Base-excision repair pathway shapes 5-methylcytosine deamination signatures in pan-cancer genomes (CUT&RUN)
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ABSTRACT:
PROVIDER: PRJNA1149724 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR30296153_1.fastq.gz | Fastqsanger.gz | |||
| SRR30296153_2.fastq.gz | Fastqsanger.gz | |||
| SRR30296154_1.fastq.gz | Fastqsanger.gz | |||
| SRR30296154_2.fastq.gz | Fastqsanger.gz | |||
| SRR30296155_1.fastq.gz | Fastqsanger.gz |
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Project description:Transition of cytosine to thymine in CpG dinucleotides is the most frequent type of mutation in cancer. This increased mutability is commonly attributed to the spontaneous deamination of 5-methylcytosine (5mC), which is normally repaired by the base-excision repair (BER) pathway. However, the contribution of 5mC deamination in the increasing diversity of cancer mutational signatures remains poorly explored. Here, we integrate mutational signatures analysis in a large series of tumor whole genomes with lineage-specific epigenomic data to draw a detailed view on 5mC deamination in cancer. We uncover tumor type-specific patterns of 5mC deamination signatures in CpG and non-CpG contexts. We demonstrate that the BER glycosylase MBD4 preferentially binds to active chromatin and early replicating DNA, which correlates with lower mutational burden in these domains. We validate our findings by modeling BER deficiencies in isogenic cell models. Overall, we establish MBD4 as the main actor responsible for 5mC deamination repair in humans.
2024-09-12 | GSE275181 | GEO
Project description:N 4-methylcytosine (4mC) is a natural DNA modification occurring in thermophiles and plays important roles in restriction-modification (R-M) systems in bacterial genomes. However, the precise location and sequence context of 4mC in the whole genome are limited. In this study, we developed an APOBEC3A-mediated deamination sequencing (4mC-AMD-seq) method for genome-wide mapping of 4mC at single-base resolution. In the 4mC-AMD-seq method, cytosine and 5-methylcytosine (5mC) are deaminated by APOBEC3A (A3A) protein to generate uracil and thymine, both of which are read as thymine in sequencing, while 4mC is resistant to deamination and therefore read as cytosine. Thus, the readouts of cytosines from sequencing could manifest the original 4mC sites in genomes. With the 4mC-AMD-seq method, we achieved the genome-wide mapping of 4mC in Deinococcus radiodurans (D. radiodurans). In addition, we confirmed that 4mC, but not 5mC, was the major modification in the D. radiodurans genome. We identified 1586 4mC sites in the genome of D. radiodurans, among which 564 sites were located in the CCGCGG motif. The average methylation levels in the CCGCGG motif and non-CCGCGG sequence were 70.0% and 22.8%, respectively. We envision that the 4mC-AMD-seq method will facilitate the investigation of 4mC functions, including the 4mC-involved R-M systems, in uncharacterized but potentially useful strains.
2021-12-15 | GSE189965 | GEO
Project description:Genome-wide mapping of N4-methylcytosine at single-base resolution by APOBEC3A-mediated deamination sequencing
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Project description:5-Hydroxymethylcytosine (5hmC) is an important epigenetic mark that can regulate gene expression. While some methods were developed to detect 5hmC, direct genome-wide mapping of 5hmC at base resolution are still highly desirable. Herein, we proposed a single-step deamination sequencing (SSD-seq) method for the genome-wide mapping of 5hmC at single-base resolution. This method capitalizes on a screened engineered human apolipoprotein B mRNA-editing catalytic polypeptide-like 3A (A3A) protein to produce differential deamination activity toward cytosine (C), 5-methylcytosine (5mC), and 5hmC. In SSD-seq, an engineered A3A protein (eA3A-v10) can adequately deaminate C and 5mC, but not 5hmC. The original C and 5mC in DNA are deaminated by eA3A-v10 to form uracil (U) and thymine (T), both of which are read as T during sequencing. However, 5hmC is resistant to the deamination by eA3A-v10 and is still read as C during sequencing. Therefore, the remaining C in the sequence reads manifests the original 5hmC. Applying SSD-seq to generate a base-resolution map of 5hmC in human lung tissue, we found that 5hmC was almost entirely confined to CpG dinucleotides. The base-resolution map of 5hmC from human lung tissue generated by SSD-seq correlated strongly with that generated by prior ACE-seq. Taken together, the SSD-seq method is single-step, bisulfite-free and does not require DNA glycosylation or chemical treatment, which offers a valuable tool for the direct and quantitative detection of 5hmC in genomes at single-base resolution.
