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.
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
Project description:We discovered 364 peaks enrichment change in wild-type C.elegans genome by 6mA MeDIP-seq, which indicated those genes response to mitochondrial stress and may function in mitochondrial stress response.
