Quantitative profiling of pseudouridylation dynamics in native RNAs with nanopore sequencing

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-methylatio...

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Detalhes bibliográficos
Autores: Begik, Oguzhan, Lucas, Morghan C., Pryszcz, Leszek Piotr, 1985-, Ramirez, Jose Miguel, Medina, Rebeca, Milenkovic, Ivan, Cruciani, Sonia, Liu, Huanle, Santos Vieira, Helaine Graziele, Sas Chen, Aldema, Mattick, John S., Schwartz, Schraga, Novoa, Eva Maria
Tipo de documento: artigo
Estado:Versión aceptada para publicación
Data de publicação:2021
País:España
Recursos:Varias* (Consorci de Biblioteques Universitáries de Catalunya, Centre de Serveis Científics i Acadèmics de Catalunya)
Repositório:Recercat. Dipósit de la Recerca de Catalunya
OAI Identifier:oai:recercat.cat:10230/48205
Acesso em linha:http://hdl.handle.net/10230/48205
http://dx.doi.org/10.1038/s41587-021-00915-6
Access Level:Acceso aberto
Palavra-chave:RNA modification
RNA sequencing
Software
Descrição
Resumo: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.

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