The role of sequence context, nucleotide pool balance and stress in 2′-deoxynucleotide misincorporation in viral, bacterial and mammalian RNA
Name
Wang-2016-The role of sequence.pdf
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2.06 MB
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Author(s)
Wang, Jin
Dong, Hongping
Chionh, Yok Hian
McBee, Megan E.
Sirirungruang, Sasilada
Cunningham, Richard P.
Shi, Pei-Yong
Dedon, Peter C
Date Issued
June 2016
Journal
Nucleic Acids Research
Publisher
Oxford University Press
Citation
Wang, Jin et al. “The Role of Sequence Context, Nucleotide Pool Balance and Stress in 2′-Deoxynucleotide Misincorporation in Viral, Bacterial and Mammalian RNA.” Nucleic Acids Research 44.18 (2016): 8962–8975.
Version
Final published version
Abstract
The misincorporation of 2′-deoxyribonucleotides (dNs) into RNA has important implications for the function of non-coding RNAs, the translational fidelity of coding RNAs and the mutagenic evolution of viral RNA genomes. However, quantitative appreciation for the degree to which dN misincorporation occurs is limited by the lack of analytical tools. Here, we report a method to hydrolyze RNA to release 2′-deoxyribonucleotide-ribonucleotide pairs (dNrN) that are then quantified by chromatography-coupled mass spectrometry (LC-MS). Using this platform, we found misincorporated dNs occurring at 1 per 10[superscript 3] to 10[superscript 5] ribonucleotide (nt) in mRNA, rRNAs and tRNA in human cells, Escherichia coli, Saccharomyces cerevisiae and, most abundantly, in the RNA genome of dengue virus. The frequency of dNs varied widely among organisms and sequence contexts, and partly reflected the in vitro discrimination efficiencies of different RNA polymerases against 2′-deoxyribonucleoside 5′-triphosphates (dNTPs). Further, we demonstrate a strong link between dN frequencies in RNA and the balance of dNTPs and ribonucleoside 5′-triphosphates (rNTPs) in the cellular pool, with significant stress-induced variation of dN incorporation. Potential implications of dNs in RNA are discussed, including the possibilities of dN incorporation in RNA as a contributing factor in viral evolution and human disease, and as a host immune defense mechanism against viral infections.
MIT Department
Massachusetts Institute of Technology. Center for Environmental Health Sciences
Massachusetts Institute of Technology. Department of Biological Engineering
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Creative Commons Attribution 4.0 International License
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DOI of Published Version
https://doi.org/10.1093/nar/gkw572
