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DNA N6-adenine methylation in Arabidopsis thaliana (RNA)

ABSTRACT: DNA methylation on N6-adenine (6mA), the most prevalent DNA modification in prokaryotes, has recently been found as a potentially new epigenetic mark in several unicellular and multicellular eukaryotes. However, the distribution patterns and potential functions of 6mA in land plants, which are primary producers for most ecosystems, remain completely unknown. Here we report global profiling of 6mA sites at single-nucleotide resolution in the genomes of Arabidopsis thaliana Columbia-0 (Col), using single-molecule real-time sequencing. DNA methylome analysis shows that 6mA sites are widely distributed across the Col genomes and enriched over the pericentromeric heterochromatin regions. Nearly 30% of 6mA sites are present in gene bodies with a trend of enrichment around the transcriptional start site. In addition to a common consensus 6mA site found in other eukaryotes, novel 6mA sites were found, indicating that 6mA could evolve new functions in land plants. Further analysis of 6mA methylome and RNA-sequencing data demonstrates that 6mA positively correlates with the gene expression level in Col plants. Consistently, DNA affinity chromatography coupled with mass spectrometry reveals that histone variants associated with actively expressed genes interact with 6mA DNA. Our results uncover 6mA as a DNA mark associated with actively expressed genes in Arabidopsis, indicating that 6mA could serve as a potentially novel epigenetic mark in land plants.

ORGANISM(S): Arabidopsis thaliana

PROVIDER: GSE81594 | GEO | 2018/04/16

REPOSITORIES: GEO

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DNA N6-adenine methylation in Arabidopsis thaliana (methylation)

Project description:DNA methylation on N6-adenine (6mA), the most prevalent DNA modification in prokaryotes, has recently been found as a potentially new epigenetic mark in several unicellular and multicellular eukaryotes. However, the distribution patterns and potential functions of 6mA in land plants, which are primary producers for most ecosystems, remain completely unknown. Here we report global profiling of 6mA sites at single-nucleotide resolution in the genomes of Arabidopsis thaliana Columbia-0 (Col), using single-molecule real-time sequencing. DNA methylome analysis shows that 6mA sites are widely distributed across the Col genomes and enriched over the pericentromeric heterochromatin regions. Nearly 30% of 6mA sites are present in gene bodies with a trend of enrichment around the transcriptional start site. In addition to a common consensus 6mA site found in other eukaryotes, novel 6mA sites were found, indicating that 6mA could evolve new functions in land plants. Further analysis of 6mA methylome and RNA-sequencing data demonstrates that 6mA positively correlates with the gene expression level in Col plants. Consistently, DNA affinity chromatography coupled with mass spectrometry reveals that histone variants associated with actively expressed genes interact with 6mA DNA. Our results uncover 6mA as a DNA mark associated with actively expressed genes in Arabidopsis, indicating that 6mA could serve as a potentially novel epigenetic mark in land plants.

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DNA N6-adenine methylation in Arabidopsis thaliana [SMRT-seq]

Project description:DNA methylation on N6-adenine (6mA) has recently been found as a potentially new epigenetic mark in several unicellular and multicellular eukaryotes. However, its distribution patterns and potential functions in land plants, which are primary producers for most ecosystems, remain completely unknown. Here we report global profiling of 6mA sites at single-nucleotide resolution in the genome of Arabidopsis thaliana using single-molecule real-time sequencing. 6mA sites are widely distributed across the Arabidopsis genome and enriched over the pericentromeric heterochromatin regions. Nearly 30% of 6mA sites are present in gene bodies. Further analysis of 6mA methylome and RNA-sequencing data demonstrates that 6mA frequency positively correlates with the gene expression level in Arabidopsis. Consistently, histone variants associated with actively expressed genes interact with 6mA DNA. Our results uncover 6mA as a DNA mark associated with actively expressed genes in Arabidopsis, suggesting that 6mA serves as a novel epigenetic mark in land plants.

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DNA N6-adenine methylation in 3-week-old Arabidopsis thaliana [RNA-seq]

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Characterization of eukaryotic DNA N6-methyladenine by a highly sensitive restriction enzyme-assisted sequencing

Project description:To interrogate single-base resolution 6mA sites in the genome-wide, we develop DA-6mA-seq (DpnI-Assisted N6-methylAdenine sequencing), an optimized sequencing method taking advantage of restriction enzyme DpnI, which exclusively cleaves methylated adenine sites. We find DpnI also recognizes other sequence motifs besides the canonical GATC restriction sites, largely expanding the application range of this method. DA-6mA-seq requires less starting material and lower sequencing depth than previous methods, but achieves higher sensitivity, providing a good strategy to identify 6mA in large genome with a low abundance of 6mA. We rebuild the 6mA maps of Chlamydomonas by DA-6mA-seq and then apply this method to another two eukaryotic organisms, Plasmodium and Penicillium. Further analysis reveals most 6mA sites are symmetric at various sequence contexts, suggesting 6mA may function as a new heritable epigenetic mark in eukaryotes. A new sequencing method is developed to detect 6mA in eukaryotes

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N6-Methyldeoxyadenosine Marks Active Transcription Start Sites in Chlamydomonas

