dc.contributor.author |
Lucas, Morghan C. |
dc.contributor.author |
Pryszcz, Leszek Piotr, 1985- |
dc.contributor.author |
Medina, Rebeca |
dc.contributor.author |
Milenkovic, Ivan |
dc.contributor.author |
Camacho, Noelia |
dc.contributor.author |
Marchand, Virginie |
dc.contributor.author |
Motorin, Yuri |
dc.contributor.author |
Ribas de Pouplana, Lluís |
dc.contributor.author |
Novoa, Eva Maria |
dc.date.accessioned |
2023-06-19T06:05:42Z |
dc.date.available |
2023-06-19T06:05:42Z |
dc.date.issued |
2024 |
dc.identifier.citation |
Lucas MC, Pryszcz LP, Medina R, Milenkovic I, Camacho N, Marchand V, et al. Quantitative analysis of tRNA abundance and modifications by nanopore RNA sequencing. Nat Biotechnol. 2024 Jan;42(1):72-86. DOI: 10.1038/s41587-023-01743-6 |
dc.identifier.issn |
1087-0156 |
dc.identifier.uri |
http://hdl.handle.net/10230/57222 |
dc.description.abstract |
Transfer RNAs (tRNAs) play a central role in protein translation. Studying them has been difficult in part because a simple method to simultaneously quantify their abundance and chemical modifications is lacking. Here we introduce Nano-tRNAseq, a nanopore-based approach to sequence native tRNA populations that provides quantitative estimates of both tRNA abundances and modification dynamics in a single experiment. We show that default nanopore sequencing settings discard the vast majority of tRNA reads, leading to poor sequencing yields and biased representations of tRNA abundances based on their transcript length. Re-processing of raw nanopore current intensity signals leads to a 12-fold increase in the number of recovered tRNA reads and enables recapitulation of accurate tRNA abundances. We then apply Nano-tRNAseq to Saccharomyces cerevisiae tRNA populations, revealing crosstalks and interdependencies between different tRNA modification types within the same molecule and changes in tRNA populations in response to oxidative stress. |
dc.description.sponsorship |
We thank all the members of the Novoa laboratory for their valuable insights and discussions. We thank V. Malhotra for providing us with the S. cerevisiae Pus4 KO strains used in this work. We thank the CRG Protein Technologies Unit for producing recombinant E. coli T4 RNA Ligase 2. We thank A. Delgado-Tejedor for help setting up simulations on bulk data from S. cerevisiae Pus1 and Pus7 KO Nano-tRNAseq runs. M.C.L. is supported by an FPI Severo Ochoa fellowship by the Spanish Ministry of Economy, Industry, and Competitiveness (MEIC). L.P.P. is supported by funding from the European Union’s H2020 Research and Innovation Programme under Marie Sklodowska-Curie grant agreement number 754422. I.M. is supported by ‘la Caixa’ InPhINIT PhD fellowship (LCF/BQ/DI18/11660028). This work was supported by funds from the Spanish Ministry of Economy, Industry, and Competitiveness (MEIC) (PID2021-128193NB-100 to E.M.N.). This project has received funding from the ERCEA program (ERC-StG-2021 under grant agreement number 101042103 to E.M.N.). We acknowledge the support of the MEIC to the EMBL partnership, Centro de Excelencia Severo Ochoa and CERCA Programme/Generalitat de Catalunya. |
dc.format.mimetype |
application/pdf |
dc.language.iso |
eng |
dc.publisher |
Nature Research |
dc.relation.ispartof |
Nat Biotechnol. 2024 Jan;42(1):72-86 |
dc.rights |
© The Author(s) 2023. Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. |
dc.rights.uri |
http://creativecommons.org/licenses/by/4.0/ |
dc.title |
Quantitative analysis of tRNA abundance and modifications by nanopore RNA sequencing |
dc.type |
info:eu-repo/semantics/article |
dc.identifier.doi |
http://dx.doi.org/10.1038/s41587-023-01743-6 |
dc.subject.keyword |
RNA sequencing |
dc.subject.keyword |
Small RNAs |
dc.relation.projectID |
info:eu-repo/grantAgreement/EC/H2020/754422 |
dc.relation.projectID |
info:eu-repo/grantAgreement/ES/3PE/PID2021-128193NB-100 |
dc.rights.accessRights |
info:eu-repo/semantics/openAccess |
dc.type.version |
info:eu-repo/semantics/publishedVersion |