Landscape Of Histone Modifications Research Articles

P989: DYNAMIC CHANGES IN THE HISTONE H3 LYSINE 27 TRIMETHYLATION EPIGENETIC LANDSCAPE FOLLOWING RUXOLITINIB ADMINISTRATION

Background: Dysregulation of normal epigenetic processes is increasingly recognised as a feature of myeloid malignancy generally and implicated in the pathogenesis of MPN. Epigenetic changes may directly result from the mutation of key epigenetic regulators including TET2, ASXL1 and EZH2 or indirectly from the activation of intracellular signalling cascades. We have previously observed that ruxolitinib, a JAK1/2 inhibitor, may alter the epigenetic landscape of histone modifications in MPN. CUT&RUN is a novel experimental approach providing high resolution profiling of epigenetic modifications and transcription factor binding. Aims: To comprehensively profile the H3K27me3 epigenetic landscape of JAK2 V617F positive cells resulting from JAK inhibition and determine the dynamic changes occurring between short term and persistent ruxolitinib therapy to identify potential genetic contributors to pathogenesis and options for future therapeutics. Methods:JAK2 V617F positive UKE1 cells were cultured in duplicate in the presence of 1000nM ruxolitinib or DMSO 0.1% vehicle control continuously for approximately five weeks. Samples were harvested at 37 days for CUT&RUN analysis using H3K27me3, EZH2 and IgG (negative control) antibodies. CUT&RUN performed using Epicypher CUTANA Kit according to manufacturer instructions. Libraries prepared using NEBNext® Ultra™ II DNA Library Prep kit with 50 base pair, paired end sequencing undertaken on Illumina NovaSeq 6000. Resulting FASTQ files were trimmed using Trim Galore! and aligned to reference genome hg38 using Bowtie2. Duplicate reads were removed using SAMtools rmdup and peaks called with MACS2 callpeak on the usegalaxy.eu platform. Differential peak analysis undertaken with Diffbind package in R. Samples were normalised to library size prior to differential analysis with DESeq2 methodology. Bigwig files generated using deepTools bamcoverage for visualisation. Results: UKE1 cells remain sensitive to ruxolitinib after a period of treatment with high dose ruxolitinib lasting over 5 weeks. CUT&RUN libraries were sequenced to a minimum depth of 3.6 million unique reads per sample. Initial comparisons between Day 37 high dose treated cells and vehicle control revealed 1004 differentially enriched H3K27me3 peaks. Of these, 545 (54.2%) peaks were increased in the ruxolitinib treated cells and 459 (45.7%) peaks were increased in the control cells. Analysis of the difference between day 37 and day 1 high dose ruxolitinib treated cells revealed 290 peaks differentially enriched. Of these peaks, 279 (96.2%) were increased and 11 (3.8%) were decreased at day 37. Analysis of the overlap between these results identified 33 peaks increased at D37 in comparison to DMSO control and day 1 treatment samples. Analysis of the differential binding patterns and direct visualisation of the results in a genome browser revealed particular genetic locations of interest including the promoter region of SPRY1, a potential regulator of erythropoiesis and JAK signalling, UGCG and BTBD11. Figure 1 shows an example. Differential H3K27me3 was also noted at regions associated with PLXNA1 and NRP2 genes which act as co-receptors for class III semaphorin signalling. Image:Summary/Conclusion: The dynamic changes in the H3K27me3 epigenetic landscape of JAK2 V617F positive cells following persistent high dose ruxolitinib administration has potential therapeutic implications and may sensitise cells to epigenetic therapies. Specific gene loci identified demonstrating differential H3K27me3 may offer some insight into JAK2 V617F mediated pathology and require further investigation.

