038\/s12276-023-01145-3","article-title":"HIRA vs. DAXX: The Two Axes Shaping the Histone H3.3 Landscape","volume":"56","author":"Choi","year":"2024","journal-title":"Exp. Mol. Med."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"559","DOI":"10.1074\/jbc.M509266200","article-title":"Expression Patterns and Post-Translational Modifications Associated with Mammalian Histone H3 Variants","volume":"281","author":"Hake","year":"2006","journal-title":"J. Biol. Chem."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"309","DOI":"10.1016\/j.molcel.2006.08.019","article-title":"PTMs on H3 Variants before Chromatin Assembly Potentiate Their Final Epigenetic State","volume":"24","author":"Loyola","year":"2006","journal-title":"Mol. Cell"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"94","DOI":"10.1016\/j.cell.2013.08.029","article-title":"Transcription Recovery after DNA Damage Requires Chromatin Priming by the H3.3 Histone Chaperone HIRA","volume":"155","author":"Adam","year":"2013","journal-title":"Cell"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"547","DOI":"10.1016\/j.molcel.2016.01.019","article-title":"PARP1 Links CHD2-Mediated Chromatin Expansion and H3.3 Deposition to DNA Repair by Non-Homologous End-Joining","volume":"61","author":"Luijsterburg","year":"2016","journal-title":"Mol. Cell"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1253","DOI":"10.1101\/gad.566910","article-title":"The Death-Associated Protein DAXX Is a Novel Histone Chaperone Involved in the Replication-Independent Deposition of H3.3","volume":"24","author":"Ouararhni","year":"2010","journal-title":"Genes Dev."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"97","DOI":"10.1016\/j.tcb.2005.12.002","article-title":"Daxx: Death or Survival Protein?","volume":"16","author":"Salomoni","year":"2006","journal-title":"Trends Cell Biol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"14075","DOI":"10.1073\/pnas.1008850107","article-title":"Daxx Is an H3.3-Specific Histone Chaperone and Cooperates with ATRX in Replication-Independent Chromatin Assembly at Telomeres","volume":"107","author":"Lewis","year":"2010","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Lovejoy, C.A., Li, W., Reisenweber, S., Thongthip, S., Bruno, J., de Lange, T., De, S., Petrini, J.H.J., Sung, P.A., and Jasin, M. (2012). Loss of ATRX, Genome Instability, and an Altered DNA Damage Response Are Hallmarks of the Alternative Lengthening of Telomeres Pathway. PLoS Genet., 8.","DOI":"10.1371\/journal.pgen.1002772"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"11","DOI":"10.1016\/j.molcel.2018.05.014","article-title":"ATRX Promotes DNA Repair Synthesis and Sister Chromatid Exchange during Homologous Recombination","volume":"71","author":"Elbakry","year":"2018","journal-title":"Mol. Cell"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1136","DOI":"10.1093\/nar\/gkad1141","article-title":"DAXX Promotes Centromeric Stability Independently of ATRX by Preventing the Accumulation of R-Loop-Induced DNA Double-Stranded Breaks","volume":"52","author":"Pinto","year":"2024","journal-title":"Nucleic Acids Res."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"132","DOI":"10.1038\/s41586-021-03824-5","article-title":"DAXX Represents a New Type of Protein-Folding Enabler","volume":"597","author":"Huang","year":"2021","journal-title":"Nature"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"1193","DOI":"10.1038\/s41467-017-01206-y","article-title":"Structural and Mechanistic Insights into ATRX-Dependent and -Independent Functions of the Histone Chaperone DAXX","volume":"8","author":"Hoelper","year":"2017","journal-title":"Nat. Commun."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"724","DOI":"10.1038\/sj.cdd.4401559","article-title":"Daxx Is Required for Stress-Induced Cell Death and JNK Activation","volume":"12","author":"Khelifi","year":"2005","journal-title":"Cell Death Differ."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"171","DOI":"10.1080\/15548627.2019.1677323","article-title":"Histone H3F3\/H3.3 Chaperone DAXX Converts to Modulate SQSTM1 Phase Condensation for NFE2L2 Activation","volume":"16","author":"Yang","year":"2020","journal-title":"Autophagy"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"379","DOI":"10.1007\/978-1-61779-998-3_27","article-title":"I-SceI-Based Assays to Examine Distinct Repair Outcomes of Mammalian Chromosomal Double Strand Breaks","volume":"920","author":"Gunn","year":"2012","journal-title":"Methods Mol. Biol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"101604","DOI":"10.1016\/j.isci.2020.101604","article-title":"Targeting SMYD3 to Sensitize Homologous Recombination-Proficient Tumors to PARP-Mediated Synthetic Lethality","volume":"23","author":"Sanese","year":"2020","journal-title":"iScience"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"995","DOI":"10.1038\/nmeth947","article-title":"Direct Observation of Individual Endogenous Protein Complexes in Situ by Proximity Ligation","volume":"3","author":"Gullberg","year":"2006","journal-title":"Nat. Methods"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"339","DOI":"10.1007\/s00018-024-05380-3","article-title":"R-Loops and Impaired Autophagy Trigger cGAS-Dependent Inflammation via Micronuclei Formation in Senataxin-Deficient Cells","volume":"81","author":"Zannini","year":"2024","journal-title":"Cell Mol. Life Sci."