{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,2]],"date-time":"2026-07-02T19:12:57Z","timestamp":1783019577458,"version":"3.54.6"},"reference-count":55,"publisher":"Public Library of Science (PLoS)","issue":"7","license":[{"start":{"date-parts":[[2026,7,2]],"date-time":"2026-07-02T00:00:00Z","timestamp":1782950400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000057","name":"National Institute of General Medical Sciences","doi-asserted-by":"publisher","award":["R01GM133810"],"award-info":[{"award-number":["R01GM133810"]}],"id":[{"id":"10.13039\/100000057","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS One"],"abstract":"<jats:p>\n                    The Epstein\u2013Barr virus (EBV) infects over 95% of adults, establishing lifelong latency and contributing to the development of various malignancies, including Burkitt lymphoma and nasopharyngeal carcinoma. However, the RNA structures regulating the splicing of the critical EBV gene, latent membrane protein 1 (\n                    <jats:italic>LMP1<\/jats:italic>\n                    ), remain uncharacterized. To identify these regulatory elements, we applied spliceosome inhibition with RNA probing and sequencing (SIRP-seq) to the BJAB-B1 cell line. By utilizing the spliceosome inhibitor pladienolide B, we enriched pre-mRNA species, enabling the detection of structural features within both the full-length pre-mRNA (\n                    <jats:italic>LMP1-FL<\/jats:italic>\n                    ) and an alternatively spliced isoform retaining intron 2 (\n                    <jats:italic>LMP1-AS<\/jats:italic>\n                    ). The resulting chemical probing datasets informed the RNA folding algorithms RNAfold and ScanFold to generate the first high-resolution secondary structure models for the\n                    <jats:italic>LMP1<\/jats:italic>\n                    pre-mRNA, encompassing both exonic and intronic regions. Our results identify 11 novel, thermodynamically stable RNA structures, with several key elements positioned near splice junctions. Notably, three structures (Structures 8, 9, and 10) were identified near the 3\u2032 splice site of intron 2, appearing in alternative conformations that may influence splicing accessibility. Furthermore, these structures map to regions containing disease-relevant mutations associated with patient survival in Burkitt lymphoma. This structural framework provides new insights into how\n                    <jats:italic>LMP1<\/jats:italic>\n                    splicing may be regulated by RNA structure and identifies potential novel therapeutic targets for mitigating EBV-associated diseases.\n                  <\/jats:p>","DOI":"10.1371\/journal.pone.0345208","type":"journal-article","created":{"date-parts":[[2026,7,2]],"date-time":"2026-07-02T17:58:23Z","timestamp":1783015103000},"page":"e0345208","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":0,"title":["DMS-informed secondary structure modeling of Epstein\u2013Barr Virus LMP-1 pre-mRNA defines novel elements spanning introns"],"prefix":"10.1371","volume":"21","author":[{"given":"Taylor O.","family":"Eich","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Evelyn C.","family":"Coppenbarger","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Abdelraouf O.","family":"Dapour","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6419-5570","authenticated-orcid":true,"given":"Walter N.","family":"Moss","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2026,7,2]]},"reference":[{"key":"pone.0345208.ref001","volume-title":"Burkitt Lymphoma","author":"PT Naing","year":"2025"},{"issue":"1","key":"pone.0345208.ref002","doi-asserted-by":"crossref","first-page":"31","DOI":"10.1007\/s00432-021-03824-y","article-title":"Estimating the global burden of Epstein-Barr virus-related cancers","volume":"148","author":"Y Wong","year":"2022","journal-title":"J Cancer Res Clin Oncol"},{"issue":"1","key":"pone.0345208.ref003","doi-asserted-by":"crossref","first-page":"75","DOI":"10.1016\/j.beha.2012.01.005","article-title":"EBV-associated lymphomas in adults","volume":"25","author":"M Roschewski","year":"2012","journal-title":"Best