Active endogenous retroviral elements in human pluripotent stem cells play a role in regulating host gene expression

Human endogenous retroviruses, also called LTR elements, can be bound by transcription factors and marked by different histone modifications in different biological contexts. Recently, individual LTR or certain subclasses of LTRs such as LTR7/HERVH and LTR5_Hs/HERVK families have been identified as cis-regulatory elements. However, there are still many LTR elements with unknown functions. Here, we dissected the landscape of histone modifications and regulatory map of LTRs by integrating 98 ChIP-seq data in human embryonic stem cells (ESCs), and annotated the active LTRs enriching enhancer/promoter-related histone marks. Notably, we found that MER57E3 functionally acted as proximal regulatory element to activate respective ZNF gene. Additionally, HERVK transcript could mainly function in nucleus to activate the adjacent genes. Since LTR5_Hs/LTR5 was bound by many early embryo-specific transcription factors, we further investigated the expression dynamics in different pluripotent states. LTR5_Hs/LTR5/HERVK exhibited higher expression level in naïve ESCs and extended pluripotent stem cells (EPSCs). Functionally, the LTR5_Hs/LTR5 with high activity could serve as a distal enhancer to regulate the host genes. Ultimately, our study not only provides a comprehensive regulatory map of LTRs in human ESCs, but also explores the regulatory models of MER57E3 and LTR5_Hs/LTR5 in host genome.

Histone Modification Landscapes as a Roadmap for Malaria Parasite Development.

Plasmodium falciparum remains the deadliest parasite species in the world, responsible for 229 million cases of human malaria in 2019. The ability of the P. falciparum parasite to progress through multiple life cycle stages and thrive in diverse host and vector species hinges on sophisticated mechanisms of epigenetic regulation of gene expression. Emerging evidence indicates such epigenetic control exists in concentric layers, revolving around core histone post-translational modification (PTM) landscapes. Here, we provide a necessary update of recent epigenome research in malaria parasites, focusing specifically on the ability of dynamic histone PTM landscapes to orchestrate the divergent development and differentiation pathways in P. falciparum parasites. In addition to individual histone PTMs, we discuss recent findings that imply functional importance for combinatorial PTMs in P. falciparum parasites, representing an operational histone code. Finally, this review highlights the remaining gaps and provides strategies to address these to obtain a more thorough understanding of the histone modification landscapes that are at the center of epigenetic regulation in human malaria parasites.

Chromatin domain alterations linked to 3D genome organization in a large cohort of schizophrenia and bipolar disorder brains.

Chromosomal organization, scaling from the 147 base pair nucleosome to megabase-ranging domains encompassing multiple transcriptional units including heritability loci for psychiatric traits, remains largely unexplored in the human brain. Here, we construct promoter and enhancer enriched nucleosomal histone modification landscapes for adult prefrontal cortex (PFC) from H3-lysine 27 acetylation and H3-lysine 4 trimethylation profiles, generated from (n=739) 388 controls and 351 subjects diagnosed with schizophrenia (SCZ) or bipolar disorder (BD). We mapped thousands of cis-regulatory domains (CRDs), revealing fine-grained, 104-106 bp chromosomal organization, firmly integrated into Hi-C topologically associating domain (TAD) stratification by open/repressive chromosomal environments and nuclear topography. Large clusters of hyperacetylated CRDs were enriched for SCZ heritability, with prominent representation of regulatory sequences governing fetal development and glutamatergic neuron signaling. Therefore, SCZ and BD brains show coordinated dysregulation of risk-associated regulatory sequences assembled into kilo- to megabase-scaling chromosomal domains.

Therapeutical interference with the epigenetic landscape of germ cell tumors: a comparative drug study and new mechanistical insights

BackgroundType II germ cell tumors (GCT) are the most common solid cancers in males of age 15 to 35 years. Treatment of these tumors includes cisplatin-based therapy achieving high cure rates, but also leading to late toxicities. As mainly young men are suffering from GCTs, late toxicities play a major role regarding life expectancy, and the development of therapy resistance emphasizes the need for alternative therapeutic options. GCTs are highly susceptible to interference with the epigenetic landscape; therefore, this study focuses on screening of drugs against epigenetic factors as a treatment option for GCTs.ResultsWe present seven different epigenetic inhibitors efficiently decreasing cell viability in GCT cell lines including cisplatin-resistant subclones at low concentrations by targeting epigenetic modifiers and interactors, like histone deacetylases (Quisinostat), histone demethylases (JIB-04), histone methyltransferases (Chaetocin), epigenetic readers (MZ-1, LP99) and polycomb-repressive complexes (PRT4165, GSK343). Mass spectrometry-based analyses of the histone modification landscape revealed effects beyond the expected mode-of-action of each drug, suggesting a wider spectrum of activity than initially assumed. Moreover, we characterized the effects of each drug on the transcriptome of GCT cells by RNA sequencing and found common deregulations in gene expression of ion transporters and DNA-binding factors. A kinase array revealed deregulations of signaling pathways, like cAMP, JAK-STAT and WNT.ConclusionOur study identified seven drugs against epigenetic modifiers to treat cisplatin-resistant GCTs. Further, we extensively analyzed off-target effects and modes-of-action, which are important for risk assessment of the individual drugs.

