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Single-cell 5-formylcytosine Landscapes of Mammalian Early Embryos at Single-base Resolution

ABSTRACT: Single-cell 5-formylcytosine Landscapes of Mammalian Early Embryos at Single-base Resolution

PROVIDER: PRJNA1095445 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR28533002_1.fastq.gz	Fastqsanger.gz
	SRR28533002_2.fastq.gz	Fastqsanger.gz
	SRR28533003_1.fastq.gz	Fastqsanger.gz
	SRR28533003_2.fastq.gz	Fastqsanger.gz
	SRR28533004_1.fastq.gz	Fastqsanger.gz

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Single-cell 5-formylcytosine Landscapes of Mammalian Early Embryos at Single-base Resolution

Project description:Active DNA demethylation in mammals involves Ten-eleven translocation (TET) family proteins-mediated oxidation of 5-methylcytosine (5mC). However, base-resolution landscapes of 5-formylcytosine (5fC, an oxidized derivative of 5mC) at the single-cell level remains unexplored. Here we present “CLEVER-seq”, which is a single-cell, single-base resolution 5fC sequencing technology, based on biocompatible, selective chemical labeling of 5fC and subsequent C-to-T conversion during amplification and sequencing. CLEVER-seq of mouse early embryos reveals the highly patterned genomic distribution and parental specific dynamics of 5fC during mouse early pre-implantation development and synergistic 5fC production in paternal and maternal genomes. Integrated analysis demonstrates that promoter 5fC production precedes the expression upregulation of a clear set of developmentally and metabolically critical genes. Collectively, our work reveals the dynamics of active DNA demethylation during early mouse pre-implantation development and provides an important resource for further functional studies of epigenetic reprogramming in single cells.

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Quantitative sequencing of 5-formylcytosine in DNA at single-base resolution

Project description:Cytosine base modifications 5-methylcytosine (5mC), 5-hydroxymethylcytosine (5hmC) and 5-formylcytosine (5fC) are present in mammalian DNA. Here, reduced bisulfite sequencing is developed for quantitatively sequencing 5fC at single-base resolution. This method is then applied with oxidative bisulfite sequencing to gain a map of 5mC, 5hmC and 5fC in mouse embryonic stem cells. 12 samples, reduced representation bisulphite treatment: 4 replicates each for bisulphite (BS), oxidative BS (oxBS) and reduced BS (redBS) for the detection of 5mC, 5hmC and 5fC. Mouse (strain B6C) embryonic stem cells.

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Single-Cell 5-Hydroxymethylcytosine Landscapes of Mammalian Early Embryos at Single-Base Resolution

Project description:Single-Cell 5-Hydroxymethylcytosine Landscapes of Mammalian Early Embryos at Single-Base Resolution

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Project description:Mapping genome-wide 5-hydroxymethylcytosine (5hmC) and 5-formylcytosine (5fC) at single-base resolution is important to understand their biological functions. We present a cost-efficient mapping method that combines 5hmC-specific restriction enzyme PvuRts1I with a 5hmC enrichment method. The sensitive method enables detection of low abundant 5hmC sites, providing a more complete 5hmC landscape than available bisulfite-based methods. This method generated the first genome-wide 5fC map at single-base resolution. Parallel analyses revealed that 5hmC and 5fC existed with lower abundance and more dynamically in non-CpG context than in CpG context. In the genic region, distribution of 5hmCpG and 5fCpG differed from 5hmCH and 5fCH (H=A, T, C). 5hmC and 5fC were distributed distinctly at regulatory protein-DNA binding sites, depleted in permissive transcription factor binding sites, and enriched at active and poised enhancers. This sensitive bisulfite-conversion free method can be applied to biological samples with limited starting material or low abundance of cytosine modifications. Sensitive mapping of genome-wide 5-hydroxymethylcytosine and 5-formylcytosine in mouse embryonic stem cell at single-base resolution by combining 5-hydroxymethylcytosine specific restriction enzyme PvuRts1I and 5-hydroxymethylcytosine enrichment method (selective chemical labeling or SEAL)

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Project description:DNA methylation and hydroxymethylation are extensively reprogrammed in mammalian early embryogenesis. However, a comprehensive picture of DNA hydroxymethylation map across early embryo stages is lacking. Here, we develop “schmC-CATCH” (single-cell 5hmC chemical-assisted C-to-T conversion-enabled sequencing) to obtain quantitative, genome-wide landscape of the base-resolution DNA hydroxymethylome in mouse gametes and pre-implantation embryos from one-cell (PN1-PN5) to blastocyst stages. We revealed the dynamics of DNA hydroxymethylation and observed dramatically different 5hmC patterns on the paternal and maternal genomes. We found hotspots of DNA hydroxymethylation during mouse early embryo development, which are highly associated with young retroelements including LINE1. 5hmC is also associated with regulatory elements, indicating a potential epigenetic function during early embryogenesis. In addition, 5hmC is asymmetrically distributed at the chromosome level and can be used to track the cell and strand lineages during the first two embryo cleavages. Collectively, our work reveals the dynamics of active DNA demethylation during mouse pre-implantation development and provides an important resource for functional studies of epigenetic reprogramming in single cells.

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