Pract Res Clin Haematol"},{"issue":"10","key":"pone.0345208.ref004","doi-asserted-by":"crossref","first-page":"1673","DOI":"10.1200\/JCO.1987.5.10.1673","article-title":"Lymphoproliferative diseases in immunocompromised hosts: the role of Epstein-Barr virus","volume":"5","author":"AF List","year":"1987","journal-title":"J Clin Oncol"},{"issue":"5","key":"pone.0345208.ref005","first-page":"1511","article-title":"Association of Epstein-Barr virus (EBV) with Sj\u00f6gren\u2019s syndrome: differential EBV expression between epithelial cells and lymphocytes in salivary glands","volume":"149","author":"S Wen","year":"1996","journal-title":"Am J Pathol"},{"issue":"4","key":"pone.0345208.ref006","doi-asserted-by":"crossref","first-page":"1018","DOI":"10.1084\/jem.147.4.1018","article-title":"Lymphocytes transformed by Epstein-Barr virus. Induction of nuclear antigen reactive with antibody in rheumatoid arthritis","volume":"147","author":"MA Alspaugh","year":"1978","journal-title":"J Exp Med"},{"issue":"6","key":"pone.0345208.ref007","doi-asserted-by":"crossref","first-page":"695","DOI":"10.1289\/ehp.11092","article-title":"Recent advances and opportunities in research on lupus: environmental influences and mechanisms of disease","volume":"116","author":"GS Cooper","year":"2008","journal-title":"Environ Health Perspect"},{"issue":"6578","key":"pone.0345208.ref008","doi-asserted-by":"crossref","first-page":"296","DOI":"10.1126\/science.abj8222","article-title":"Longitudinal analysis reveals high prevalence of Epstein-Barr virus associated with multiple sclerosis","volume":"375","author":"K Bjornevik","year":"2022","journal-title":"Science"},{"key":"pone.0345208.ref009","article-title":"EBV: immunobiology and host response","author":"DJ Moss","year":"2007"},{"key":"pone.0345208.ref010","doi-asserted-by":"crossref","first-page":"29","DOI":"10.1146\/annurev-pathmechdis-012418-013023","article-title":"Epstein-Barr Virus and Cancer","volume":"14","author":"PJ Farrell","year":"2019","journal-title":"Annu Rev Pathol"},{"issue":"3","key":"pone.0345208.ref011","doi-asserted-by":"crossref","first-page":"1031","DOI":"10.1128\/jvi.63.3.1031-1039.1989","article-title":"Distinction between Epstein-Barr virus type A (EBNA 2A) and type B (EBNA 2B) isolates extends to the EBNA 3 family of nuclear proteins","volume":"63","author":"M Rowe","year":"1989","journal-title":"J Virol"},{"issue":"2","key":"pone.0345208.ref012","doi-asserted-by":"crossref","DOI":"10.1371\/journal.ppat.1010254","article-title":"Epstein-Barr virus oncoprotein-driven B cell metabolism remodeling","volume":"18","author":"EM Burton","year":"2022","journal-title":"PLoS Pathog"},{"issue":"12","key":"pone.0345208.ref013","doi-asserted-by":"crossref","DOI":"10.3390\/v13122344","article-title":"Molecular basis of Epstein-Barr virus latency establishment and lytic reactivation","volume":"13","author":"T Murata","year":"2021","journal-title":"Viruses"},{"issue":"1","key":"pone.0345208.ref014","doi-asserted-by":"crossref","first-page":"165","DOI":"10.1128\/JVI.01512-10","article-title":"A new model of Epstein-Barr virus infection reveals an important role for early lytic viral protein expression in the development of lymphomas","volume":"85","author":"S-D Ma","year":"2011","journal-title":"J Virol"},{"issue":"11","key":"pone.0345208.ref015","doi-asserted-by":"crossref","first-page":"691","DOI":"10.1038\/s41579-019-0249-7","article-title":"Latency and lytic replication in Epstein-Barr virus-associated oncogenesis","volume":"17","author":"C M\u00fcnz","year":"2019","journal-title":"Nat Rev Microbiol"},{"issue":"7","key":"pone.0345208.ref016","doi-asserted-by":"crossref","first-page":"4415","DOI":"10.1128\/JVI.79.7.4415-4424.2005","article-title":"Epstein-Barr virus transforming protein LMP1 plays a critical role in virus production","volume":"79","author":"N Ahsan","year":"2005","journal-title":"J Virol"},{"issue":"2","key":"pone.0345208.ref017","doi-asserted-by":"crossref","first-page":"528","DOI":"10.1128\/jvi.53.2.528-535.1985","article-title":"Two related but differentially expressed potential membrane proteins encoded by the EcoRI