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- Title
Quantitative profiling of pseudouridylation dynamics in native RNAs with nanopore sequencing.
- Authors
Begik, Oguzhan; Lucas, Morghan C.; Pryszcz, Leszek P.; Ramirez, Jose Miguel; Medina, Rebeca; Milenkovic, Ivan; Cruciani, Sonia; Liu, Huanle; Vieira, Helaine Graziele Santos; Sas-Chen, Aldema; Mattick, John S.; Schwartz, Schraga; Novoa, Eva Maria
- 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. Nanopore sequencing detects pseudouridine and 2′-O-methylation modifications in cellular RNAs.
- Publication
Nature Biotechnology, 2021, Vol 39, Issue 10, p1278
- ISSN
1087-0156
- Publication type
Article
- DOI
10.1038/s41587-021-00915-6