{"doi":"10.64898/2026.03.16.712100","title":"Molecular basis of tRNA modification by the human m\n                  <sup>5</sup>\n                  C methyltransferase NSUN2","abstract":"<jats:title>ABSTRACT</jats:title>\n                <jats:p>\n                  RNA 5-methylcytidine (m\n                  <jats:sup>5</jats:sup>\n                  C) is a prevalent modification that drives RNA stability and function. In humans, m\n                  <jats:sup>5</jats:sup>\n                  C is deposited on distinct RNA substrates by DNMT2/TRDMT1 and the NSUN family, to regulate diverse cellular processes, but how m\n                  <jats:sup>5</jats:sup>\n                  C writers recognise their substrates remains unclear. NSUN2 is a major m\n                  <jats:sup>5</jats:sup>\n                  C methyltransferase with broad roles in cell physiology and strong links to cancer and neurodevelopmental disorders\n                  <jats:sup>1</jats:sup>\n                  . Here, we reconstitute an active human NSUN2-tRNA complex and capture its post-catalytic, tRNA-bound structure at 3.1 Å resolution. Using an integrated approach combining biochemistry, cryo-electron microscopy, crosslinking mass spectrometry and molecular dynamics simulations, we show that NSUN2 remodels the tRNA to access the variable-loop target cytidine. Recognition is driven by RNA architecture, with NSUN2 exploiting the L-shaped tRNA scaffold to position the target base in the catalytic centre. We further show that Gly679 at the NSUN2-tRNA interface is important for the stability of the complex, providing a mechanistic basis for how the disease-associated Gly679Arg substitution can impair tRNA binding. Together, these findings establish an RNA-structure-guided mechanism for NSUN2 substrate recognition and methylation and provide general principles for m\n                  <jats:sup>5</jats:sup>\n                  C deposition on cellular RNAs and their fundamental role in disease.\n                </jats:p>","journal":null,"year":null,"id":630219,"datarank":0.10397207708399181,"base_score":0.6931471805599453,"endowment":0.6931471805599453,"self_citation_contribution":0.10397207708399181,"citation_network_contribution":0.0,"self_endowment_contribution":0.10397207708399181,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":1,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":null,"is_data_producer":false,"deposit_databanks":null,"is_oa":false,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":null,"fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1632516,"name":"Michelangelo Lassandro","orcid":"0009-0006-7265-0602","position":1,"is_corresponding":false},{"id":1632517,"name":"Arianna Di Fazio","orcid":"0000-0002-5921-3495","position":2,"is_corresponding":false},{"id":1632518,"name":"Alessio Di Ianni","orcid":"0000-0002-2902-3797","position":3,"is_corresponding":false},{"id":1632519,"name":"Kanhaya Lal","orcid":"0000-0001-8555-7948","position":4,"is_corresponding":false},{"id":1632520,"name":"Jon Rodríguez-Villa","orcid":"0009-0002-7291-5763","position":5,"is_corresponding":false},{"id":810039,"name":"Alice Rossi","orcid":"0000-0003-2643-028X","position":6,"is_corresponding":false},{"id":1016990,"name":"Andrea Graziadei","orcid":"0000-0001-7709-6002","position":7,"is_corresponding":false},{"id":1632521,"name":"Monika Gullerova","orcid":"0000-0002-4512-2779","position":8,"is_corresponding":false},{"id":369403,"name":"Ana Casañal","orcid":"0000-0002-0334-0591","position":9,"is_corresponding":false},{"id":638325,"name":"Elodie Leroy","orcid":"0000-0001-7902-2162","position":0,"is_corresponding":false}],"reference_count":0,"raw_metadata":{"has_enrichment":true,"resolved":true,"title":"Molecular basis of tRNA modification by the human m\n                  <sup>5</sup>\n                  C methyltransferase NSUN2","abstract":"<jats:title>ABSTRACT</jats:title>\n                <jats:p>\n                  RNA 5-methylcytidine (m\n                  <jats:sup>5</jats:sup>\n                  C) is a prevalent modification that drives RNA stability and function. In humans, m\n                  <jats:sup>5</jats:sup>\n                  C is deposited on distinct RNA substrates by DNMT2/TRDMT1 and the NSUN family, to regulate diverse cellular processes, but how m\n                  <jats:sup>5</jats:sup>\n                  C writers recognise their substrates remains unclear. NSUN2 is a major m\n                  <jats:sup>5</jats:sup>\n                  C methyltransferase with broad roles in cell physiology and strong links to cancer and neurodevelopmental disorders\n                  <jats:sup>1</jats:sup>\n                  . Here, we reconstitute an active human NSUN2-tRNA complex and capture its post-catalytic, tRNA-bound structure at 3.1 Å resolution. Using an integrated approach combining biochemistry, cryo-electron microscopy, crosslinking mass spectrometry and molecular dynamics simulations, we show that NSUN2 remodels the tRNA to access the variable-loop target cytidine. Recognition is driven by RNA architecture, with NSUN2 exploiting the L-shaped tRNA scaffold to position the target base in the catalytic centre. We further show that Gly679 at the NSUN2-tRNA interface is important for the stability of the complex, providing a mechanistic basis for how the disease-associated Gly679Arg substitution can impair tRNA binding. Together, these findings establish an RNA-structure-guided mechanism for NSUN2 substrate recognition and methylation and provide general principles for m\n                  <jats:sup>5</jats:sup>\n                  C deposition on cellular RNAs and their fundamental role in disease.\n                </jats:p>","is_dataset_classified":null,"base_score":0.0,"endowment":0.0,"datacite_reuse_total":0,"file_count":0,"downloads":0,"views":0,"has_version_chain":false,"is_dataset":false,"is_oa":false,"pmid":null,"pmcid":null,"openalex_id":null,"authors":[],"funders":[{"funder_name":"","grant_id":"BVR025900","title":null},{"funder_name":"EPA Trust Fund","grant_id":"BVR01670","title":null},{"funder_name":"EMBO Long Term Fellowship","grant_id":"ALTF 894-202","title":null}],"total_grants":3,"fwci":null,"citation_percentile":null,"influential_citations":0,"citation_trend":[],"oa_status":"closed","license":"https://www.biorxiv.org/about/FAQ#license","oa_locations":[{"url":"https://syndication.highwire.org/content/doi/10.64898/2026.03.16.712100","host_type":"publisher"}],"fields_of_study":[],"mesh_terms":[],"keywords":[],"sdg_mappings":[],"linked_datasets":[],"clinical_trials":[],"software_tools":[],"database_accessions":[],"source":"live","citation_network_status":"fetched"},"created_at":"2026-08-05T20:42:20.946725Z","pmid":null,"pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}