TY  - JOUR
A1  - Kaiser, Steffen
A1  - Byrne, Shane R.
A1  - Ammann, Gregor
A1  - Asadi-Atoi, Paria
A1  - Borland, Kayla
A1  - Brecheisen, Roland
A1  - DeMott, Michael S.
A1  - Gehrke, Tim Herbert
A1  - Hagelskamp, Felix
A1  - Heiß, Matthias
A1  - Yoluç, Yasemin
A1  - Liu, Lili
A1  - Zhang, Qinghua
A1  - Dedon, Peter C.
A1  - Cao, Bo
A1  - Kellner, Stefanie
T1  - Strategies to avoid artifacts in mass spectrometry-based epitranscriptome analyses
T2  - Angewandte Chemie
N2  - 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.
KW  - digestion artifact
KW  - mass spectrometry
KW  - nucleoside analysis
KW  - RNA modification
KW  - RNA PT
Y1  - 2021
UR  - http://publikationen.ub.uni-frankfurt.de/frontdoor/index/index/docId/63940
UR  - https://nbn-resolving.org/urn:nbn:de:hebis:30:3-639403
SN  - 1521-3773
N1  - This work was supported by Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) grants 255344185-SPP 1784, 325871075-SFB 1309, and KE1943/3-1 (SK), US NIH NIEHS Core Center Grant ES0002109 (CEHS Bioanalytical Core), a grant from the National Research Foundation of Singapore in support of the Antimicrobial Resistance Interdisciplinary Research Group (PD), and by National Natural Science Foundation of China grant 32070629 (CB). Shane Byrne acknowledges support from NIEHS Training Grant in Environmental Toxicology T32-ES007020. Open Access funding enabled and organized by Projekt DEAL.
VL  - 60
IS  - 44
SP  - 23885
EP  - 23893
PB  - Wiley-VCH
CY  - Weinheim
ER  -