Base-resolution profiling of 5-glyceryl-methylcytosine in Chlamydomonas reinhardtii via DEA-sequencing [WGBS]
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PROVIDER: PRJNA1210647 | ENA |
REPOSITORIES: ENA
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Project description:5gmC (5-glyceryl-methylcytosine), a vitamin C-derived hypermodified base, was identified in the genome of Chlamydomonas reinhardtii. Despite its potential involvement in active DNA demethylation, the global distribution of 5gmC and its function as a stable epigenetic mark remain poorly understood. In this study, we employed a commercial DNA deaminase to distinguish 5gmC from 5mC (5-methylcytosine) and C, enabling precise profiling of 5gmC across the entire genome. This DEA-sequencing method demonstrates superior performance compared to the previously proposed approach, which relies on TET dioxygenase-coupled bisulfite treatment. Using both methods, we identified numerous confident 5gmC sites. Unlike 5mC, which predominantly occurs at CpG sites, 5gmC is more evenly distributed across both CpG and non-CpG contexts. Remarkably, 5gmC and 5mC are largely mutually exclusive, with less than 2% of the sites overlapping between the two modifications. Additionally, 5gmC shows significant enrichment at transcriptional start sites (TSS) and transcriptional end sites (TES), contrasting with the more extensive localization of 5mC in intergenic and gene body regions. Importantly, 5gmC levels are positively correlated with transcription, while 5mC typically exhibits an inverse relationship with gene expression. Together, these findings suggest that 5gmC is not only an intermediate in active DNA demethylation but also functions as a stable epigenetic mark, potentially influencing transcriptional regulation independently of 5mC.
2025-07-11 | GSE287000 | GEO
Project description:Base-resolution profiling of 5-glyceryl-methylcytosine in Chlamydomonas reinhardtii via DEA-sequencing [RNA-seq]
| PRJNA1210646 | ENA
Project description:Whole-genome bisulfite sequencing (WGBS) is currently the gold standard for DNA methylation (5-methylcytosine, 5mC) profiling, however the destructive nature of sodium bisulfite results in DNA fragmentation and subsequent biases in sequencing data. Such issues have led to the development of bisulfite-free methods for 5mC detection. Nanopore sequencing is a long read non-destructive approach that directly analyzes DNA and RNA fragments in real time. Recently, computational tools have been developed that enable base-resolution detection of 5mC from Oxford Nanopore sequencing data. In this chapter we provide a detailed protocol for preparation, sequencing, read assembly and analysis of genome-wide 5mC using Nanopore sequencing technologies.
2021-12-03 | GSE179673 | GEO
Project description:5gmC (5-glyceryl-methylcytosine), a vitamin C-derived hypermodified base, was identified in the genome of Chlamydomonas reinhardtii. Despite its potential involvement in active DNA demethylation, the global distribution of 5gmC and its function as a stable epigenetic mark remain poorly understood. In this study, we employed a commercial DNA deaminase to distinguish 5gmC from 5mC (5-methylcytosine) and C, enabling precise profiling of 5gmC across the entire genome. This DEA-sequencing method demonstrates superior performance compared to the previously proposed approach, which relies on TET dioxygenase-coupled bisulfite treatment. Using both methods, we identified numerous confident 5gmC sites. Unlike 5mC, which predominantly occurs at CpG sites, 5gmC is more evenly distributed across both CpG and non-CpG contexts. Remarkably, 5gmC and 5mC are largely mutually exclusive, with less than 2% of the sites overlapping between the two modifications. Additionally, 5gmC shows significant enrichment at transcriptional start sites (TSS) and transcriptional end sites (TES), contrasting with the more extensive localization of 5mC in intergenic and gene body regions. Importantly, 5gmC levels are positively correlated with transcription, while 5mC typically exhibits an inverse relationship with gene expression. Together, these findings suggest that 5gmC is not only an intermediate in active DNA demethylation but also functions as a stable epigenetic mark, potentially influencing transcriptional regulation independently of 5mC.
2025-07-11 | GSE286999 | GEO
Project description:Direct enzymatic sequencing of 5-methylcytosine at single-base resolution
| PRJNA938125 | ENA
Project description:Single-base resolution methylome of 5-methylcytosine in mammalian transcriptome
| PRJNA301456 | ENA
Project description:N4-methylcytosine is a major DNA modification integral to restriction-modification (R-M) systems in bacterial genomes. Here we describe 4mC-Tet-Assisted Bisulfite-sequencing (4mC-TAB-seq), a method that accurately and rapidly reveals the genome-wide locations of N4-methylcytosines at single-base resolution. By coupling Tet-mediated oxidation with a modified sodium bisulfite conversion reaction, unmodified cytosines and 5-methylcytosines are read out as thymines, whereas N4-methylcytosines are read out as cytosines revealing their positions throughout the genome. 4mC-TAB-seq
2015-07-11 | E-GEOD-63371 | biostudies-arrayexpress
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 NaegleriaTET-assisted deaminase sequencing (NTD-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 NTD-seq to HEK293T cells, generating a base-resolution map of 5mC that exhibits strong concordance with maps generated by conventional BS-seq. NTD-seq emerges as a powerful, bisulfite-free approach for the single-base resolution mapping of 5mC stoichiometry in genomic DNA.
2025-04-02 | GSE263426 | GEO
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
Project description:Direct enzymatic sequencing of 5-methylcytosine at single-base resolution [3]
| PRJNA938127 | ENA