Multi-omics approaches for revealing the epigenetic regulation of histone H3.1 during spermatogonial stem cell differentiation in vitro
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE222617 | GEO | 2023/03/01
REPOSITORIES: GEO
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Project description:Nucleosomes package eukaryotic DNA and are composed of four different histone proteins, H3, H4, H2A and H2B. Histone H3 has two main variants, H3.1 and H3.3, which show different genomic localization patterns in animals. We profiled H3.1 and H3.3 variants in the genome of the plant Arabidopsis thaliana and show that the localization of these variants shows broad similarity in plants and animals, in addition to some unique features. H3.1 was enriched in silent areas of the genome including regions containing the repressive chromatin modifications H3 lysine 27 methylation, H3 lysine 9 methylation, and DNA methylation. In contrast, H3.3 was enriched in actively transcribed genes, especially peaking at the 3’ end of genes, and correlated with histone modifications associated with gene activation such as histone H3 lysine 4 methylation, and H2B ubiquitylation, as well as by RNA Pol II occupancy. Surprisingly, both H3.1 and H3.3 were enriched on defined origins of replication, as was overall nucleosome density, suggesting a novel characteristic of plant origins. Our results are broadly consistent with the hypothesis that H3.1 acts as the canonical histone that is incorporated during DNA replication, whereas H3.3 acts as the replacement histone that can be incorporated outside of S-phase during chromatin disrupting processes like transcription. ChIP-seq - 4 samples: 2 experiment and 2 controls RNA-seq - 1 sample
2012-03-05 | E-GEOD-34840 | biostudies-arrayexpress
Project description:The histone H3.1 variant is specifically inserted on chromatin during DNA replication in eukaryotes. A single amino acid variation (position 31) in the N-terminal tail of H3.1 compared to replication-independent H3.3 is conserved in plants and animals, but no function has been assigned to amino acid 31 to justify a role for H3.1 during replication. In this study, we demonstrate that the protein TONSOKU (TSK/TONSL), which participates in rescuing broken replication forks, specifically interacts with H3.1 via recognition of alanine 31 by its tetratricopeptide repeat (TPR) domain. We show that genomic amplification in the absence of H3K27me1 in plants depends on H3.1, TSK and DNA polymerase theta (Pol θ). Overall, our study reveals similar strategies in plants and animals for regulating TSK activity and post-replicative chromatin maturation, with both clades using the H3.1 variant and different histone mono-methyltransferases (ATXR5/ATXR6 or SET8) to achieve this.
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Project description:Nucleosomes package eukaryotic DNA and are composed of four different histone proteins, H3, H4, H2A and H2B. Histone H3 has two main variants, H3.1 and H3.3, which show different genomic localization patterns in animals. We profiled H3.1 and H3.3 variants in the genome of the plant Arabidopsis thaliana and show that the localization of these variants shows broad similarity in plants and animals, in addition to some unique features. H3.1 was enriched in silent areas of the genome including regions containing the repressive chromatin modifications H3 lysine 27 methylation, H3 lysine 9 methylation, and DNA methylation. In contrast, H3.3 was enriched in actively transcribed genes, especially peaking at the 3’ end of genes, and correlated with histone modifications associated with gene activation such as histone H3 lysine 4 methylation, and H2B ubiquitylation, as well as by RNA Pol II occupancy. Surprisingly, both H3.1 and H3.3 were enriched on defined origins of replication, as was overall nucleosome density, suggesting a novel characteristic of plant origins. Our results are broadly consistent with the hypothesis that H3.1 acts as the canonical histone that is incorporated during DNA replication, whereas H3.3 acts as the replacement histone that can be incorporated outside of S-phase during chromatin disrupting processes like transcription.