Chromatin Immunoprecipitation Followed by Next-Generation Sequencing (ChIP-Seq) Analysis in Ewing Sarcoma.

ChIP-seq is the method of choice for profiling protein-DNA interactions, and notably for characterizing the landscape of transcription factor binding and histone modifications. This technique has been widely used to study numerous aspects of tumor biology and led to the development of several promising cancer therapies. In Ewing sarcoma research, ChIP-seq provided important insights into the mechanism of action of the major oncogenic fusion protein EWSR1-FLI1 and related epigenetic and transcriptional changes. In this chapter, we provide a detailed pipeline to analyze ChIP-seq experiments from the preprocessing of raw data to tertiary analysis of detected binding sites. We also advise on best practice to prepare tumor samples prior to sequencing.

Abstract PO-001: xCT expression alters the epigenome and induces genomic instability

Abstract Metabolic reprogramming, remodeling of the epigenetic landscape, and genomic instability are three processes that drive tumorigenesis. While each has been investigated individually, there is a growing interest in the interplay between each process during early cancer development. High expression of the cystine/glutamate antiporter SLC7A11 (xCT) is frequently seen in lung cancer. Our lab previously demonstrated overexpression of xCT in normal bronchial airway epithelial cells is sufficient to induce metabolic reprogramming. In this study, we investigated the effects of xCT overexpression in normal bronchial airway epithelial cells. We hypothesize that xCT overexpression both induces genomic instability and alters the epigenome in normal bronchial airway epithelial cells through the perturbation of intracellular metabolite levels. Through utilizing an alkaline comet assay, we discovered that xCT overexpression induces DNA damage, increasing the percentage of DNA in the comet tail by 15.1% (P=0.002). Furthermore, we found that xCT overexpression induces alterations to the histone modification landscape, most notably increasing H3K9me3 and H3K9ac by 3.2 and 3.9-fold (P=0.08) respectively. Finally, xCT overexpression shows higher tumorigenic potential through increased colony formation in a soft agar assay. Taken together, xCT overexpression in normal bronchial airway epithelial cells increases tumorigenic potential, induces metabolic reprogramming, causes higher levels of DNA damage, and alters the epigenetic landscape. Future experiments elucidating the links between these processes, including ChIP-seq and whole-genome bisulfite sequencing are currently underway. Establishing causal links between metabolic reprogramming, epigenetic alterations, and genomic instability are imperative in elucidating an effective, potentially multifaceted approach to treating early-stage lung cancer. Citation Format: Dalton Hill, Jamshedur Rahman, Christien Kluwe, Pierre P. Massion. xCT expression alters the epigenome and induces genomic instability [abstract]. In: Abstracts: AACR Special Virtual Conference on Epigenetics and Metabolism; October 15-16, 2020; 2020 Oct 15-16. Philadelphia (PA): AACR; Cancer Res 2020;80(23 Suppl):Abstract nr PO-001.

H4K20 Methylation Is Differently Regulated by Dilution and Demethylation in Proliferating and Cell-Cycle-Arrested Xenopus Embryos.

DNA replication during cell division leads to dilution of histone modifications and can thus affect chromatin-mediated gene regulation, raising the question of how the cell-cycle shapes the histone modification landscape, particularly during embryogenesis. We tackled this problem by manipulating the cell cycle during early Xenopus laevis embryogenesis and analyzing invivo histone H4K20 methylation kinetics. The global distribution of un-, mono-, di-, and tri-methylated histone H4K20 was measured by mass spectrometry in normal and cell-cycle-arrested embryos over time. Using multi-start maximum likelihood optimization and quantitative model selection, we found that three specific biological methylation rate constants were required to explain the measured H4K20 methylation state kinetics. While demethylation is essential for regulating H4K20 methylation kinetics in non-cycling cells, demethylation is very likely dispensable in rapidly dividing cells of early embryos, suggesting that cell-cycle-mediated dilution of H4K20 methylation is an essential regulatory component for shaping its epigenetic landscape during early development. A record of this paper's transparent peer review process is included in the Supplemental Information.