Project description:N6-methyldeoxyadenosine (6mA or m6A) is a DNA modification preserved in prokaryotes to eukaryotes. It is widespread in bacteria, and functions in DNA mismatch repair, chromosome segregation, and virulence regulation. In contrast, the distribution and function of 6mA in eukaryotes have been unclear. Here we present a comprehensive analysis of the 6mA landscape in the genome of Chlamydomonas using new sequencing approaches. We identified the 6mA modification in 84% of genes in Chlamydomonas. We found that 6mA mainly locates at ApT dinucleotides around transcription start sites (TSS) with a bimodal distribution, and appears to mark active genes. A periodic pattern of 6mA deposition was also observed at base resolution, which is associated with nucleosome distribution near the TSS, suggesting a possible role in nucleosome positioning. The new genome-wide mapping of 6mA and its unique distribution in the Chlamydomonas genome suggest potential regulatory roles of 6mA in gene expression in eukaryotic organisms. Multiple sequencing methods are developed to profile the distribution of 6mA in Chlamydomonas including MeDIP-Seq, enzyme-treated DNA-Seq, MNase-Seq and RNA-Seq.

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Project description:N6-methyladenine (6mA) DNA modification in eukaryotic genomes has emerged as a potential epigenetic mark. However, little is known about how 6mA epigenetic codes are read and interpreted in higher eukaryotes. Here we investigate 6mA genome-wide distributions in multiple higher eukaryotes. Using Drosophila as a pioneer system, we show that 6mA exhibits defined patterns and preferentially marks zygotic genes in early embryos. Moreover, we identify that the Fox-family protein, Jumu, is a "6mA-DNA reader" and functions in concert with DMAD to contribute to zygotic gene activation. Further methylome analysis reveals that 6mA modification has common features in genomic DNA from mouse, rat and monkey, and that "forkhead-domain-binding motifs" are enriched in 6mA-marked DNA of these genomes, in a way similar to Drosophila. Collectively, our findings identify the 6mA DNA reader protein and suggest a conserved 6mA-based mechanism in higher eukaryotes.

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N6-methyldeoxyadenosine directs nucleosome positioning in Tetrahymena DNA

Project description:N6-methyldeoxyadenosine (6mA or m6dA) was shown more than 40 years ago to exist in simple eukaryotes, yet functional studies have been limited. Recent investigations in multiple eukaryotes suggest 6mA as a potential DNA epigenetic mark that plays regulatory roles in gene regulation. Here we use Tetrahymena thermophila as a model to examine the effects of 6mA on nucleosome positioning. We have employed independent methods to identify genome-wide 6mA distribution, which revealed the enrichment after transcription start sites with a periodic pattern and a mutually exclusive relationship with the positions of nucleosomes. The exclusive distribution pattern of 6mA and nucleosome can be recapitulated by in vitro nucleosome assembly on native Tetrahymena genomic DNA, but not on DNA without 6mA. Model DNA containing artificially installed 6mA resists nucleosome assembling compared to unmodified DNA in vitro. Computational simulation revealed that 6mA increases dsDNA rigidity, which disfavors nucleosome wrapping. Knockout of a potential 6mA methyltransferase disturbs the nucleosome positioning in Tetrahymena, leading to the transcriptome-wide change of gene expression. These findings uncover a new mechanism by which DNA 6mA assists to shape the chromatin topology in order to stabilize gene expression.

2018-10-17 | GSE104699 | GEO

N6-Methyldeoxyadenosine Marks Active Transcription Start Sites in Chlamydomonas

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Identification and analysis of adenine N6-methylation sites in the rice genome

Project description:We report the application of 6mA IP-sequencing technology for high-throughput profiling in rice tissues of 12 days' leaf. By obtaining over 20 million high quality clean mapping sequenced reads from immunoprecipitated DNA, we generate genome-wide 6mA maps of rice 12 days' leaf. Using mass spectrometry and immunoprecipitation and validation with analysis of single-molecule real-time sequencing, we observed that about 0.2% of all adenines are 6mA-methylated in the rice genome. 6mA occurs most frequently at GAGG motifs and is mapped to about 20% of genes and 14% of transposable elements (TEs). In promoters, 6mA marks silent genes, but in bodies correlates with gene activity. 6mA overlaps with 5-methylated cytosine (5mC) at CG sites in gene bodies and is complementary to 5mC at CHH sites in TEs. We show that OsALKBH1 may be potentially involved in 6mA demethylation in rice. The results suggest that 6mA is complementary to 5mC as an epigenomic mark in rice and reinforces a distinct role for 6mA as a gene-expression associated epigenomic mark in eukaryotes.

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Identification of N6-adenine methylation in C. elegans DNA

Project description:In mammalian cells, DNA methylation on the 5th position of cytosine (5mC) plays an important role as an epigenetic mark. However, DNA methylation was considered to be absent in C. elegans because of the lack of detectable 5mC as well as homologs of the cytosine DNA methyltransferases. Here, using multiple approaches, we demonstrate the presence of adenine N6-methylation (6mA) in C. elegans DNA. We further demonstrate that this modification increases trans-generationally in a paradigm of epigenetic inheritance. Importantly, we identify a DNA demethylase, NMAD-1, and a potential DNA methyltransferase, DAMT-1, which regulate 6mA levels and crosstalk between methylation of histone H3K4me2 and 6mA, and control the epigenetic inheritance of phenotypes associated with the loss of the H3K4me2 demethylase spr-5. Together, these data identify a novel DNA modification in C. elegans and raise the exciting possibility that 6mA may be a carrier of heritable epigenetic information in eukaryotes. SMRT-sequencing for a mixed cell population of wildtype worms 6mA ChIP-Seq for a mixed cell population of wildtype worms

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