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Project description:We developed a new sequencing assay to track the de novo deposition of the histone H3 variants H3.1 and H3.3 during S phase. We use cells stably expressing H3.1-SNAP or H3.3-SNAP, and synchronize them in G1/S by double-thymidine block. The SNAP-tag enables to discriminate newly synthesized histones from preexisting ones, via a quench-chase-capture strategy. We applied this strategy to isolate new H3.1 and H3.3 after releasing cells into S phase, and probed their distribution by MNase digestion and sequencing. We could thus characterize H3.1 and H3.3 dynamics from early to mid S phase at genome-wide resolution. We further applied our method to investigate the consequences of perturbations upon deletion of the H3.3 chaperone HIRA. We used HIRA knockout and control cells, and compared H3.1 and H3.3 distribution to early replication patterns by EdU labeling and sequencing of nascent DNA.
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Project description:The replication-dependent canonical histone genes are the only genes found in multicellular organisms whose messenger RNA (mRNA) does not terminate with a poly(A) tail at the 3’ end. We previously demonstrated that arsenic exposure induces polyadenylation of canonical histone H3.1 mRNA in tissue culture system. We report that arsenic is capable of inducing high levels of H3.1 mRNA polyadenylation and marked loss of the Stem-loop binding protein (SLBP) in vivo, which is critical for canonical histone pre-mRNA processing. Ectopic expression of polyadenylated H3.1 mRNA enhances anchorage-independent cell growth and tumor formation in nude mice. Moreover, polyadenylation of H3.1 mRNA causes transcriptional deregulation, G2/M arrest, chromosome aneuploidy and aberration. Importantly, overexpression of H3.3 attenuates arsenic-induced cell transformation. Through ChIP-Seq and RNA-Seq, we try to decipher if polyadenylated H3.1 mRNA could result in depletion of histone variant H3.3 at critical gene regulatory regions. Our study uncovers polyadenylation of H3.1 and resulting displacement of variant H3.3 as a potential new mechanism for arsenic-induced carcinogenesis.
2020-09-02 | GSE135637 | GEO
Project description:Establishment of a proper chromatin landscape is central to genome function. Here, we explain H3 variant distribution by specific targeting and dynamics of deposition involving the CAF-1 and HIRA histone chaperones. Impairing replicative H3.1 incorporation via CAF-1 enables an alternative H3.3 deposition at replication sites via HIRA. Conversely, the H3.3 incorporation throughout the cell cycle via HIRA cannot be replaced by H3.1. ChIP-seq analyses reveal correlation between HIRA-dependent H3.3 accumulation and RNA pol II at transcription sites and specific regulatory elements, further supported by their biochemical association. Remarkably, the HIRA complex shows unique DNA binding properties and depleting HIRA increases DNA sensitivity to nucleases. We propose that protective gap-filling of naked DNA by HIRA leads to a broad distribution of H3.3, and HIRA association with Pol II ensures local H3.3 enrichment at specific sites. Examination of genome-wide localization of two histone H3 variants.
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Project description:We present here evidence that histones H3.1 and H4 can be imported into the nucleus as monomers in human cells. Using a tether-and-release system to study the cytosolic phase and import dynamics of newly synthesised histones, we find that H3.1 and H4 can be maintained as stable monomers in the cytosol in a tethered state. Cytosolically tethered histones are bound tightly to Importin- proteins (predominantly IPO4), but not to the histone specific chaperones NASP, ASF1a, RbAp46 (RBBP7) or HAT1, which reside in the nucleus in interphase cells. Release of monomeric histones from their cytosolic tether results in rapid nuclear translocation, dissociation with IPO4 and incorporation into chromatin at sites of replication. Quantitative analysis of histones bound to individual chaperones under steady-state conditions reveals an excess of H3 specifically associated with sNASP, suggesting that NASP can maintain a soluble, monomeric pool of H3 within the nucleus and may act as a nuclear receptor for newly imported histone. In summary, we propose that histones H3 and H4 are rapidly imported as monomeric units, forming heterodimers in the nucleus rather than the cytosol, with sNASP acting as a potential nuclear receptor for monomeric histone H3.
