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Gcn5-mediated histone acetylation governs nucleosome dynamics in spermiogenesis [MNase-Seq]

ABSTRACT: During mammalian spermatogenesis, male germ cells undergo dramatic reorganization of chromatin, whereby 90-99% of histones are evicted and replaced by protamines. This reorganization prominently features histone acetylation to loosen chromatin structure. Given the potential role of retained histones in fertility and early embryonic development, the genomic location of retained nucleosomes is of great interest. However, the ultimate position and mechanisms underlying nucleosome eviction/retention are poorly understood, including several studies utilizing MNase-seq methodologies, but reporting remarkably dissimilar locations. Here, we utilized ATAC-seq to determine the location of retained nucleosomes in mouse sperm and found enrichment at promoters, but also retention at inter- and intragenic regions, and repetitive elements. We further investigated nucleosome eviction/retention by generating pre-meiotic, germ cell specific, conditional knockout mice for the histone acetyltransferase Gcn5, a key histone acetyltransferase. Gcn5cKO germ cells exhibited abnormal chromatin dynamics during spermiogenesis, including diminished nucleosome eviction leading to increased histone retention in sperm. These mice exhibited severe reproductive phenotypes: abnormal sperm production, sperm morphology, and ultimately, male infertility. Our findings demonstrate Gcn5 mediated histone acetylation promotes chromatin accessibility and nucleosome eviction in spermiogenesis, and that loss of histone acetylation leads to defects that disrupt male fertility and potentially, early embryogenesis.

ORGANISM(S): Mus musculus

PROVIDER: GSE134560 | GEO | 2019/11/07

REPOSITORIES: GEO

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Project description:To achieve the extreme nuclear condensation necessary for sperm function, most histones are replaced with protamines during spermiogenesis in mammals. Mature sperm retain only a small fraction of nucleosomes, which are, in part, enriched on gene regulatory sequences, and recent findings suggest that these retained histones provide epigenetic information that regulates expression of a subset of genes involved in embryo development after fertilization. We addressed this tantalizing hypothesis by analyzing two mouse models exhibiting abnormal histone positioning in mature sperm due to impaired poly(ADP-ribose) (PAR) metabolism during spermiogenesis and identified altered sperm histone retention in specific gene loci genome-wide using MNase digestion-based enrichment of mononucleosomal DNA. We then set out to determine the extent to which expression of these genes was altered in embryos generated with these sperm. For control sperm, most genes showed some degree of histone association, unexpectedly suggesting that histone retention in sperm genes is not an all-or-none phenomenon and that a small number of histones may remain associated with genes throughout the genome. The amount of retained histones, however, was altered in many loci when PAR metabolism was impaired. To ascertain whether sperm histone association and embryonic gene expression are linked, the transcriptome of individual 2-cell embryos derived from such sperm was determined using microarrays and RNA sequencing. Strikingly, a moderate but statistically significant portion of the genes that were differentially expressed in these embryos also showed different histone retention in the corresponding gene loci in sperm of their fathers. These findings provide new evidence for the existence of a linkage between sperm histone retention and gene expression in the embryo. Mnase sensitivity of sperm DNA, indicating nucleosomal, not protamine, packaging was altered in mice by manipulating poly(ADP-ribose) metabolism in adult males using a specific PARP inhibitor for 6 weeks. Abnormal sperm nucleosomal organization of males analyzed by these tiling arrays was compared with differential gene expression in 2 cell embryos fathered by these males analyzed by separate gene expression arrays and RNA sequencing.

Project description:To achieve the extreme nuclear condensation necessary for sperm function, most histones are replaced with protamines during spermiogenesis in mammals. Mature sperm retain only a small fraction of nucleosomes, which are, in part, enriched on gene regulatory sequences, and recent findings suggest that these retained histones provide epigenetic information that regulates expression of a subset of genes involved in embryo development after fertilization. We addressed this tantalizing hypothesis by analyzing two mouse models exhibiting abnormal histone positioning in mature sperm due to impaired poly(ADP-ribose) (PAR) metabolism during spermiogenesis and identified altered sperm histone retention in specific gene loci genome-wide using MNase digestion-based enrichment of mononucleosomal DNA. We then set out to determine the extent to which expression of these genes was altered in embryos generated with these sperm. For control sperm, most genes showed some degree of histone association, unexpectedly suggesting that histone retention in sperm genes is not an all-or-none phenomenon and that a small number of histones may remain associated with genes throughout the genome. The amount of retained histones, however, was altered in many loci when PAR metabolism was impaired. To ascertain whether sperm histone association and embryonic gene expression are linked, the transcriptome of individual 2-cell embryos derived from such sperm was determined using microarrays and RNA sequencing. Strikingly, a moderate but statistically significant portion of the genes that were differentially expressed in these embryos also showed different histone retention in the corresponding gene loci in sperm of their fathers. These findings provide new evidence for the existence of a linkage between sperm histone retention and gene expression in the embryo. Mnase sensitivity of sperm DNA, indicating nucleosomal, not protamine, packaging was altered in mice by manipulating poly(ADP-ribose) metabolism in adult males using Parg gene disruption (Parg(110)-/-). Abnormal sperm nucleosomal organization of males analyzed by these tiling arrays was compared with differential gene expression in individual 2 cell embryos fathered by these males analyzed in separate expression microarrays.