2023-12-04 | GSE236353 | 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:APOBEC (apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like) is a family of enzymes that deaminates cytosine (C) to uracil (U) on nucleic acid. APOBEC3B (A3B) functions in innate immunity against intrinsic and invading retroelements and viruses. A3B can also induce genomic DNA mutations to cause cancer. A3B contains two cytosine deaminase domains (CD1, CD2), and there are conflicting reports about whether both domains are active. Here we demonstrate that only CD2 of A3B (A3BCD2) has C deamination activity. We also reveal that both A3B and A3BCD2 can deaminate methylcytosine (mC). Guided by structural and functional analysis, we successfully engineered A3BCD2 to gain over two orders of magnitude higher activity for mC deamination. Important determinants that contribute to the activity and selectivity for mC deamination have been identified, which reveals that multiple elements, rather than single ones, contribute to the mC deamination activity and selectivity in A3BCD2 and possibly other APOBECs.
| S-EPMC4613526 | biostudies-literature
Project description:DNA methylation in mammals is an epigenetic mark necessary for normal embryogenesis. During development active loss of methylation occurs in the male pronucleus during the first cell cycle after fertilisation. This is accompanied by major chromatin remodelling and generates a marked asymmetry between the paternal and maternal genomes. The mechanism(s) by which this is achieved implicate, among others, base excision repair (BER) components and more recently a major role for TET3 hydroxylase. To investigate these methylation dynamics further we have analysed DNA methylation and hydroxymethylation in fertilised mouse oocytes by indirect immunofluorescence (IF) and evaluated the relative contribution of different candidate factors for active demethylation in knock-out zygotes by three-dimensional imaging and IF semi-quantification.We find two distinct phases of loss of paternal methylation in the zygote, one prior to and another coincident with, but not dependent on, DNA replication. TET3-mediated hydroxymethylation is limited to the replication associated second phase of demethylation. Analysis of cytosine deaminase (AID) null fertilised oocytes revealed a role for this enzyme in the second phase of loss of paternal methylation, which is independent from hydroxymethylation. Investigation into the possible repair pathways involved supports a role for AID-mediated cytosine deamination with subsequent U-G mismatch long-patch BER by UNG2 while no evidence could be found for an involvement of TDG.There are two observable phases of DNA demethylation in the mouse zygote, before and coincident with DNA replication. TET3 is only involved in the second phase of loss of methylation. Cytosine deamination and long-patch BER mediated by UNG2 appear to independently contribute to this second phase of active demethylation. Further work will be necessary to elucidate the mechanism(s) involved in the first phase of active demethylation that will potentially involve activities required for early sperm chromatin remodelling.
| S-EPMC4037648 | biostudies-literature
Project description:Bisulfite treatment damages the DNA leading to fragmentation and loss of long-range methylation information. To overcome this limitation of bisulfite treated DNA we applied a new enzymatic deamination approach, termed EM-seq (Enzymatic Methyl-seq) to long-range sequencing technologies. Our methodology, named LR-EM-seq (Long Range Enzymatic Methyl-seq) preserves the integrity of DNA allowing long-range methylation profiling of 5-mC and 5-hmC) over several kilobases of genomic DNA. Applied to known differentially methylated regions (DMR), LR-EM-seq achieves phasing of over 5 kb resulting in much broader and better defined DMRs than previously reported.
2020-09-01 | GSE141908 | GEO
Project description:Transcriptome-wide deamination analysis
| PRJNA951692 | ENA
Project description:DNA cytosine methylation (5-methylcytosine, 5mC) is a predominant epigenetic modification that plays critical roles in a variety of biological and pathological processes in mammals. In active DNA demethylation, the ten-eleven translocation (TET) dioxygenases can sequentially oxidize 5mC to generate three modified forms of cytosine, 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC). Beyond being a demethylation intermediate, recent studies have shown that 5fC has regulatory functions in gene expression and chromatin organization. While some methods have been developed to detect 5fC, genome-wide mapping of 5fC at base resolution are still highly desirable. Herein, we propose a Chemical Labeling Enrichment and Deamination sequencing (CLED-seq) method for detecting 5fC in genomic DNA at single-base resolution. The CLED-seq method utilizes selective labeling and enrichment of 5fC-containing DNA fragments, followed by deamination mediated by A3A and sequencing. In the CLED-seq process, while all C, 5mC, and 5hmC are interpreted as T during sequencing, 5fC is still read as C, enabling the precise detection of 5fC in DNA. Using the proposed CLED-seq method, we accomplished genome-wide mapping of 5fC in mouse embryonic stem cells. The mapping study revealed that promoter regions enriched with 5fC overlapped with H3K4me1, H3K4me3, and H3K27ac marks. These findings suggest a correlation between 5fC marks and active gene expression in mouse embryonic stem cells. In conclusion, CLED-seq is a straightforward, bisulfite-free method that offers a valuable tool for detecting 5fC in genomes at a single-base resolution.
2023-08-31 | GSE239943 | GEO