The two-step purification method ViREn identifies a single NSUN6-mediated 5-methylcytosine modification promoting dengue virus RNA genome turnover.
| Title: | The two-step purification method ViREn identifies a single NSUN6-mediated 5-methylcytosine modification promoting dengue virus RNA genome turnover. |
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| Authors: | Wu CC; Department of Infectious Diseases, Molecular Virology, Center for Integrative Infectious Disease Research, Heidelberg University, Medical Faculty Heidelberg, 69120 Heidelberg, Germany.; Naarmann-de Vries IS; Klaus Tschira Institute for Integrative Computational Cardiology, Heidelberg University, Medical Faculty Heidelberg, 69120 Heidelberg, Germany.; German Centre for Cardiovascular Research (DZHK)-Partner Site Heidelberg/Mannheim, 69120 Heidelberg, Germany.; Hartmann J; Department of Infectious Diseases, Molecular Virology, Center for Integrative Infectious Disease Research, Heidelberg University, Medical Faculty Heidelberg, 69120 Heidelberg, Germany.; Nidoieva Z; Institute of Pharmaceutical and Biomedical Sciences, Johannes Gutenberg University Mainz, 55128 Mainz, Germany.; Kopietz K; Institute of Pharmaceutical and Biomedical Sciences, Johannes Gutenberg University Mainz, 55128 Mainz, Germany.; Marchand V; Université de Lorraine, SMP IBSLor, EpiRNA-Seq Core Facility, 54000 Nancy, France.; Université de Lorraine, CNRS, UMR7365 IMoPA, 54000 Nancy, France.; Özrendeci Z; Institute of Pharmaceutical and Biomedical Sciences, Johannes Gutenberg University Mainz, 55128 Mainz, Germany.; Lindner D; Division of Biochemistry, Mannheim Institute for Innate Immunoscience (MI3) and Mannheim Cancer Center (MCC), Heidelberg University, Medical Faculty Mannheim, 68167 Mannheim, Germany.; Center for Molecular Biology of Heidelberg University (ZMBH), German Cancer Research Center (DKFZ)-ZMBH Alliance, 69120 Heidelberg, Germany.; Schelchshorn S; Department of Infectious Diseases, Molecular Virology, Center for Integrative Infectious Disease Research, Heidelberg University, Medical Faculty Heidelberg, 69120 Heidelberg, Germany.; Flad S; Division of Mechanisms Regulating Gene Expression, German Cancer Research Center-Deutsches Krebsforschungszentrum (DKFZ), 69120 Heidelberg, Germany.; Frye M; Division of Mechanisms Regulating Gene Expression, German Cancer Research Center-Deutsches Krebsforschungszentrum (DKFZ), 69120 Heidelberg, Germany.; Papavasiliou FN; Division of Immune Diversity, German Cancer Research Center-Deutsches Krebsforschungszentrum (DKFZ), 69120 Heidelberg, Germany.; Schirmeister T; Institute of Pharmaceutical and Biomedical Sciences, Johannes Gutenberg University Mainz, 55128 Mainz, Germany.; Stoecklin G; Division of Biochemistry, Mannheim Institute for Innate Immunoscience (MI3) and Mannheim Cancer Center (MCC), Heidelberg University, Medical Faculty Mannheim, 68167 Mannheim, Germany.; Center for Molecular Biology of Heidelberg University (ZMBH), German Cancer Research Center (DKFZ)-ZMBH Alliance, 69120 Heidelberg, Germany.; Schott J; Division of Biochemistry, Mannheim Institute for Innate Immunoscience (MI3) and Mannheim Cancer Center (MCC), Heidelberg University, Medical Faculty Mannheim, 68167 Mannheim, Germany.; Center for Molecular Biology of Heidelberg University (ZMBH), German Cancer Research Center (DKFZ)-ZMBH Alliance, 69120 Heidelberg, Germany.; Motorin Y; Université de Lorraine, SMP IBSLor, EpiRNA-Seq Core Facility, 54000 Nancy, France.; Université de Lorraine, CNRS, UMR7365 IMoPA, 54000 Nancy, France.; Tuorto F; Division of Biochemistry, Mannheim Institute for Innate Immunoscience (MI3) and Mannheim Cancer Center (MCC), Heidelberg University, Medical Faculty Mannheim, 