GENOME-WIDE MAPPING OF NUCLEOSOME POSITIONING AND DNA METHYLATION WITHIN INDIVIDUAL DNA MOLECULES
We established a novel biochemical method, called Nome-seq which allows us to measure methylation AND nucleosome occupancy along the genomic sequence. These two pieces of information are crucial to understanding the regulatory context of the genome.
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.
RELEVANCE TO OC
Nome-seq allows us to understand methylation and its effects on gene regulation, and through these concepts we expect to learn much about the regulation of genes in ovarian cancer progression.