Quantitative profiling of pseudouridylation dynamics in native RNAs with nanopore sequencing
Top Cited Papers
- 13 May 2021
- journal article
- research article
- Published by Springer Science and Business Media LLC in Nature Biotechnology
- Vol. 39 (10), 1278-1291
- https://doi.org/10.1038/s41587-021-00915-6
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.Keywords
This publication has 97 references indexed in Scilit:
- Comprehensive Analysis of mRNA Methylation Reveals Enrichment in 3′ UTRs and near Stop CodonsCell, 2012
- The SNORD115 (H/MBII-52) and SNORD116 (H/MBII-85) gene clusters at the imprinted Prader-Willi locus generate canonical box C/D snoRNAsNucleic Acids Research, 2012
- rRNA Pseudouridylation Defects Affect Ribosomal Ligand Binding and Translational Fidelity from Yeast to Human CellsMolecular Cell, 2011
- Requirement of rRNA Methylation for 80S Ribosome Assembly on a Cohort of Cellular Internal Ribosome Entry SitesMolecular and Cellular Biology, 2011
- Loss of ribosomal RNA modification causes developmental defects in zebrafishNucleic Acids Research, 2011
- Integrative genomics viewerNature Biotechnology, 2011
- U2 snRNA is inducibly pseudouridylated at novel sites by Pus7p and snR81 RNPThe EMBO Journal, 2010
- The Sequence Alignment/Map format and SAMtoolsBioinformatics, 2009
- A deletion of the HBII-85 class of small nucleolar RNAs (snoRNAs) is associated with hyperphagia, obesity and hypogonadismHuman Molecular Genetics, 2009
- Prader-Willi phenotype caused by paternal deficiency for the HBII-85 C/D box small nucleolar RNA clusterNature Genetics, 2008