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A bioinformatic pipeline for analysis of M.EcoGII methylation footprint PacBio long-read sequence data

ABSTRACT: Recent studies have combined DNA methyltransferase footprinting of genomic DNA in nuclei with long-read sequencing, resulting in detailed chromatin maps for multi-kilobase stretches of genomic DNA from one cell. Nucleosome footprints and nucleosome-depleted regions can be identified, yielding unprecedented information concerning their degree of correlation within the same cell. The enzyme of choice is M.EcoGII, which methylates adenines in any sequence context, potentially resulting in very high resolution. However, in practice, the methylation efficiency (defined as the fraction of each genomic adenine that is methylated), is quite low, resulting in false footprints predicted by random unmethylated gaps between methylated adenines. We report PacBio long-read sequence data for budding yeast nuclei treated with M.EcoGII. We present a bioinformatic pipeline which accounts for methylation efficiency, as well as an observed bias against methylation as a function of increasing AT content. It also accounts for our observation that some methylation occurs within nucleosomes, breaking up their footprints. Comparison of long reads for each gene indicates the extent of chromatin heterogeneity within the cell population. Although the population average is consistent with that derived using other techniques, we observe a wide range of heterogeneity in nucleosome positions at the single-molecule level.

ORGANISM(S): Escherichia coli Saccharomyces cerevisiae

PROVIDER: GSE243114 | GEO | 2024/03/15

REPOSITORIES: GEO

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Project description:Probing epigenetic features on long molecules of DNA has tremendous potential to advance our understanding of the phased epigenome. In this study, we evaluate CpG methylation and chromatin accessibility simultaneously on long strands of DNA using GpC methyltransferase to exogenously label open chromatin, coupled with nanopore sequencing technology. We performed nanopore sequencing of Nucleosome Occupancy and Methylome (nanoNOMe) on four human cell lines (GM12878, MCF-10A, MCF-7, MDA-MB-231), and demonstrate the ability to directly measure methylation and chromatin accessibility in genomic features such as structural variations and repetitive elements. The long single-molecule resolution allows footprinting of protein and nucleosome binding and determining the combinatorial promoter epigenetic state on individual molecules. Long-read sequencing makes it possible to robustly assign reads to haplotypes, enabling allele-specific epigenetic analysis across the genome. We use existing SNV data on GM12878 to present the first fully phased human Probing epigenetic features on long molecules of DNA has tremendous potential to advance our understanding of the phased epigenome. We evaluate CpG methylation and chromatin accessibility simultaneously on long strands of DNA using GpC methyltransferase to exogenously label open chromatin, coupled with nanopore sequencing technology. We performed nanopore sequencing of Nucleosome Occupancy and Methylome (nanoNOMe) on four human cell lines (GM12878, MCF-10A, MCF-7, MDA-MB-231), and demonstrate the ability to directly measure methylation and chromatin accessibility in genomic features such as structural variations and repetitive elements. The long single-molecule resolution allows footprinting of protein and nucleosome binding and determining the combinatorial promoter epigenetic state on individual molecules. Long-read sequencing makes it possible to robustly assign reads to haplotypes, enabling allele-specific epigenetic analysis across the genome. We use existing SNV data on GM12878 to present the first fully phased human epigenome, consisting of chromosome-level allele-specific profiles of CpG methylation and chromatin accessibility.mosome-level allele-specific profiles of CpG methylation and chromatin accessibility.

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A bioinformatic pipeline for analysis of M.EcoGII methylation footprint PacBio long-read sequence data

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