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Strategies to Avoid Artifacts in Mass Spectrometry‐Based Epitranscriptome Analyses
In this report, we perform structure validation of recently reported RNA phosphorothioate (PT) modifications, a new set of epitranscriptome marks found in bacteria and eukaryotes including humans. By comparing synthetic PT‐containing diribonucleotides with native species in RNA hydrolysates by high‐...
Autores principales: | , , , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
John Wiley and Sons Inc.
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8597057/ https://www.ncbi.nlm.nih.gov/pubmed/34339593 http://dx.doi.org/10.1002/anie.202106215 |
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author | Kaiser, Steffen Byrne, Shane R. Ammann, Gregor Asadi Atoi, Paria Borland, Kayla Brecheisen, Roland DeMott, Michael S. Gehrke, Tim Hagelskamp, Felix Heiss, Matthias Yoluç, Yasemin Liu, Lili Zhang, Qinghua Dedon, Peter C. Cao, Bo Kellner, Stefanie |
author_facet | Kaiser, Steffen Byrne, Shane R. Ammann, Gregor Asadi Atoi, Paria Borland, Kayla Brecheisen, Roland DeMott, Michael S. Gehrke, Tim Hagelskamp, Felix Heiss, Matthias Yoluç, Yasemin Liu, Lili Zhang, Qinghua Dedon, Peter C. Cao, Bo Kellner, Stefanie |
author_sort | Kaiser, Steffen |
collection | PubMed |
description | In this report, we perform structure validation of recently reported RNA phosphorothioate (PT) modifications, a new set of epitranscriptome marks found in bacteria and eukaryotes including humans. By comparing synthetic PT‐containing diribonucleotides with native species in RNA hydrolysates by high‐resolution mass spectrometry (MS), metabolic stable isotope labeling, and PT‐specific iodine‐desulfurization, we disprove the existence of PTs in RNA from E. coli, S. cerevisiae, human cell lines, and mouse brain. Furthermore, we discuss how an MS artifact led to the initial misidentification of 2′‐O‐methylated diribonucleotides as RNA phosphorothioates. To aid structure validation of new nucleic acid modifications, we present a detailed guideline for MS analysis of RNA hydrolysates, emphasizing how the chosen RNA hydrolysis protocol can be a decisive factor in discovering and quantifying RNA modifications in biological samples. |
format | Online Article Text |
id | pubmed-8597057 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | John Wiley and Sons Inc. |
record_format | MEDLINE/PubMed |
spelling | pubmed-85970572021-11-22 Strategies to Avoid Artifacts in Mass Spectrometry‐Based Epitranscriptome Analyses Kaiser, Steffen Byrne, Shane R. Ammann, Gregor Asadi Atoi, Paria Borland, Kayla Brecheisen, Roland DeMott, Michael S. Gehrke, Tim Hagelskamp, Felix Heiss, Matthias Yoluç, Yasemin Liu, Lili Zhang, Qinghua Dedon, Peter C. Cao, Bo Kellner, Stefanie Angew Chem Int Ed Engl Research Articles In this report, we perform structure validation of recently reported RNA phosphorothioate (PT) modifications, a new set of epitranscriptome marks found in bacteria and eukaryotes including humans. By comparing synthetic PT‐containing diribonucleotides with native species in RNA hydrolysates by high‐resolution mass spectrometry (MS), metabolic stable isotope labeling, and PT‐specific iodine‐desulfurization, we disprove the existence of PTs in RNA from E. coli, S. cerevisiae, human cell lines, and mouse brain. Furthermore, we discuss how an MS artifact led to the initial misidentification of 2′‐O‐methylated diribonucleotides as RNA phosphorothioates. To aid structure validation of new nucleic acid modifications, we present a detailed guideline for MS analysis of RNA hydrolysates, emphasizing how the chosen RNA hydrolysis protocol can be a decisive factor in discovering and quantifying RNA modifications in biological samples. John Wiley and Sons Inc. 2021-09-29 2021-10-25 /pmc/articles/PMC8597057/ /pubmed/34339593 http://dx.doi.org/10.1002/anie.202106215 Text en © 2021 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH https://creativecommons.org/licenses/by/4.0/This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Articles Kaiser, Steffen Byrne, Shane R. Ammann, Gregor Asadi Atoi, Paria Borland, Kayla Brecheisen, Roland DeMott, Michael S. Gehrke, Tim Hagelskamp, Felix Heiss, Matthias Yoluç, Yasemin Liu, Lili Zhang, Qinghua Dedon, Peter C. Cao, Bo Kellner, Stefanie Strategies to Avoid Artifacts in Mass Spectrometry‐Based Epitranscriptome Analyses |
title | Strategies to Avoid Artifacts in Mass Spectrometry‐Based Epitranscriptome Analyses |
title_full | Strategies to Avoid Artifacts in Mass Spectrometry‐Based Epitranscriptome Analyses |
title_fullStr | Strategies to Avoid Artifacts in Mass Spectrometry‐Based Epitranscriptome Analyses |
title_full_unstemmed | Strategies to Avoid Artifacts in Mass Spectrometry‐Based Epitranscriptome Analyses |
title_short | Strategies to Avoid Artifacts in Mass Spectrometry‐Based Epitranscriptome Analyses |
title_sort | strategies to avoid artifacts in mass spectrometry‐based epitranscriptome analyses |
topic | Research Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8597057/ https://www.ncbi.nlm.nih.gov/pubmed/34339593 http://dx.doi.org/10.1002/anie.202106215 |
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