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Transcriptome-Wide 2’-O-Methylation Mapping at Single-base Resolution by Nm-REP-seq

ABSTRACT: Transcriptome-Wide 2’-O-Methylation Mapping at Single-base Resolution by Nm-REP-seq

PROVIDER: PRJNA663360 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR12639600_1.fastq.gz	Fastqsanger.gz
	SRR12639600_2.fastq.gz	Fastqsanger.gz
	SRR12639601_1.fastq.gz	Fastqsanger.gz
	SRR12639601_2.fastq.gz	Fastqsanger.gz
	SRR12639602_1.fastq.gz	Fastqsanger.gz

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Transcriptome-Wide 2’-O-Methylation Mapping at Single-base Resolution by Nm-REP-seq

Project description:2’-O-methylation (Nm) is one of the most abundant RNA epigenetic modification and plays vital roles in the post-transcriptional regulation of gene expression. Current Nm mapping approaches are normally limited to highly abundant RNAs and have significant technical hurdles in mRNAs or relatively rare non-coding RNAs (ncRNAs). Here, we developed a new method for enriching Nm sites by using RNA exoribonuclease (M. genitalium RNase R, MgR) and periodate oxidation reactivity to eliminate 2’-hydroxylated (2’-OH) nucleosides, coupled with sequencing (Nm-REP-seq). We revealed several novel classes of Nm-containing ncRNAs as well as mRNAs in humans, mice, and drosophila. We found that some novel Nm sites are present at fixed positions in different tRNAs and are potential substrates of fibrillarin (FBL) methyltransferase mediated by snoRNAs. Importantly, we discovered, for the first time, that Nm located at the 3’-end of various types of ncRNAs and fragments derived from them. Our approach precisely redefines the genome-wide distribution of Nm and provides new technologies for functional studies of Nm-mediated gene regulation.

2021-03-25 | GSE157930 | GEO

Nm-Mut-seq: The base-resolution quantitative method for mapping transcriptome-wide 2’-O-methylations

Project description:2’-O-methylation (Nm) is a prevalent post-transcriptional RNA modification present in many cellular RNAs and plays a critical role in modulating both the physical properties and regulation of eukaryotic RNAs. Studies of Nm modifications in RNA have long been hampered by a lack of effective mapping methods. Previously reported approaches can work well for detecting Nm modifications on abundant RNAs, but face challenges when applied to low-abundant RNAs, such as mRNA, lack stoichiometric information, and are challenged by issues of RNA sample degradation due to chemical treatment. Here, we present Nm-Mut-seq, a mutation signature-based Nm mapping method, which uses a custom reverse transcriptase (RT) that installs mutations at Am, Cm, and Gm-modified sites (Um is undetectable by this method). Our work provides a much-needed approach to detect Nm at base resolution in low abundant RNAs and to estimate the stoichiometry of each modified site transcriptome-wide.

2023-06-01 | GSE174518 | GEO

Nm-REP-seq Reveals 2’-O-Methylation Landscape at the 3'-end of Noncoding RNAs with Single-base Resolution

Project description:Many non-coding RNAs (ncRNAs), such as piRNAs, miRNAs and siRNAs, contain Nm at the 3'-end nucleotides, protecting the 3’-ends form terminal nucleotide addition or exonucleolytic degradation, but whether other kinds of RNAs contain a 3'-terminal Nm remain unknown. Here, we developed Nm-REP-seq to globally profile the 3'end-Nm sites and we revealed several novel classes of Nm-containing ncRNAs in mice and drosophila. Importantly, we discovered that Nm distribute at the end of miRNA, snoRNA, snRNA, tRNA as well as fragments derived from them. Collectively, our approach precisely redefines the genome-wide distribution of Nm and provides new technologies for functional studies of Nm-mediated gene regulation. Method: We developed Nm-REP-seq to globally profile the 3'-end Nm sites. Results: We discovered that Nm distribute at the end of miRNA, snoRNA, snRNA, tRNA as well as fragments derived from them. Conclusion:Collectively, our approach precisely redefines the genome-wide distribution of Nm and provides new technologies for functional studies of Nm-mediated gene regulation.