Dynamic regulation of histone modifications and long-range chromosomal interactions during postmitotic transcriptional reactivation.

During mitosis, transcription of genomic DNA is dramatically reduced, before it is reactivated during nuclear reformation in anaphase/telophase. Many aspects of the underlying principles that mediate transcriptional memory and reactivation in the daughter cells remain unclear. Here, we used ChIP-seq on synchronized cells at different stages after mitosis to generate genome-wide maps of histone modifications. Combined with EU-RNA-seq and Hi-C analyses, we found that during prometaphase, promoters, enhancers, and insulators retain H3K4me3 and H3K4me1, while losing H3K27ac. Enhancers globally retaining mitotic H3K4me1 or locally retaining mitotic H3K27ac are associated with cell type-specific genes and their transcription factors for rapid transcriptional activation. As cells exit mitosis, promoters regain H3K27ac, which correlates with transcriptional reactivation. Insulators also gain H3K27ac and CCCTC-binding factor (CTCF) in anaphase/telophase. This increase of H3K27ac in anaphase/telophase is required for posttranscriptional activation and may play a role in the establishment of topologically associating domains (TADs). Together, our results suggest that the genome is reorganized in a sequential order, in which histone methylations occur first in prometaphase, histone acetylation, and CTCF in anaphase/telophase, transcription in cytokinesis, and long-range chromatin interactions in early G1. We thus provide insights into the histone modification landscape that allows faithful reestablishment of the transcriptional program and TADs during cell division.

Domain Model Explains Propagation Dynamics and Stability of Histone H3K27 and H3K36 Methylation Landscapes

Chromatin states must be maintained during cell proliferation to uphold cellular identity and genome integrity. Inheritance of histone modifications is central in this process. However, the histone modification landscape is challenged by incorporation of new unmodified histones during each cell cycle, and the principles governing heritability remain unclear. We take a quantitative computational modeling approach to describe propagation of histone H3K27 and H3K36 methylation states. We measure combinatorial H3K27 and H3K36 methylation patterns by quantitative mass spectrometry on subsequent generations of histones. Using model comparison, we reject active global demethylation and invoke the existence of domains defined by distinct methylation endpoints. We find that H3K27me3 on pre-existing histones stimulates the rate of de novo H3K27me3 establishment, supporting a read-write mechanism in timely chromatin restoration. Finally, we provide a detailed quantitative picture of the mutual antagonism between H3K27 and H3K36 methylation and propose that it stabilizes epigenetic states across cell division.

Maintenance of active chromatin states by HMGN2 is required for stem cell identity in a pluripotent stem cell model

BackgroundMembers of the HMGN protein family modulate chromatin structure and influence epigenetic modifications. HMGN1 and HMGN2 are highly expressed during early development and in the neural stem/progenitor cells of the developing and adult brain. Here, we investigate whether HMGN proteins contribute to the chromatin plasticity and epigenetic regulation that is essential for maintaining pluripotency in stem cells.ResultsWe show that loss of Hmgn1 or Hmgn2 in pluripotent embryonal carcinoma cells leads to increased levels of spontaneous neuronal differentiation. This is accompanied by the loss of pluripotency markers Nanog and Ssea1, and increased expression of the pro-neural transcription factors Neurog1 and Ascl1. Neural stem cells derived from these Hmgn-knockout lines also show increased spontaneous neuronal differentiation and Neurog1 expression. The loss of HMGN2 leads to a global reduction in H3K9 acetylation, and disrupts the profile of H3K4me3, H3K9ac, H3K27ac and H3K122ac at the Nanog and Oct4 loci. At endodermal/mesodermal genes, Hmgn2-knockout cells show a switch from a bivalent to a repressive chromatin configuration. However, at neuronal lineage genes whose expression is increased, no epigenetic changes are observed and their bivalent states are retained following the loss of HMGN2.ConclusionsWe conclude that HMGN1 and HMGN2 maintain the identity of pluripotent embryonal carcinoma cells by optimising the pluripotency transcription factor network and protecting the cells from precocious differentiation. Our evidence suggests that HMGN2 regulates active and bivalent genes by promoting an epigenetic landscape of active histone modifications at promoters and enhancers.

