Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Genome-wide mapping of nucleosome positioning and DNA methylation within Individual DNA molecules

ABSTRACT: DNA methylation and nucleosome positioning work together to generate chromatin structures that regulate gene expression. Nucleosomes are typically mapped using nuclease digestion requiring significant amounts of material and varying enzyme concentrations. We have developed a method (NOMe-seq) that uses a GpC methyltransferase (M.CviPI) and next generation sequencing to generate a high resolution footprint of nucleosome positioning genome-wide using less than 1 million cells while retaining endogenous DNA methylation information from the same DNA strand. Using a novel bioinformatics pipeline we show a striking anti-correlation between nucleosome occupancy and DNA methylation at CTCF regions, that is not present at promoters. We further show that the extent of nucleosome depletion at promoters is directly correlated to expression level and can accommodate multiple nucleosomes and provide genome-wide evidence that expressed non-CpG island promoters are nucleosome depleted. Importantly, NOMe-seq obtains DNA methylation and nucleosome positioning information from the same DNA molecule, giving the first genome-wide DNA methylation and nucleosome positioning correlation at the single molecule and thus, single cell level that can be used to monitor disease progression and response to therapy. Nucleosome Occupancy and Methylome-Sequencing (NOMe-Seq) on IMR90 cell line and 2 GBM cell lines

ORGANISM(S): Homo sapiens

SUBMITTER: Yaping Liu

PROVIDER: E-GEOD-40770 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:DNA cytosine methylation is a hallmark of epigenetic gene silencing. DNA demethylation requires ROS1, a bifunctional DNA glycosylase/lyases and open chromatin status mediated by IDM1. HDA6 is a RDP3-like histone deacetylase and was confirmed to mediate DNA methylation. In previous screening for ros1 suppressor, we identified two hda6 mutants reverting ros1-caused hypermethylation at RD29A and 35S promoters respectively, indicating the antagonization of DNA methylation between HDA6 and ROS1. To learn antagonized effects between HDA6 and ROS1 at DNA cytosine methylation genome-wide, we performed whole genome bisulfite sequencing to search antagonized targets of HDA6 and ROS1 and their specific targets to evaluate their roles on DNA methylation. To evaluate HDA6’s roles in sRNA biogenesis and nucleosome positioning, we also performed small RNA sequencing and genome-wide mapping of nucleosome positioning of C24, ros1, hda6-9 and hda6-10. Our results indicate that around 43% ros1-caused CG hypermethylation, 74.5% and 84.5% ros1-caused CHG and CHH hypermethylation were reverted by the two hda6 alleles in the ros1 background respectively. These results indicate that most of ROS1-demethylated targets are also HDA6-mediated DNA methylated targets. In addition, we observed that HDA6-affected DNA methylation targets are far more than ROS1-demethylation targets at CHG and CHH context, but not at CG context. sRNA analysis showed that HDA6 inhibits LTR/Gypsy and TE gene 24nt siRNA accumulation, while promotes RC/Helitron 24nt siRNA accumulation. Our Mono-nucleosome positioning data further showed that the two hda6 alleles have striking difference on nucleosome positioning, hda6-10 obviously have different nucleosome positioning patterns with hda6-9. Our results indicate HDA6 not only mediates overall DNA methylation at CG, CHG and CHH context, and antagonizes with ROS1 mediated DNA demethylation, but also regulates nucleosome positioning and small RNA accumulation at some genome specific regions.

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Genome-wide mapping of nucleosome positioning and DNA methylation within Individual DNA molecules

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