68167 Mannheim, Germany.; Center for Molecular Biology of Heidelberg University (ZMBH), German Cancer Research Center (DKFZ)-ZMBH Alliance, 69120 Heidelberg, Germany.; Dieterich C; Klaus Tschira Institute for Integrative Computational Cardiology, Heidelberg University, Medical Faculty Heidelberg, 69120 Heidelberg, Germany.; German Centre for Cardiovascular Research (DZHK)-Partner Site Heidelberg/Mannheim, 69120 Heidelberg, Germany.; Helm M; Institute of Pharmaceutical and Biomedical Sciences, Johannes Gutenberg University Mainz, 55128 Mainz, Germany.; Ruggieri A; Department of Infectious Diseases, Molecular Virology, Center for Integrative Infectious Disease Research, Heidelberg University, Medical Faculty Heidelberg, 69120 Heidelberg, Germany. |
| Source: | Nucleic acids research [Nucleic Acids Res] 2026 Jan 14; Vol. 54 (2). |
| Publication Type: | Journal Article |
| Language: | English |
| Journal Info: | Publisher: Oxford University Press Country of Publication: England NLM ID: 0411011 Publication Model: Print Cited Medium: Internet ISSN: 1362-4962 (Electronic) Linking ISSN: 03051048 NLM ISO Abbreviation: Nucleic Acids Res Subsets: MEDLINE |
| Imprint Name(s): | Publication: 1992- : Oxford : Oxford University Press; Original Publication: London, Information Retrieval ltd. |
| MeSH Terms: | Dengue Virus*/genetics ; RNA, Viral*/metabolism ; RNA, Viral*/isolation & purification ; RNA, Viral*/genetics ; RNA, Viral*/chemistry ; 5-Methylcytosine*/metabolism ; Methyltransferases*/metabolism ; Methyltransferases*/genetics ; Genome, Viral*; Humans ; RNA Stability ; RNA Processing, Post-Transcriptional |
| Abstract: | Chemical modifications on cellular and viral RNAs are new layers of post-transcriptional regulation of cellular processes, including RNA stability and translation. Although advances in analytical methods have improved the detection of RNA modifications, precise mapping at single-base resolution remains challenging. Requirements for sensitivity and purity limit accuracy and reproducibility, especially for low abundant viral RNAs extracted from infected cells. Here, we report a two-step method, ViREn, for the enrichment of the genomic RNA (gRNA) of dengue virus (DENV), a positive-sense single-stranded RNA virus. This approach enabled the preparation of gRNA with significantly increased purity and led to the identification of a single high-confidence 5-methylcytosine (m5C) site in DENV gRNA at position 1218. This finding was orthogonally validated by Illumina-based bisulfite sequencing and by Nanopore Oxford Technologies direct RNA sequencing. Strikingly, m5C1218 was detected exclusively in gRNA extracted from infected cells but not in gRNA extracted from viral particles. We identified NSUN6 as the host methyltransferase catalyzing this modification and demonstrated a role for m5C in regulating DENV gRNA turnover. ViREn thus enables the mapping of m5C on low-abundance viral gRNA with unprecedented precision and sensitivity and facilitates future mechanistic studies into the role of RNA modifications in virus replication.; (© The Author(s) 2026. Published by Oxford University Press.) |
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| Grant Information: | 439669440 Deutsche Forschungsgemeinschaft; 445549683 Deutsche Forschungsgemeinschaft; French Grand Est Region Project; Heidelberg University |
| Substance Nomenclature: | 0 (RNA, Viral); 6R795CQT4H (5-Methylcytosine); EC 2.1.1.- (Methyltransferases) |
| Entry Date(s): | Date Created: 20260121 Date Completed: 20260121 Latest Revision: 20260125 |
| Update Code: | 20260130 |
| PubMed Central ID: | PMC12820529 |
| DOI: | 10.1093/nar/gkag003 |
| PMID: | 41562255 |
| Database: | MEDLINE |
Journal Article