Dhet region of Epstein-Barr virus B95-8","volume":"53","author":"GS Hudson","year":"1985","journal-title":"J Virol"},{"issue":"1","key":"pone.0345208.ref018","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1006\/viro.1997.8638","article-title":"Early detection of the lytic LMP-1 protein in EBV-infected B-cells suggests its presence in the virion","volume":"234","author":"KD Erickson","year":"1997","journal-title":"Virology"},{"issue":"5","key":"pone.0345208.ref019","doi-asserted-by":"crossref","first-page":"2637","DOI":"10.1128\/jvi.67.5.2637-2645.1993","article-title":"Biochemical, genetic, and functional analyses of the phosphorylation sites on the Epstein-Barr virus-encoded oncogenic latent membrane protein LMP-1","volume":"67","author":"RK Moorthy","year":"1993","journal-title":"J Virol"},{"issue":"26","key":"pone.0345208.ref020","doi-asserted-by":"crossref","first-page":"15595","DOI":"10.1073\/pnas.2136756100","article-title":"Epstein-Barr virus latent membrane protein 1 activation of NF-kappaB through IRAK1 and TRAF6","volume":"100","author":"M Luftig","year":"2003","journal-title":"Proc Natl Acad Sci U S A"},{"issue":"2","key":"pone.0345208.ref021","doi-asserted-by":"crossref","DOI":"10.3390\/cells10020239","article-title":"Aberrant splicing events and epigenetics in viral oncogenomics: current therapeutic strategies","volume":"10","author":"FZ Francies","year":"2021","journal-title":"Cells"},{"issue":"2","key":"pone.0345208.ref022","doi-asserted-by":"crossref","DOI":"10.1128\/JVI.01178-17","article-title":"c-Myc Represses Transcription of Epstein-Barr Virus Latent Membrane Protein 1 Early after Primary B Cell Infection","volume":"92","author":"AM Price","year":"2018","journal-title":"J Virol"},{"issue":"5","key":"pone.0345208.ref023","doi-asserted-by":"crossref","first-page":"506","DOI":"10.1016\/j.bbagrm.2014.03.013","article-title":"Myc and cell cycle control","volume":"1849","author":"G Bretones","year":"2015","journal-title":"Biochim Biophys Acta"},{"issue":"2","key":"pone.0345208.ref024","doi-asserted-by":"crossref","first-page":"178","DOI":"10.1002\/gcc.20063","article-title":"Level of MYC overexpression in pediatric Burkitt\u2019s lymphoma is strongly dependent on genomic breakpoint location within the MYC locus","volume":"41","author":"M Wilda","year":"2004","journal-title":"Genes Chromosomes Cancer"},{"issue":"11","key":"pone.0345208.ref025","doi-asserted-by":"crossref","DOI":"10.1016\/j.heliyon.2019.e02624","article-title":"The interplay between Epstein-Bar virus (EBV) with the p53 and its homologs during EBV associated malignancies","volume":"5","author":"K Chatterjee","year":"2019","journal-title":"Heliyon"},{"issue":"1","key":"pone.0345208.ref026","doi-asserted-by":"crossref","first-page":"25","DOI":"10.1016\/j.molcel.2005.02.029","article-title":"Structure of the p53 binding domain of HAUSP\/USP7 bound to Epstein-Barr nuclear antigen 1 implications for EBV-mediated immortalization","volume":"18","author":"V Saridakis","year":"2005","journal-title":"Mol Cell"},{"issue":"9","key":"pone.0345208.ref027","doi-asserted-by":"crossref","first-page":"2215","DOI":"10.1016\/j.celrep.2017.08.012","article-title":"The Functional Impact of Alternative Splicing in Cancer","volume":"20","author":"H Climente-Gonz\u00e1lez","year":"2017","journal-title":"Cell Rep"},{"issue":"1","key":"pone.0345208.ref028","doi-asserted-by":"crossref","first-page":"29","DOI":"10.1186\/s12985-019-1137-5","article-title":"The Epstein-Barr virus EBNA1 protein modulates the alternative splicing of cellular genes","volume":"16","author":"S Boudreault","year":"2019","journal-title":"Virol J"},{"issue":"1","key":"pone.0345208.ref029","doi-asserted-by":"crossref","first-page":"6381","DOI":"10.1038\/s41467-025-61870-3","article-title":"Comprehensive resolution and classification of the Epstein Barr virus transcriptome","volume":"16","author":"TD Nguyen","year":"2025","journal-title":"Nat Commun"},{"issue":"4","key":"pone.0345208.ref030","article-title":"Thermodynamic and structural characterization of an EBV infected B-cell lymphoma transcriptome","volume":"4","author":"CA O\u2019Leary","year":"2022","journal-title":"NAR