N6-adenine DNA-immunoprecipitation-based genomic profiling in eukaryotes
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA811954 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR18191400_1.fastq.gz | Fastqsanger.gz | |||
| SRR18191400_2.fastq.gz | Fastqsanger.gz | |||
| SRR18191401_1.fastq.gz | Fastqsanger.gz | |||
| SRR18191401_2.fastq.gz | Fastqsanger.gz | |||
| SRR18191380_1.fastq.gz | Fastqsanger.gz |
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Project description:DNA methylation in the form of 5-methylcytosine (5mC) is widespread in eukaryotes, while the presence of N6-methyladenine (6mA) has sparked considerable debate. Methodological disparities in quantifying and mapping 6mA in genomic DNA have fueled this controversy. Yet, the distantly related early branching fungi, ciliates and the algae Chlamydomonas reinhardtii exhibit robust 6mA methylation patterns, but the origin and evolution of 6mA remain unknown. Here we use Oxford Nanopore modified base calling to profile 6mA at base pair resolution in 18 unicellular eukaryotes spanning all major eukaryotic supergroups. Our results reveal that only species encoding the adenine methyltransferase AMT1 display robust genomic 6mA patterns. Notably, 6mA consistently accumulates downstream of transcriptional start sites, aligning with H3K4me3-enriched nucleosomes, suggesting a conserved role in placing transcriptionally permissive nucleosomes. Intriguingly, the recurrent loss of the 6mA pathway across eukaryotes, particularly in major multicellular lineages, implies a convergent alteration in the dual methylation system of the Last Eukaryotic Common Ancestor, which featured transcription-associated 6mA and repression-associated 5mC.
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Project description:DNA methylation is a key mechanism for regulation of DNA repair, DNA replication, and gene expression. In bacteria, DNA is modified by methylation on C5-cytosine (5mC), N6-adenine (6mA) and N4-cytosine (4mC). Metazoans were thought to only use 5mC to regulate gene expression until the recent discovery of N6-adenine methylation in DNA of diverse metazoan species, including mammals. Here we show by dot blot, immunoprecipitation with 4mC-specific antibodies, and ultra-high performance liquid chromatography coupled with triple-quadrupole mass spectrometry that 4mC also occurs in most eukaryotes. We further characterize 4mC sites on a near genome-wide scale by single molecule real time (SMRT) sequencing in the nematode C. elegans. In C. elegans, 4mC is found in all chromosomes but is relatively sparse on the X chromosome. 4mC is relatively enriched on introns, depleted on exons, and co-localizes with 6mA. In human cells, 4mC is anti-correlated with 5mC. Moreover, 4mC modifications are associated with increased gene expression. Transfection of human cells with a 4mC-modified luciferase reporter plasmid increases luciferase expression as compared to the unmethylated plasmid. These data suggest that 4mC promotes gene transcription, playing an opposing role to 5mC, which generally represses transcription. Thus 4mC modification of DNA is a previously unrecognized mechanism of gene regulation in eukaryotes, providing evidence for a complex epigenetic DNA code.
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