2022-10-25 | GSE198748 | GEO

Nm-Mut-seq: The base-resolution quantitative method for mapping transcriptome-wide 2’-O-methylations

Project description:Nm-Mut-seq: The base-resolution quantitative method for mapping transcriptome-wide 2’-O-methylations

| PRJNA730412 | ENA

Nm-REP-seq Reveals 2’-O-Methylation Landscape at the 3'-end of Noncoding RNAs with Single-base Resolution

Project description:Nm-REP-seq Reveals 2’-O-Methylation Landscape at the 3'-end of Noncoding RNAs with Single-base Resolution

| PRJNA816727 | ENA

Nm-seq finds thousands of modified 2’-O-methylation sites in mRNA with base precision

Project description:Nm-seq maps 2'-O-methylation sites in human mRNA with base precision The ribose of rna nucleotides can be 2′-O-methylated (nm). despite advances in high-throughput detection, the inert chemical nature of nm still limits sensitivity and precludes mapping in mrna. We leveraged the differential reactivity of 2′-O-methylated and 2′-hydroxylated nucleosides to periodate oxidation to develop nm-seq, a sensitive method for transcriptome-wide mapping of nm with base precision. nm-seq uncovered thousands of nm sites in human mrna with features suggesting functional roles.

2017-05-15 | GSE90164 | GEO

High-throughput single-base resolution mapping of 2’-O-Methylated residues

Project description:Functional characterization of the transcriptome requires tools for the systematic investigation of RNA post-transcriptional modifications. 2’-O-Methylation of the ribose moiety is one of the most abundant post-transcriptional modifications of the RNA. We describe here a high-throughput method that enables fast and accurate mapping at single-base resolution, and quantitation, of 2’-OMe modified residues. Our approach expands the actual repertoire of methods for transcriptome-wide mapping of RNA post-transcriptional modifications.

2016-09-13 | GSE73065 | GEO

Epigenome characterization at single base-pair resolution

Project description:We have combined standard micrococcal (MNase) digestion of nuclei with a modified protocol for construction paired-end DNA sequencing libraries to map both nucleosomes and subnucleosome-sized particles at single base-pair resolution throughout the budding yeast genome. We found that partially unwrapped nucleosomes and subnucleosome-sized particles can occupy the same position within a cell population, suggesting dynamic behavior. By varying the time of MNase digestion, we have been able to observe changes that reflect differential sensitivity of particles, including eviction of nucleosomes. Our protocol and mapping method provide a general strategy for characterizing full epigenomes. We used micrococcal nuclease mapping, chromatin immunoprecipitation and paired-end sequencing to determine the structure of yeast centromeres at single base-pair resolution.

2011-10-26 | E-GEOD-30551 | biostudies-arrayexpress

High-throughput single-base resolution mapping of 2’-O-Methylated residues

Project description:High-throughput single-base resolution mapping of 2’-O-Methylated residues

| PRJNA295991 | ENA

Bisulfite-Free Whole-Genome Mapping of 5-Methylcytosine at Single-Base Resolution by TAD-seq

Project description:5-Methylcytosine (5mC) is a crucial epigenetic modification plays a significant role in the regulation of gene expression. Accurate and quantitative detection of 5mC at single-base resolution is essential for understanding its epigenetic functions within genomes. In this study, we develop a novel nTET-assisted deaminase sequencing (TAD-seq) method for the base-resolution and quantitative detection of 5mC in genomic DNA. The TAD-seq method utilizes a Naegleria TET-like dioxygenase (nTET) to oxidize 5mC, generating 5-methylcytosine oxidation products (5moC). We also engineered a variant of the human apolipoprotein B mRNA-editing catalytic polypeptide-like 3A (A3A), creating an A3A mutant (A3Am). Treatment with A3Am results in the conversion of cytosine to uracil, while 5moC remains unchanged. Consequently, TAD-seq enables the direct deamination of cytosine to uracil by A3Am, which is sequenced as thymine, whereas 5mC, once oxidized to 5moC by nTET, resists deamination and is sequenced as cytosine. Therefore, the cytosines that persist in the sequencing data represent the original 5mC sites. We applied TAD-seq to HEK293T cells, generating a base-resolution map of 5mC that exhibits strong concordance with maps generated by conventional BS-seq. TAD-seq emerges as a powerful, bisulfite-free approach for the single-base resolution mapping of 5mC stoichiometry in genomic DNA.

2024-06-30 | GSE254203 | GEO

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