Inositol phosphates and core subunits of the Sin3L/Rpd3L histone deacetylase (HDAC) complex up-regulate deacetylase activity

The constitutively nuclear histone deacetylases (HDACs) 1, 2, and 3 erase acetyl marks on acetyllysine residues, alter the landscape of histone modifications, and modulate chromatin structure and dynamics and thereby crucially regulate gene transcription in higher eukaryotes. Nuclear HDACs exist as at least six giant multiprotein complexes whose nonenzymatic subunits confer genome targeting specificity for these enzymes. The deacetylase activity of HDACs has been shown previously to be enhanced by inositol phosphates, which also bridge the catalytic domain in protein-protein interactions with SANT (Swi3, Ada2, N-Cor, and TFIIIB) domains in all HDAC complexes except those that contain the Sin3 transcriptional corepressors. Here, using purified recombinant proteins, coimmunoprecipitation and HDAC assays, and pulldown and NMR experiments, we show that HDAC1/2 deacetylase activity in one of the most ancient and evolutionarily conserved Sin3L/Rpd3L complexes is inducibly up-regulated by inositol phosphates but involves interactions with a zinc finger motif in the Sin3-associated protein 30 (SAP30) subunit that is structurally unrelated to SANT domains, indicating convergent evolution at the functional level. This implies that this mode of regulation has evolved independently multiple times and provides an evolutionary advantage. We also found that constitutive association with another core subunit, Rb-binding protein 4 chromatin-binding factor (RBBP4), further enhances deacetylase activity, implying both inducible and constitutive regulatory mechanisms within the same HDAC complex. Our results indicate that inositol phosphates stimulate HDAC activity and that the SAP30 zinc finger motif performs roles similar to that of the unrelated SANT domain in promoting the SAP30-HDAC1 interaction and enhancing HDAC activity.

Abstract 1842: A novel therapeutic approach to overcome resistance to glutaminase inhibition in NF1 driven malignant peripheral nerve sheath tumors MPNST

Abstract Glutamine, one of the most abundant intracellular amino acids, plays an important role in satisfying the biosynthetic needs of proliferating cancer cells by providing carbons to produce tricarboxylic acid (TCA) cycle intermediates, glutathione, fatty acids, and nucleotides. Though recent research has focused on targeting novel pathways in cellular metabolism, targeting of dysregulated metabolic pathways in therapeutic resistance, a highly clinical relevant area in cancer research, has not been specifically addressed. In this study we hypothesized that combinatorial targeting of metabolism and epigenetics may overcome therapeutic resistance to glutaminase (GLS1) inhibitor, CB-839, in NF1 driven MPNST cells. To test this, we cultured NF1-null MPNST cell line (ST8814) sensitive to growth inhibition by CB-839 with the drug in a dose-escalating manner until the emergence of resistant cell populations. Cell viability assays and western blot analysis demonstrated that CB-839-induced inhibition of cell growth and apoptosis (PARP cleavage) were indeed abrogated in resistant ST8814 cells. To rule out the possibility that acquired resistance was due to changes in the uptake, metabolism or target inhibition of CB-839, we also confirmed that the resistant cells were no longer sensitive to growth inhibition and cell death induced by siRNA knockdown of GLS1. We profiled global landscape of histone modifications in ST8814 parental and resistant cells and found that there was a marked decrease in the repressive histone methylation marks H3K9me3 and H3K27me3 and an increase in H3K9ac, a modification associated with active transcription. Accordingly, levels of c-Myc were significantly elevated in resistant cells. RNAi-mediated c-Myc knockdown selectively inhibited growth factor signaling and cell proliferation in CB-839 resistant cells, suggesting that these cells are addicted to the reprogrammed epigenetic and transcriptional state. Expression levels of c-Myc and histone acetylation are connected by the Bromo-domain protein BRD4. Consistently, CB-839 resistant cells demonstrated augmented sensitivity towards treatment with JQ1, a small molecule inhibitor of BRD4. We also carried out receptor tyrosine kinase (RTK) array and immunoblotting analysis which demonstrated strong overexpression and activation of c-Met (HGF-R) in CB-839 resistant cells, compared to the parental cells. Conversely, combination treatment with inhibitors of c-Myc and c-MET resulted in decreased cell viability, and induction of apoptotic markers such as cleaved PARP and cleaved Caspase3 in the resistant ST8814 cells. Taken together, our data strongly indicates that combinatorial targeting of metabolism and epigenetics is a novel potential approach to overcome CB-839 resistance and merits evaluation in the treatment of drug resistant NF1 driven soft tissue sarcoma. Citation Format: Tahir Sheikh, Parag Patwardhan, Chao Lu, Gary K. Schwartz. A novel therapeutic approach to overcome resistance to glutaminase inhibition in NF1 driven malignant peripheral nerve sheath tumors MPNST [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1842.

