The Transcriptome-Wide Mapping of 2-Methylthio-N6-isopentenyladenosine at Single-Base Resolution.
J Am Chem Soc
; 145(9): 5467-5473, 2023 03 08.
Article
em En
| MEDLINE
| ID: mdl-36820840
ABSTRACT
Hundreds of modified bases have been identified and enzymatically modified to transfer RNAs (tRNAs) to regulate RNA function in various organisms. 2-Methylthio-N6-isopentenyladenosine (ms2i6A), a hypermodified base found at tRNA position 37, exists in both prokaryotes and eukaryotes. ms2i6A is traditionally identified by separating and digesting each tRNA from total RNA using RNA mass spectrometry. A transcriptome-wide and single-base resolution method that enables absolute mapping of ms2i6A along with analysis of its distribution in different RNAs is lacking. Here, through chemoselective methylthio group bioconjugation, we introduce a new approach (redox activated chemical tagging sequencing, ReACT-seq) to detect ms2i6A transcriptome-wide at single-base resolution. Using the chemoselectivity between the methylthio group and oxaziridine group, ms2i6A is bio-orthogonally tagged with an azide group without interference of canonical nucleotides, advancing enrichment of methylthio group modified RNAs prior to sequencing. ReACT-seq was demonstrated on nine known tRNAs and proved to be highly accurate, and the reverse transcription stop (RT-stop) character enables ReACT-seq detection at single-base resolution. In addition, ReACT-seq identified that the modification of ms2i6A is conservative and may not exist in other RNAs.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Transcriptoma
/
Isopenteniladenosina
Idioma:
En
Ano de publicação:
2023
Tipo de documento:
Article