{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,10]],"date-time":"2026-06-10T14:33:53Z","timestamp":1781102033604,"version":"3.54.1"},"reference-count":74,"publisher":"National Academy of Sciences","issue":"14","license":[{"start":{"date-parts":[[2026,4,3]],"date-time":"2026-04-03T00:00:00Z","timestamp":1775174400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001691","name":"MEXT | Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["JP22H00449"],"award-info":[{"award-number":["JP22H00449"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001691","name":"MEXT | Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["JP22H00449"],"award-info":[{"award-number":["JP22H00449"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001691","name":"MEXT | Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["JP 25H01028"],"award-info":[{"award-number":["JP 25H01028"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001691","name":"MEXT | Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["JP 25H01028"],"award-info":[{"award-number":["JP 25H01028"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001691","name":"MEXT | Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["JP22K06202"],"award-info":[{"award-number":["JP22K06202"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["www.pnas.org"],"crossmark-restriction":true},"short-container-title":["Proc. Natl. Acad. Sci. U.S.A."],"published-print":{"date-parts":[[2026,4,7]]},"abstract":"<jats:p>\n                    Class switch recombination (CSR) and somatic hypermutation are essential mechanisms of effective antibody production, dependent on the enzyme activation-induced cytidine deaminase (AID). Since AID lacks the intrinsic ability to cleave DNA, topoisomerase 1 (TOP1) has been hypothesized to mediate AID- and transcription-dependent DNA cleavage. However, the molecular mechanism underlying the formation of an irreversible TOP1\u2013DNA cleavage complex (TOP1-CC) following cytidine to uridine (C-to-U) editing by AID remains undefined. To unveil this mechanism, we tested antisense oligonucleotides (ASOs) targeting noncoding germline transcripts (GLTs) that form R-loops across DNA cleavage sites during CSR. These studies revealed that the anti-GLT-ASOs increased DNA cleavage frequency, concomitant with a decrease in R-loops. Mechanistically, ASO-mediated R-loop disruption led to increased TOP1-CC formation specifically within the GLT region during CSR. Furthermore, an in vitro transcription assay demonstrated that RNase H-mediated R-loop degradation enhanced positive supercoiling and double-strand breaks in the presence of TOP1, even in the absence of AID. Moreover, knockout of Tyrosyl-DNA phosphodiesterase 1 (\n                    <jats:italic toggle=\"yes\">Tdp1<\/jats:italic>\n                    ), a canonical TOP1-CC processing enzyme, revealed that TDP1 suppresses CSR, indicating that accumulated TOP1-CCs in\n                    <jats:italic toggle=\"yes\">Tdp1<\/jats:italic>\n                    knockout cells serve as substrates for CSR through an alternative TOP1-CC processing pathway. Collectively, these results suggest the R-loop destabilization as a critical, hitherto unrecognized intermediary step linking C-to-U editing to TOP1-CC formation. AID-mediated C-to-U editing within R-loops introduces a G:U wobble base pair, which we hypothesize renders R-loops unstable and sensitive to nuclear RNases, leading to R-loop disruption; this disruption consequently induces the TOP1-CCs and results in CSR, as confirmed in this study.\n                  <\/jats:p>","DOI":"10.1073\/pnas.2533648123","type":"journal-article","created":{"date-parts":[[2026,4,3]],"date-time":"2026-04-03T17:36:37Z","timestamp":1775237797000},"update-policy":"https:\/\/doi.org\/10.1073\/pnas.cm10313","source":"Crossref","is-referenced-by-count":0,"title":["R-loop disruption triggers irreversible topoisomerase 1\u2013DNA cleavage complex formation during AID-dependent IgH gene diversification"],"prefix":"10.1073","volume":"123","author":[{"given":"Maki","family":"Kobayashi","sequence":"first","affiliation":[{"name":"Department of Immunology and Genomic Medicine, Center for Cancer Immunotherapy and Immunobiology, Graduate School of Medicine, Kyoto 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Kobayashi X. Chen T. Honjo Source data and uncropped images for \u201cR-loop disruption triggers irreversible topoisomerase 1-DNA cleavage complex formation during AID-dependent IgH gene diversification.\u201d Figshare. https:\/\/doi.org\/10.6084\/m9.figshare.31800808. Deposited 18 March 2026."}],"container-title":["Proceedings of the National Academy of Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/pnas.org\/doi\/pdf\/10.1073\/pnas.2533648123","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,4,3]],"date-time":"2026-04-03T17:36:50Z","timestamp":1775237810000},"score":1,"resource":{"primary":{"URL":"https:\/\/pnas.org\/doi\/10.1073\/pnas.2533648123"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,4,3]]},"references-count":74,"journal-issue":{"issue":"14","published-print":{"date-parts":[[2026,4,7]]}},"alternative-id":["10.1073\/pnas.2533648123"],"URL":"https:\/\/doi.org\/10.1073\/pnas.2533648123","relation":{},"ISSN":["0027-8424","1091-6490"],"issn-type":[{"value":"0027-8424","type":"print"},{"value":"1091-6490","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,4,3]]},"assertion":[{"value":"2025-11-23","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2026-02-18","order":2,"name":"accepted","label":"Accepted","group":{"name":"publication_history","label":"Publication History"}},{"value":"2026-04-03","order":3,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}],"article-number":"e2533648123"}}