Genom Bioinform"},{"issue":"4","key":"pone.0345208.ref031","article-title":"Intronic RNA secondary structural information captured for the human MYC pre-mRNA","volume":"6","author":"TO Eich","year":"2024","journal-title":"NAR Genom Bioinform"},{"key":"pone.0345208.ref032","doi-asserted-by":"crossref","DOI":"10.7717\/peerj.6136","article-title":"ScanFold: an approach for genome-wide discovery of local RNA structural elements-applications to Zika virus and HIV","volume":"6","author":"RJ Andrews","year":"2018","journal-title":"PeerJ"},{"key":"pone.0345208.ref033","doi-asserted-by":"crossref","first-page":"26","DOI":"10.1186\/1748-7188-6-26","article-title":"ViennaRNA Package 2.0","volume":"6","author":"R Lorenz","year":"2011","journal-title":"Algorithms in Molecular Biology"},{"issue":"17","key":"pone.0345208.ref034","doi-asserted-by":"crossref","first-page":"168549","DOI":"10.1016\/j.jmb.2024.168549","article-title":"NNDB: An Expanded Database of Nearest Neighbor Parameters for Predicting Stability of Nucleic Acid Secondary Structures","volume":"436","author":"A Mittal","year":"2024","journal-title":"J Mol Biol"},{"issue":"19","key":"pone.0345208.ref035","doi-asserted-by":"crossref","first-page":"7287","DOI":"10.1073\/pnas.0401799101","article-title":"Incorporating chemical modification constraints into a dynamic programming algorithm for prediction of RNA secondary structure","volume":"101","author":"DH Mathews","year":"2004","journal-title":"Proc Natl Acad Sci U S A"},{"issue":"4","key":"pone.0345208.ref036","doi-asserted-by":"crossref","first-page":"1531","DOI":"10.1128\/jvi.63.4.1531-1539.1989","article-title":"Influence of Burkitt\u2019s lymphoma and primary B cells on latent gene expression by the nonimmortalizing P3J-HR-1 strain of Epstein-Barr virus","volume":"63","author":"C Rooney","year":"1989","journal-title":"J Virol"},{"issue":"10","key":"pone.0345208.ref037","doi-asserted-by":"crossref","first-page":"4929","DOI":"10.1128\/JVI.75.10.4929-4935.2001","article-title":"Differential regulation of Epstein-Barr virus (EBV) latent gene expression in Burkitt lymphoma cells infected with a recombinant EBV strain","volume":"75","author":"P Trivedi","year":"2001","journal-title":"J Virol"},{"issue":"12","key":"pone.0345208.ref038","doi-asserted-by":"crossref","DOI":"10.1371\/journal.pone.0224953","article-title":"The spliceosome inhibitors isoginkgetin and pladienolide B induce ATF3-dependent cell death","volume":"15","author":"EJ Vanzyl","year":"2020","journal-title":"PLoS One"},{"key":"pone.0345208.ref039","doi-asserted-by":"crossref","first-page":"219","DOI":"10.1007\/978-1-0716-1158-6_13","article-title":"DMS-MaPseq for Genome-Wide or Targeted RNA Structure Probing In Vitro and In Vivo","volume":"2254","author":"P Tomezsko","year":"2021","journal-title":"Methods Mol Biol"},{"issue":"1","key":"pone.0345208.ref040","doi-asserted-by":"crossref","first-page":"75","DOI":"10.1038\/nmeth.4057","article-title":"DMS-MaPseq for genome-wide or targeted RNA structure probing in vivo","volume":"14","author":"M Zubradt","year":"2017","journal-title":"Nat Methods"},{"issue":"16","key":"pone.0345208.ref041","doi-asserted-by":"crossref","first-page":"2078","DOI":"10.1093\/bioinformatics\/btp352","article-title":"The Sequence Alignment\/Map format and SAMtools","volume":"25","author":"H Li","year":"2009","journal-title":"Bioinformatics"},{"issue":"16","key":"pone.0345208.ref042","doi-asserted-by":"crossref","DOI":"10.1093\/nar\/gky486","article-title":"RNA Framework: an all-in-one toolkit for the analysis of RNA structures and post-transcriptional modifications","volume":"46","author":"D Incarnato","year":"2018","journal-title":"Nucleic Acids Res"},{"issue":"14","key":"pone.0345208.ref043","doi-asserted-by":"crossref","DOI":"10.1093\/nar\/gkx533","article-title":"Structure-seq2: sensitive and accurate genome-wide profiling of RNA structure in vivo","volume":"45","author":"LE Ritchey","year":"2017","journal-title":"Nucleic Acids Res"},{"issue":"1","key":"pone.0345208.ref044","doi-asserted-by":"crossref","DOI":"10.1093\/bioinformatics\/btac830","article-title":"igv.js: an