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Nucleic Acids Res. 2019 Sep 2. pii: gkz751. doi: 10.1093/nar/gkz751. [Epub ahead of print]

Quantifying the RNA cap epitranscriptome reveals novel caps in cellular and viral RNA.

Author information

1
State Key Laboratory of Reproductive Regulation and Breeding of Grassland Livestock, Inner Mongolia University, Hohhot, People's Republic of China.
2
School of Life Sciences, Inner Mongolia University, Hohhot, People's Republic of China.
3
Antimicrobial Resistance Interdisciplinary Research Group, Singapore-MIT Alliance for Research and Technology, Singapore.
4
Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore.
5
NTU Institute of Health Technologies, Interdisciplinary Graduate Programme, Nanyang Technological University, Singapore.
6
Shanghai Blueray Biopharma, Shanghai, People's Republic of China.
7
Cancer Science Institute of Singapore, Singapore.
8
School of Biological Sciences, Nanyang Technological University, Singapore.
9
Department of Microbiology, National University of Singapore, Singapore.
10
Department of Biology, Saint Louis University, St. Louis, MO, USA.
11
Departments of Biochemistry & Molecular Biology and Pharmacology & Toxicology, and Sealy Center for Structural Biology & Molecular Biophysics, University of Texas Medical Branch, Galveston, TX, USA.
12
Synthetic Biology Center, Departments of Biological Engineering and Electrical Engineering and Computer Science, Massachusetts Institute of Technology, Cambridge, MA, USA.
13
Department of Pharmacology, Weill Medical College, Cornell University, New York, NY, USA.
14
Dept. of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA.

Abstract

Chemical modification of transcripts with 5' caps occurs in all organisms. Here, we report a systems-level mass spectrometry-based technique, CapQuant, for quantitative analysis of an organism's cap epitranscriptome. The method was piloted with 21 canonical caps-m7GpppN, m7GpppNm, GpppN, GpppNm, and m2,2,7GpppG-and 5 'metabolite' caps-NAD, FAD, UDP-Glc, UDP-GlcNAc, and dpCoA. Applying CapQuant to RNA from purified dengue virus, Escherichia coli, yeast, mouse tissues, and human cells, we discovered new cap structures in humans and mice (FAD, UDP-Glc, UDP-GlcNAc, and m7Gpppm6A), cell- and tissue-specific variations in cap methylation, and high proportions of caps lacking 2'-O-methylation (m7Gpppm6A in mammals, m7GpppA in dengue virus). While substantial Dimroth-induced loss of m1A and m1Am arose with specific RNA processing conditions, human lymphoblast cells showed no detectable m1A or m1Am in caps. CapQuant accurately captured the preference for purine nucleotides at eukaryotic transcription start sites and the correlation between metabolite levels and metabolite caps.

PMID:
31504804
DOI:
10.1093/nar/gkz751

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