Project description:To achieve the extreme nuclear condensation necessary for sperm function, most histones are replaced with protamines during spermiogenesis in mammals. Mature sperm retain only a small fraction of nucleosomes, which are, in part, enriched on gene regulatory sequences, and recent findings suggest that these retained histones provide epigenetic information that regulates expression of a subset of genes involved in embryo development after fertilization. We addressed this tantalizing hypothesis by analyzing two mouse models exhibiting abnormal histone positioning in mature sperm due to impaired poly(ADP-ribose) (PAR) metabolism during spermiogenesis and identified altered sperm histone retention in specific gene loci genome-wide using MNase digestion-based enrichment of mononucleosomal DNA. We then set out to determine the extent to which expression of these genes was altered in embryos generated with these sperm. For control sperm, most genes showed some degree of histone association, unexpectedly suggesting that histone retention in sperm genes is not an all-or-none phenomenon and that a small number of histones may remain associated with genes throughout the genome. The amount of retained histones, however, was altered in many loci when PAR metabolism was impaired. To ascertain whether sperm histone association and embryonic gene expression are linked, the transcriptome of individual 2-cell embryos derived from such sperm was determined using microarrays and RNA sequencing. Strikingly, a moderate but statistically significant portion of the genes that were differentially expressed in these embryos also showed different histone retention in the corresponding gene loci in sperm of their fathers. These findings provide new evidence for the existence of a linkage between sperm histone retention and gene expression in the embryo. 9 WT samples in 3 groups of 3. Each group consists of 3 eggs fertilized by the same father. 9 KO samples in the same setup. Samples are single 2-cell embryos. All mice are in a 129SVE (=129S6/SvEvTac, Taconic) background.

Project description:To achieve the extreme nuclear condensation necessary for sperm function, most histones are replaced with protamines during spermiogenesis in mammals. Mature sperm retain only a small fraction of nucleosomes, which are, in part, enriched on gene regulatory sequences, and recent findings suggest that these retained histones provide epigenetic information that regulates expression of a subset of genes involved in embryo development after fertilization. We addressed this tantalizing hypothesis by analyzing two mouse models exhibiting abnormal histone positioning in mature sperm due to impaired poly(ADP-ribose) (PAR) metabolism during spermiogenesis and identified altered sperm histone retention in specific gene loci genome-wide using MNase digestion-based enrichment of mononucleosomal DNA. We then set out to determine the extent to which expression of these genes was altered in embryos generated with these sperm. For control sperm, most genes showed some degree of histone association, unexpectedly suggesting that histone retention in sperm genes is not an all-or-none phenomenon and that a small number of histones may remain associated with genes throughout the genome. The amount of retained histones, however, was altered in many loci when PAR metabolism was impaired. To ascertain whether sperm histone association and embryonic gene expression are linked, the transcriptome of individual 2-cell embryos derived from such sperm was determined using microarrays and RNA sequencing. Strikingly, a moderate but statistically significant portion of the genes that were differentially expressed in these embryos also showed different histone retention in the corresponding gene loci in sperm of their fathers. These findings provide new evidence for the existence of a linkage between sperm histone retention and gene expression in the embryo. Seven WT (wild-type) samples of one 2-cell embryo each, all from fathers treated with saline (controls). Each group A, B, D consists of eggs fertilized by the same father (each sample is a single 2 cell embryo, 2CE). Ten samples of 2CE from fathers treated with PJ34 (PARP inhibitor) for 6 weeks, in the same setup (PJ-MTM11-1 to PJ34-MTM14-2, offspring from fathers MTM11, MTM13 or MTM14). All mice are wild-type, strain 129SVE (=129S6/SvEvTac, Taconic).

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Gcn5-mediated histone acetylation governs nucleosome dynamics in spermiogenesis [MNase-Seq]

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