embeddable JavaScript implementation of the Integrative Genomics Viewer (IGV)","volume":"39","author":"JT Robinson","year":"2023","journal-title":"Bioinformatics"},{"issue":"3","key":"pone.0345208.ref045","doi-asserted-by":"crossref","first-page":"249","DOI":"10.1038\/s41592-021-01075-w","article-title":"Genome-scale deconvolution of RNA structure ensembles","volume":"18","author":"E Morandi","year":"2021","journal-title":"Nat Methods"},{"issue":"1","key":"pone.0345208.ref046","doi-asserted-by":"crossref","first-page":"97","DOI":"10.1073\/pnas.0806929106","article-title":"Accurate SHAPE-directed RNA structure determination","volume":"106","author":"KE Deigan","year":"2009","journal-title":"Proc Natl Acad Sci U S A"},{"key":"pone.0345208.ref047","doi-asserted-by":"crossref","first-page":"1105","DOI":"10.1002\/bip.360290621","article-title":"The equilibrium partition function and base pair binding probabilities for RNA secondary structure","volume":"29","author":"JS McCaskill","year":"1990","journal-title":"Biopolymers"},{"issue":"15","key":"pone.0345208.ref048","doi-asserted-by":"crossref","first-page":"1974","DOI":"10.1093\/bioinformatics\/btp250","article-title":"VARNA: Interactive drawing and editing of the RNA secondary structure","volume":"25","author":"K Darty","year":"2009","journal-title":"Bioinformatics"},{"key":"pone.0345208.ref049","article-title":"Introducing the Bacterial and Viral Bioinformatics Resource Center (BV-BRC): a resource combining PATRIC","volume":"51","author":"RD Olson","year":"2023","journal-title":"IRD and ViPR [Journal Article]. Nucleic Acids Res"},{"issue":"14","key":"pone.0345208.ref050","doi-asserted-by":"crossref","first-page":"3059","DOI":"10.1093\/nar\/gkf436","article-title":"MAFFT: a novel method for rapid multiple sequence alignment based on fast Fourier transform","volume":"30","author":"K Katoh","year":"2002","journal-title":"Nucleic Acids Res"},{"issue":"6","key":"pone.0345208.ref051","doi-asserted-by":"crossref","first-page":"1188","DOI":"10.1101\/gr.849004","article-title":"WebLogo: a sequence logo generator","volume":"14","author":"GE Crooks","year":"2004","journal-title":"Genome Res"},{"key":"pone.0345208.ref052","doi-asserted-by":"crossref","first-page":"527","DOI":"10.1007\/978-1-59745-514-5_33","article-title":"RNA consensus structure prediction with RNAalifold","volume":"395","author":"IL Hofacker","year":"2007","journal-title":"Methods Mol Biol"},{"key":"pone.0345208.ref053","doi-asserted-by":"crossref","first-page":"474","DOI":"10.1186\/1471-2105-9-474","article-title":"RNAalifold: improved consensus structure prediction for RNA alignments","volume":"9","author":"SH Bernhart","year":"2008","journal-title":"BMC Bioinformatics"},{"issue":"11","key":"pone.0345208.ref054","doi-asserted-by":"crossref","first-page":"2188","DOI":"10.1002\/ijc.35384","article-title":"Incorporation of Epstein-Barr viral variation implicates significance of Latent Membrane Protein 1 in survival prediction and prognostic subgrouping in Burkitt lymphoma","volume":"156","author":"IE Kim","year":"2025","journal-title":"Int J Cancer"},{"issue":"2","key":"pone.0345208.ref055","doi-asserted-by":"crossref","first-page":"893","DOI":"10.3390\/biom5020893","article-title":"RNA-Binding Proteins: Splicing Factors and Disease","volume":"5","author":"AM Fredericks","year":"2015","journal-title":"Biomolecules"}],"container-title":["PLOS One"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0345208","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,7,2]],"date-time":"2026-07-02T17:58:34Z","timestamp":1783015114000},"score":1,"resource":{"primary":{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0345208"}},"subtitle":[],"editor":[{"given":"Xiangning","family":"Zhang","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"editor"}]}],"short-title":[],"issued":{"date-parts":[[2026,7,2]]},"references-count":55,"journal-issue":{"issue":"7","published-online":{"date-parts":[[2026,7,2]]}},"URL":"https:\/\/doi.org\/10.1371\/journal.pone.0345208","relation":{},"ISSN":["1932-6203"],"issn-type":[{"value":"1932-6203","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,7,2]]}}}