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Series GSE109298 Query DataSets for GSE109298
Status Public on Feb 21, 2018
Title Enzymatic or in vivo installation of propargyl groups in combination with click chemistry enables enrichment and detection of methyltransferase target sites in RNA
Organisms Homo sapiens; synthetic construct
Experiment type Other
Summary m6A is the most abundant internal modification in eukaryotic mRNA. It is introduced by METTL3-METTL14 and tunes mRNA metabolism, impacting cell differentiation and development. Precise transcriptome-wide assignment of m6A sites is of utmost importance. However, m6A does not interfere with Watson-Crick base pairing making polymerase-based detection challenging. We developed a chemical biology approach for the precise mapping of methyltransferase (MTase) target sites based on the introduction of a bioorthogonal propargyl group in vitro and in cells. We show that propargyl can be introduced enzymatically by wild-type METTL3-METTL14. Reverse transcription terminated up to 65 % at m6A sites after bioconjugation and purification, hence enabling detection of METTL3-METTL14 target sites by next generation sequencing. Importantly, we implemented metabolic propargyl labeling of RNA MTase target sites in vivo based on propargyl-L-selenohomocysteine and validated different types of known rRNA methylation sites.
 
Overall design enrichment of methylated nucleotides by bioconjugation of propagyl groups in synthetic oligos and rRNA
 
Contributor(s) Hartstock K, Nilges BS, Ovcharenko A, Cornelissen N, Püllen N, Lawrence-Dörner A, Leidel SA, Rentmeister A
Citation(s) 29461645
Submission date Jan 17, 2018
Last update date Dec 28, 2018
Contact name Benedikt Sebastian Nilges
E-mail(s) benedikt.nilges@mpi-muenster.mpg.de
Organization name Max Planck Institute for Molecular Biomedicine
Department RNA Biology Group
Street address Von-Esmarch-Straße 54
City Muenster
ZIP/Postal code 48149
Country Germany
 
Platforms (2)
GPL15456 Illumina HiScanSQ (Homo sapiens)
GPL24519 Illumina HiScanSQ (synthetic construct)
Samples (19)
GSM2936678 1d_37T_propagylated
GSM2936679 0.1d_37T_propagylated
GSM2936680 0.01d_37T_propagylated
Relations
BioProject PRJNA430372
SRA SRP131073

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Supplementary file Size Download File type/resource
GSE109298_RAW.tar 43.6 Mb (http)(custom) TAR (of BED)
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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