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"8602","DOI":"10.1128\/MCB.20.22.8602-8612.2000","article-title":"Chromatin Association of Human Origin Recognition Complex, Cdc6, and Minichromosome Maintenance Proteins during the Cell Cycle: Assembly of Prereplication Complexes in Late Mitosis","volume":"20","author":"Stillman","year":"2000","journal-title":"Mol. Cell Biol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"1445","DOI":"10.1038\/nprot.2007.202","article-title":"Extraction, Purification and Analysis of Histones","volume":"2","author":"Shechter","year":"2007","journal-title":"Nat. Protoc."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"2901","DOI":"10.4161\/15384101.2014.946852","article-title":"Bimodal Regulation of P21(Waf1) Protein as Function of DNA Damage Levels","volume":"13","author":"Buscemi","year":"2014","journal-title":"Cell Cycle"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"167","DOI":"10.1016\/j.molcel.2008.05.017","article-title":"ATM Signaling Facilitates Repair of DNA Double-Strand Breaks Associated with Heterochromatin","volume":"31","author":"Goodarzi","year":"2008","journal-title":"Mol. Cell"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"17817","DOI":"10.18632\/oncotarget.4417","article-title":"CCAR2\/DBC1 Is Required for Chk2-Dependent KAP1 Phosphorylation and Repair of DNA Damage","volume":"6","author":"Magni","year":"2015","journal-title":"Oncotarget"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"197","DOI":"10.1038\/s41419-023-05722-2","article-title":"Sex Specific Regulation of TSPY-Like 2 in the DNA Damage Response of Cancer Cells","volume":"14","author":"Cardano","year":"2023","journal-title":"Cell Death Dis."},{"key":"ref_27","first-page":"5489","article-title":"Single-Cell Analysis of Daxx and ATRX-Dependent Transcriptional Repression","volume":"125","author":"Newhart","year":"2012","journal-title":"J. Cell Sci."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"1446","DOI":"10.1038\/emboj.2010.38","article-title":"High-Resolution Profiling of gammaH2AX around DNA Double Strand Breaks in the Mammalian Genome","volume":"29","author":"Iacovoni","year":"2010","journal-title":"EMBO J."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"18271","DOI":"10.1074\/jbc.M113.473199","article-title":"Histone Acetyltransferase 1 Promotes Homologous Recombination in DNA Repair by Facilitating Histone Turnover","volume":"288","author":"Yang","year":"2013","journal-title":"J. Biol. Chem."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1026","DOI":"10.1242\/jcs.02816","article-title":"The Number of PML Nuclear Bodies Increases in Early S Phase by a Fission Mechanism","volume":"119","author":"Dellaire","year":"2006","journal-title":"J. Cell Sci."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"730","DOI":"10.1038\/35036365","article-title":"The Function of PML in P53-Dependent Apoptosis","volume":"2","author":"Guo","year":"2000","journal-title":"Nat. Cell Biol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"114","DOI":"10.1016\/j.dnarep.2015.09.014","article-title":"Chromatin Dynamics after DNA Damage: The Legacy of the Access-Repair-Restore Model","volume":"36","author":"Polo","year":"2015","journal-title":"DNA Repair"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"7","DOI":"10.1038\/nrm3719","article-title":"Double-Strand Break Repair: 53BP1 Comes into Focus","volume":"15","author":"Panier","year":"2014","journal-title":"Nat. Rev. Mol. Cell Biol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"858","DOI":"10.1016\/j.molcel.2013.01.002","article-title":"RIF1 Is Essential for 53BP1-Dependent Nonhomologous End Joining and Suppression of DNA Double-Strand Break Resection","volume":"49","author":"Chapman","year":"2013","journal-title":"Mol. Cell"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"1203","DOI":"10.1093\/annonc\/mdy099","article-title":"RAD51 Foci as a Functional Biomarker of Homologous Recombination Repair and PARP Inhibitor Resistance in Germline BRCA-Mutated Breast Cancer","volume":"29","author":"Cruz","year":"2018","journal-title":"Ann. Oncol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"40618","DOI":"10.1074\/jbc.M112.410951","article-title":"RING Finger Nuclear Factor RNF168 Is Important for Defects in Homologous Recombination Caused by Loss of the Breast Cancer Susceptibility Factor BRCA1","volume":"287","author":"Laulier","year":"2012","journal-title":"J. Biol. Chem."