Quantitative profiling of pseudouridylation dynamics in native RNAs with nanopore sequencing

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• dc.contributor.author Begik, Oguzhan
• dc.contributor.author Lucas, Morghan C.
• dc.contributor.author Pryszcz, Leszek Piotr, 1985-
• dc.contributor.author Ramirez, Jose Miguel
• dc.contributor.author Medina, Rebeca
• dc.contributor.author Milenkovic, Ivan
• dc.contributor.author Cruciani, Sonia
• dc.contributor.author Liu, Huanle
• dc.contributor.author Santos Vieira, Helaine Graziele
• dc.contributor.author Sas Chen, Aldema
• dc.contributor.author Mattick, John S.
• dc.contributor.author Schwartz, Schraga
• dc.contributor.author Novoa, Eva Maria
• dc.date.accessioned 2021-07-16T07:14:23Z
• dc.date.issued 2021
• dc.description.abstract Nanopore RNA sequencing shows promise as a method for discriminating and identifying different RNA modifications in native RNA. Expanding on the ability of nanopore sequencing to detect N6-methyladenosine, we show that other modifications, in particular pseudouridine (Ψ) and 2'-O-methylation (Nm), also result in characteristic base-calling 'error' signatures in the nanopore data. Focusing on Ψ modification sites, we detected known and uncovered previously unreported Ψ sites in mRNAs, non-coding RNAs and rRNAs, including a Pus4-dependent Ψ modification in yeast mitochondrial rRNA. To explore the dynamics of pseudouridylation, we treated yeast cells with oxidative, cold and heat stresses and detected heat-sensitive Ψ-modified sites in small nuclear RNAs, small nucleolar RNAs and mRNAs. Finally, we developed a software, nanoRMS, that estimates per-site modification stoichiometries by identifying single-molecule reads with altered current intensity and trace profiles. This work demonstrates that Nm and Ψ RNA modifications can be detected in cellular RNAs and that their modification stoichiometry can be quantified by nanopore sequencing of native RNA.
• dc.description.sponsorship This project has received funding from the European Union’s Horizon 2020 Research and Innovation Programme under Marie Skodowska-Curie grant agreement no. 713673. This work was supported by the Australian Research Council (DP180103571 to E.M.N.) and the MEIC (PGC2018-098152-A-100 to E.M.N.). We acknowledge the support of the MEIC to the EMBL partnership, the Centro de Excelencia Severo Ochoa and the CERCA Programme/Generalitat de Catalunya.
• dc.format.mimetype application/pdf
• dc.identifier.citation Begik O, Lucas MC, Pryszcz LP, Ramirez JM, Medina R, Milenkovic I, Cruciani S, Liu H, Vieira HGS, Sas-Chen A, Mattick JS, Schwartz S, Novoa EM. Quantitative profiling of pseudouridylation dynamics in native RNAs with nanopore sequencing. Nat Biotechnol. 2021;39:1278-91. DOI: 10.1038/s41587-021-00915-6
• dc.identifier.doi http://dx.doi.org/10.1038/s41587-021-00915-6
• dc.identifier.issn 1087-0156
• dc.identifier.uri http://hdl.handle.net/10230/48205
• dc.language.iso eng
• dc.publisher Nature Research
• dc.relation.ispartof Nat Biotechnol. 2021;39:1278-91.
• dc.relation.projectID info:eu-repo/grantAgreement/EC/H2020/713673
• dc.relation.projectID info:eu-repo/grantAgreement/ES/2PE/PGC2018-098152-A-100
• dc.rights © Springer Nature Publishing AG Begik O, Lucas MC, Pryszcz LP, Ramirez JM, Medina R, Milenkovic I, Cruciani S, Liu H, Vieira HGS, Sas-Chen A, Mattick JS, Schwartz S, Novoa EM. Quantitative profiling of pseudouridylation dynamics in native RNAs with nanopore sequencing. Nat Biotechnol. 2021;39:1278-91. DOI: 10.1038/s41587-021-00915-6 [http://dx.doi.org/10.1038/s41587-021-00915-6]
• dc.rights.accessRights info:eu-repo/semantics/openAccess
• dc.subject.keyword RNA modification
• dc.subject.keyword RNA sequencing
• dc.subject.keyword Software
• dc.title Quantitative profiling of pseudouridylation dynamics in native RNAs with nanopore sequencing
• dc.type info:eu-repo/semantics/article
• dc.type.version info:eu-repo/semantics/acceptedVersion