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Validation strategies for antibodies targeting modified ribonucleotides

Chemical modifications are found on almost all RNAs and affect their coding and noncoding functions. The identification of m6A on mRNA and its important role in gene regulation stimulated the field to investigate whether additional modifications are present on mRNAs. Indeed, modifications including...

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Main Authors: Weichmann, Franziska (Author) , Dieterich, Christoph (Author) , Hett, Robert (Author) , Schepers, Aloys (Author) , Ito-Kureha, Taku (Author) , Flatley, Andrew (Author) , Slama, Kaouthar (Author) , Hastert, Florian D (Author) , Angstman, Nicholas B (Author) , Cardoso, M Cristina (Author) , König, Julian (Author) , Hüttelmaier, Stefan (Author) , Canzar, Stefan (Author) , Helm, Mark (Author) , Heissmeyer, Vigo (Author) , Feederle, Regina (Author) , Meister, Gunter (Author)
Format: Article (Journal)
Language:English
Published: July 7, 202
In: RNA
Year: 2020, Volume: 26, Issue: 10, Pages: 1489-1506
ISSN:1469-9001
DOI:10.1261/rna.076026.120
Online Access:Verlag, lizenzpflichtig, Volltext: https://doi.org/10.1261/rna.076026.120
Verlag, lizenzpflichtig, Volltext: http://rnajournal.cshlp.org/content/26/10/1489
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Author Notes:Franziska Weichmann, Robert Hett, Aloys Schepers, Taku Ito-Kureha, Andrew Flatley, Kaouthar Slama, Florian D. Hastert, Nicholas B. Angstman, M. Cristina Cardoso, Julian König, Stefan Hüttelmaier, Christoph Dieterich, Stefan Canzar, Mark Helm, Vigo Heissmeyer, Regina Feederle, and Gunter Meister
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Summary:Chemical modifications are found on almost all RNAs and affect their coding and noncoding functions. The identification of m6A on mRNA and its important role in gene regulation stimulated the field to investigate whether additional modifications are present on mRNAs. Indeed, modifications including m1A, m5C, m7G, 2′-OMe, and Ψ were detected. However, since their abundances are low and tools used for their corroboration are often not well characterized, their physiological relevance remains largely elusive. Antibodies targeting modified nucleotides are often used but have limitations such as low affinity or specificity. Moreover, they are not always well characterized and due to the low abundance of the modification, particularly on mRNAs, generated data sets might resemble noise rather than specific modification patterns. Therefore, it is critical that the affinity and specificity is rigorously tested using complementary approaches. Here, we provide an experimental toolbox that allows for testing antibody performance prior to their use.
Item Description:Gesehen am 22.10.2020
Physical Description:Online Resource
ISSN:1469-9001
DOI:10.1261/rna.076026.120

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