Comment[ArrayExpressAccession] E-GEOD-42629 MAGE-TAB Version 1.1 Public Release Date 2013-06-15 Investigation Title Density and methylation state of CpG dinucleotides define histone variant specific retention of nucleosomes in mouse spermatozoa Comment[Submitted Name] Density and methylation state of CpG dinucleotides define histone variant specific retention of nucleosomes in mouse spermatozoa Experiment Description Nucleosomes are the principal packaging units of chromatin and critical for gene regulation and genome stability. In mammals, a subset of nucleosomes fail to be replaced by protamines during spermatogenesis and are retained in mature spermatozoa providing opportunities for paternal epigenetic transmission. In humans, the remaining 10% localize at regulatory elements of genes. To assess evolutionary conservation and to dissect the molecular logic underlying nucleosome retention, we determined the genome wide nucleosome occupancy in mouse spermatozoa that only contain 1% residual histones. In striking contrast to mammalian somatic cells and haploid round spermatids, we observe high enrichment of nucleosomes at CpG-rich sequences throughout the genome, at conserved regulatory sequences as well as at intra- and intergenic regions and repetitive DNA. This preferred occupancy occurs mutually exclusive with DNA methylation both in mouse and human sperm. At unmethylated CpG-rich sequences, residing nucleosomes are largely composed of the H3.3 histone variant, and trimethylated at lysine 4 (H3K4me3). Both canonical H3.1/H3.2 and H3.3 variant histones are present at promoters marked by Polycomb-mediated H3K27me3, which is strongly predictive for gene repression in pre-implantation embryos. Our data indicate important roles of DNA sequence composition, DNA methylation, variant H3.3 and canonical H3.1/H3.2 histones and associated modifications in nucleosome retention versus eviction during the histone-to-protamine remodeling process in elongating spermatids and potentially in epigenetic inheritance by nucleosomes between generations. Identification of histone, histone variant and histone modification states in round spermatids and sperm Term Source Name ArrayExpress EFO Term Source File http://www.ebi.ac.uk/arrayexpress/ http://www.ebi.ac.uk/efo/efo.owl Person Last Name Peters Erkek Hisano Murr Schuebeler Stadler Peters Person First Name Antoine Serap Mizue Rabih Dirk Michael Antoine Person Email antoine.peters@fmi.ch Person Affiliation Friedrich Miescher Institute for Biomedical Research (FMI) Person Address Friedrich Miescher Institute for Biomedical Research (FMI), Maulbeerstrasse 66, Basel, Switzerland Person Roles submitter Protocol Name P-GSE42629-2 P-GSE42629-3 P-GSE42629-4 P-GSE42629-1 Protocol Description Low-complexity reads were filtered out on the basis of dinucleotide entropy. Read are aligned by using Bowtie. Tab and wig files were generated in the same way as described in Stadler et al., Nature 2011, pubmed ID: 22170606 Genome_build: mm9 Supplementary_files_format_and_content: Wig files show the average coverage per base in 100 bp windows in the genome. Low-complexity reads were filtered out on the basis of dinucleotide entropy. Read are aligned by using Bowtie. Tab and wig files were generated in the same way as described in Stadler et al., Nature 2011, pubmed ID: 22170606 Genome_build: mm9 Supplementary_files_format_and_content: Tab file show the total and methylated alignments overlapping individual CpGs. Mouse spermatozoa were collected by swim-up procedure from C57BL/6J mice. Round spermatids were isolated by FACS sorting using 28 day old BL6 mice. Mono-nucleosomal chromatin samples were prepared by MNase treatment. ChIP experiments for histone variants, H3.1/H3.2, H3.3, and histone modifications H3K4me3 and H3K27me3 were performed on native chromatin. ChIP-seq and MNase-seq libraries were produced using the Illumina ChIP-seq DNA Sample Prep Kit (Cat# IP-102-1001).Before preparing RNA-seq libraries, rRNA from RNA was depleted by using the Ribo-Zero rRNA removal kit (Epicentre Biotechnologies). Bisulfite libraries were prepared as described in Stadler et al., Nature 2011, pubmed ID: 22170606 Strand specific RNA-seq libraries were prepared by following the Illumina directional mRNA-seq library preparation pre-release protocol. Mouse spermatozoa were collected by swim-up procedure5 from C57BL/6J mice. Round spermatids were isolated by FACS sorting using 28 day old BL6 mice. Mono-nucleosomal chromatin samples were prepared by MNase treatment. ChIP experiments for histone variants, H3.1/H3.2, H3.3, and histone modifications H3K4me3 and H3K27me3 were performed on native chromatin. ChIP-seq and MNase-seq libraries were produced using the Illumina ChIP-seq DNA Sample Prep Kit (Cat# IP-102-1001).Before preparing RNA-seq libraries, rRNA from RNA was depleted by using the Ribo-Zero rRNA removal kit (Epicentre Biotechnologies). Bisulfite libraries were prepared as described in Stadler et al., Nature 2011, pubmed ID: 22170606 Strand specific RNA-seq libraries were prepared by following the Illumina directional mRNA-seq library preparation pre-release protocol. Protocol Type normalization data transformation protocol normalization data transformation protocol nucleic acid library construction protocol nucleic acid library construction protocol Experimental Factor Name ORGANISM PART ANTIBODY Experimental Factor Type organism part antibody Comment[SecondaryAccession] GSE42629 Comment[GEOReleaseDate] 2013-06-15 Comment[ArrayExpressSubmissionDate] 2012-11-29 Comment[GEOLastUpdateDate] 2013-06-25 Comment[AEExperimentType] RNA-seq of coding RNA Comment[AEExperimentType] ChIP-seq Comment[AEExperimentType] methylation profiling by high throughput sequencing Comment[SecondaryAccession] SRP017350 Comment[SequenceDataURI] http://www.ebi.ac.uk/ena/data/view/SRR625500-SRR625518,SRR656613-SRR656614 SDRF File E-GEOD-42629.sdrf.txt