DeepHistone: a deep learning approach to predicting histone modifications

MotivationQuantitative detection of histone modifications has emerged in the recent years as a major means for understanding such biological processes as chromosome packaging, transcriptional activation, and DNA damage. However, high-throughput experimental techniques such as ChIP-seq are usually expensive and time-consuming, prohibiting the establishment of a histone modification landscape for hundreds of cell types across dozens of histone markers. These disadvantages have been appealing for computational methods to complement experimental approaches towards large-scale analysis of histone modifications.ResultsWe proposed a deep learning framework to integrate sequence information and chromatin accessibility data for the accurate prediction of modification sites specific to different histone markers. Our method, named DeepHistone, outperformed several baseline methods in a series of comprehensive validation experiments, not only within an epigenome but also across epigenomes. Besides, sequence signatures automatically extracted by our method was consistent with known transcription factor binding sites, thereby giving insights into regulatory signatures of histone modifications. As an application, our method was shown to be able to distinguish functional single nucleotide polymorphisms from their nearby genetic variants, thereby having the potential to be used for exploring functional implications of putative disease-associated genetic variants.ConclusionsDeepHistone demonstrated the possibility of using a deep learning framework to integrate DNA sequence and experimental data for predicting epigenomic signals. With the state-of-the-art performance, DeepHistone was expected to shed light on a variety of epigenomic studies. DeepHistone is freely available in https://github.com/QijinYin/DeepHistone.

Construction of a Synthetic, Chromatin-Based Epigenetic System in Human Cells

The chromatin landscape of DNA and histone modifications represents a fundamental layer of cellular regulation. These modifications, which are implicated in gene regulation and even complex properties such as epigenetic programs, allow eukaryotic organisms to expand information content beyond DNA sequence. Despite its rich regulatory potential, our ability to manipulate this landscape in living cells remains limited. Here we exploit DNA adenine methylation (m6A), a modification rarely found in metazoan genomes, to construct a fully synthetic chromatin system in human cells. Our system is composed of three functional modules that mediate m6A operations: (1) synthetic initiator module to place m6A at specific genomic sites; (2) synthetic readout module to recognize m6A and mediate m6A-dependent transcriptional logic; (3) propagation module that implements “read-write”, a mechanism proposed to underlie chromatin spreading and epigenetic maintenance across cellular systems. Together with a quantitative model, we combine these modules to construct regulatory circuits that drive spatial m6A propagation to regulate distal genes, and enable epigenetic memory of transcriptional states. These epigenetic functions are engineered de novo and do not make use of endogenous mechanisms, providing evidence that complex behaviors like memory can be programmed using specific molecular mechanisms at the chromatin level. Our work establishes a synthetic, chromatin ‘programming language’ to expand the regulatory potential of the genome and engineer epigenetic functions in mammalian cells.

Chorioamnionitis exposure remodels the unique histone modification landscape of neonatal monocytes and alters the expression of immune pathway genes.

Chorioamnionitis is an intrauterine infection involving inflammation of the chorion, amnion, and placenta. It leads to a fetal systemic inflammatory response that can alter the transcription of neonatal immune genes. We have previously shown that neonatal monocytes gain the activating histone tail modification H3K4me3 at promoter sites of immunologically important genes as development progresses from preterm neonate to adult. In this study, we applied ChIP-seq and RNA-seq to evaluate the impact of chorioamnionitis on the neonatal monocyte H3K4me3 histone modification landscape over the course of fetal and neonatal immune system development. Chorioamnionitis exposure in neonatal monocytes resulted in a net increase in total monocyte H3K4me3, primarily in introns and intergenic regions. Immune gene expression was decreased in chorioamnionitis-exposed monocytes, with the majority of enriched transcripts falling into pathways that are not linked to the immune system. Over half of all neonatal monocyte H3K4me3 peaks, independent of their location, were associated with active gene transcription. Overall, chorioamnionitis exposure resulted in the global remodeling of the neonatal monocyte H3K4me3 landscape and changes in the expression of known immune genes. These changes resulted in a less robust inflammatory response upon exposure to a secondary challenge, which may explain why chorioamnionitis-exposed neonates have an increased risk of sepsis. DATABASE: ChIP-seq data for U30/O30/Term: GEO GSE81957 ChIP-seq data for U30C/O30C/TermC: GEO GSE111873 RNA-seq data for U/L/CU/CL: GEO GSE111927.