2018-09-10 | PXD009915 | Pride
Project description:How histone posttranslational modifications (PTMs) are inherited through cell cycle remains poorly understood. Canonical histones are made in the S phase of cell cycle. Combining mass spectrometry-based technologies and stable isotope labeling by amino acids in cell culture (SILAC), we interrogate the distributions of multiple major histone PTMs on old versus new histones in synchronized human cells. We show that histone PTMs can be grouped to three categories accordingly to their distributions. Most lysine mono-methylation and acetylation PTMs are either symmetrically distributed on old and new histones, or asymmetric on new histones. In contrast, most di- and tri-methylation PTMs are asymmetric on old histones, suggesting the inheritance of different PTMs is regulated distinctly. Intriguingly, old and new histones are distinguished in their phosphorylation status during early mitosis, in three different human cell types: Hela, 293T and human foreskin fibroblast (HFF) cells. The mitotic hallmark H3S10ph is predominantly associated with old H3 at early mitosis and become symmetric with the progression of mitosis. The same distribution pattern is found on other mitotic histone phosphorylation marks, including H3T3/T6ph, H3.1/2S28ph and H1.4S26ph, excluding S28/S31ph on the H3 variant H3.3. Although H3S10ph often associates with the neighboring K9 di- or tri-methylation, they are not required for the asymmetric distribution of S10ph on the same H3 tail. Inhibition of the kinase Aurora B does not change the distribution pattern despite significant reduction of H3S10ph levels. K9me2 abundance on the new H3 is significantly reduced after Aurora B inhibition, suggesting a crosstalk between H3S10ph and H3K9me2.
2016-06-01 | PXD003709 | Pride
Project description:Transcriptional profiling of mouse spermatogonial stem cells (SSCs) comparing control untreated SSCs with SSCs with exogenous FGF2 withdrawn and FGFR inhibitor SU5402 supplemented (-F+S). Results provide insight into the mechanisms of FGF2-supported in vitro self-renewal of SSCs. Two-condition experiment, SSCs-F+S vs. SSCs. Biological replicates: 4 control replicates, 4 -F+S replicates.
2010-08-01 | E-GEOD-21294 | biostudies-arrayexpress
Project description:Background: Spermatogonial stem cells (SSCs) sustain the process of steady-state spermatogenesis in the mammalian testis, which is critical to the ongoing production of sperm and male fertility. SSCs can both self-renew to perpetuate the SSC population, and give rise to progenitors to propel the spermatogenic differentiation pathway. SSCs are detectable as a small subpopulation of undifferentiated spermatogonia that demonstrate regenerative capacity in a transplantation assay, and can be selectively recovered on the basis of expression of an Id4-eGfp sortable marker transgene. Enriched populations of SSCs and progenitors display consistent differences in gene expression patterns suggesting they represent discrete spermatogonial subtypes. Results: Here we describe distinct spermatogonial subtype-specific epigenetic programming profiles associated with subtype-specific differential gene expression on the basis of genome-wide patterns of six different histone modifications, chromatin accessibility, and DNA methylation. We find that similarly expressed genes show no differences in epigenetic programming, whereas differentially expressed genes show distinct histone modification patterns as well as subtype-specific differences in patterns of distal intergenic low-methylated regions. Motif-enrichment analysis of differentially programmed elements predicts a set of transcription factors that may regulate this spermatogonial subtype-specific epigenetic programming, and gene-specific chromatin immunoprecipitation analyses confirm subtype-specific differences in binding of a subset of these factors to target genes. Conclusions: Taken together, these results indicate that SSCs and progenitors are discrete spermatogonial subtypes that appear to be differentially programmed to carry out mutually exclusive functions including self-renewal and maintenance of regenerative capacity in SSCs and lineage commitment and loss of regenerative capacity in progenitors.
2019-05-23 | GSE131654 | GEO