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"11135","DOI":"10.1074\/jbc.M113.457440","article-title":"RIF1 Counteracts BRCA1-Mediated End Resection during DNA Repair","volume":"288","author":"Feng","year":"2013","journal-title":"J. Biol. Chem."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"493","DOI":"10.1667\/RR0788.1","article-title":"Phosphorylation of Histone H2AX in Radiation-Induced Micronuclei","volume":"168","author":"Medvedeva","year":"2007","journal-title":"Radiat. Res."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"19855","DOI":"10.1073\/pnas.0707579104","article-title":"Profiling of UV-Induced ATM\/ATR Signaling Pathways","volume":"104","author":"Stokes","year":"2007","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"1160","DOI":"10.1126\/science.1140321","article-title":"ATM and ATR Substrate Analysis Reveals Extensive Protein Networks Responsive to DNA Damage","volume":"316","author":"Matsuoka","year":"2007","journal-title":"Science"},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"1344","DOI":"10.1126\/science.aae0065","article-title":"The Histone H3.3K36M Mutation Reprograms the Epigenome of Chondroblastomas","volume":"352","author":"Fang","year":"2016","journal-title":"Science"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"406","DOI":"10.1038\/sj.emboj.7601967","article-title":"Dynamic Histone H3 Methylation during Gene Induction: HYPB\/Setd2 Mediates All H3K36 Trimethylation","volume":"27","author":"Edmunds","year":"2008","journal-title":"EMBO J."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"366","DOI":"10.1038\/nsmb.2796","article-title":"Transcriptionally Active Chromatin Recruits Homologous Recombination at DNA Double-Strand Breaks","volume":"21","author":"Aymard","year":"2014","journal-title":"Nat. Struct. Mol. Biol."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"700","DOI":"10.1126\/science.1231573","article-title":"53BP1 Regulates DSB Repair Using Rif1 to Control 5\u2032 End Resection","volume":"339","author":"Zimmermann","year":"2013","journal-title":"Science"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"395","DOI":"10.1101\/sqb.2000.65.395","article-title":"The Interplay between Nonhomologous End-Joining and Cell Cycle Checkpoint Factors in Development, Genomic Stability, and Tumorigenesis","volume":"65","author":"Ferguson","year":"2000","journal-title":"Cold Spring Harb. Symp. Quant. Biol."},{"key":"ref_46","doi-asserted-by":"crossref","unstructured":"Tang, J., Agrawal, T., Cheng, Q., Qu, L., Brewer, M.D., Chen, J., and Yang, X. (2013). Phosphorylation of Daxx by ATM Contributes to DNA Damage-Induced P53 Activation. PLoS ONE, 8.","DOI":"10.1371\/annotation\/032b026d-5bcf-4c1b-aa15-65c226a53818"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"4318","DOI":"10.1093\/nar\/gku090","article-title":"DAXX Co-Folds with H3.3\/H4 Using High Local Stability Conferred by the H3.3 Variant Recognition Residues","volume":"42","author":"DeNizio","year":"2014","journal-title":"Nucleic Acids Res."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"402","DOI":"10.1038\/onc.2015.311","article-title":"ATM-Mediated KDM2A Phosphorylation Is Required for the DNA Damage Repair","volume":"35","author":"Cao","year":"2016","journal-title":"Oncogene"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"2006","DOI":"10.1016\/j.celrep.2014.05.026","article-title":"SETD2-Dependent Histone H3K36 Trimethylation Is Required for Homologous Recombination Repair and Genome Stability","volume":"7","author":"Pfister","year":"2014","journal-title":"Cell Rep."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"226","DOI":"10.1038\/nature10833","article-title":"Driver Mutations in Histone H3.3 and Chromatin Remodelling Genes in Paediatric Glioblastoma","volume":"482","author":"Schwartzentruber","year":"2012","journal-title":"Nature"},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"659","DOI":"10.1007\/s00401-013-1095-8","article-title":"Mutations in SETD2 and Genes Affecting Histone H3K36 Methylation Target Hemispheric High-Grade Gliomas","volume":"125","author":"Fontebasso","year":"2013","journal-title":"Acta Neuropathol."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"5699","DOI":"10.1038\/onc.2015.24","article-title":"SETD2 Loss-of-Function Promotes Renal Cancer Branched Evolution through Replication Stress and Impaired DNA Repair","volume":"34","author":"Kanu","year":"2015","journal-title":"Oncogene"}],"container-title":["Cells"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4409\/15\/2\/162\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,1,18]],"date-time":"2026-01-18T05:11:29Z","timestamp":1768713089000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4409\/15\/2\/162"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,1,16]]},"references-count":52,"journal-issue":{"issue":"2","published-online":{"date-parts":[[2026,1]]}},"alternative-id":["cells15020162"],"URL":"https:\/\/doi.org\/10.3390\/cells15020162","relation":{},"ISSN":["2073-4409"],"issn-type":[{"value":"2073-4409","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,1,16]]}}}