Insights from multidimensional analyses of the pan-cancer DNA methylome heterogeneity and the uncanonical CpG-gene associations.

Although the DNA methylome profiles have been available in large cancer cohorts such as The Cancer Genome Atlas (TCGA), integrative analysis of the DNA methylome architectures in a pan-cancer manner remains limited. In the present study, we aimed to systematically dissect the insightful features related to the inter-tumoral DNA methylome heterogeneity in a pan-cancer context of 21 cancers in TCGA. First, pan-cancer clustering of the DNA methylomes revealed convergence of cancers and, meanwhile, new classifications of cancer subtypes, which are often associated to prognostic differences. Next, within each type of cancer, we showed that the transcription factor (TF) genes tend to bear more dynamic promoter DNA methylation profiles than the other genes, which serves as a potential source of the transcriptome heterogeneity in cancers. Finally, we found unanticipated significant numbers of the non-canonical promoter CpG sites that are positively correlated with the gene expression. Distribution patterns of these CpG sites in the CpG islands, ChIP-seq, DNaseI-seq, PMD regions and histone modification landscapes suggested against a pervasive mechanism of transcriptional activation due to mCpG-dependent binding of TFs, which is not in complete agreement with previous hypothesis. In summary, our deep mining of the highly heterogeneous DNA methylome data in a pan-cancer context generated novel insights into the architecture of cancer epigenetics and provided a series of resources for further investigations in the related fields of cancer genomics and epigenetics.

Accurate Recycling of Parental Histones Reproduces the Histone Modification Landscape during DNA Replication

SummaryChromatin organization is disrupted genome-wide during DNA replication. On newly synthesized DNA, nucleosomes are assembled from new naive histones and old modified histones. It remains unknown whether the landscape of histone post-translational modifications (PTMs) is faithfully copied during DNA replication or the epigenome is perturbed. Here we develop chromatin occupancy after replication (ChOR-seq) to determine histone PTM occupancy immediately after DNA replication and across the cell cycle. We show that H3K4me3, H3K36me3, H3K79me3, and H3K27me3 positional information is reproduced with high accuracy on newly synthesized DNA through histone recycling. Quantitative ChOR-seq reveals that de novo methylation to restore H3K4me3 and H3K27me3 levels occurs across the cell cycle with mark- and locus-specific kinetics. Collectively, this demonstrates that accurate parental histone recycling preserves positional information and allows PTM transmission to daughter cells while modification of new histones gives rise to complex epigenome fluctuations across the cell cycle that could underlie cell-to-cell heterogeneity.

The Landscape of Histone Modifications in a High-Fat Diet-Induced Obese (DIO) Mouse Model

The Landscape of Histone Modifications in a High-Fat Diet-Induced Obese (DIO) Mouse Model

The Landscape of Histone Modifications in a High-Fat Diet-Induced Obese (DIO) Mouse Model.

Type 2 diabetes (T2D) is a major chronic healthcare concern worldwide. Emerging evidence suggests that a histone-modification-mediated epigenetic mechanism underlies T2D. Nevertheless, the dynamics of histone marks in T2D have not yet been carefully analyzed. Using a mass spectrometry-based label-free and chemical stable isotope labeling quantitative proteomic approach, we systematically profiled liver histone post-translational modifications (PTMs) in a prediabetic high-fat diet-induced obese (DIO) mouse model. We identified 170 histone marks, 30 of which were previously unknown. Interestingly, about 30% of the histone marks identified in DIO mouse liver belonged to a set of recently reported lysine acylation modifications, including propionylation, butyrylation, malonylation, and succinylation, suggesting possible roles of these newly identified histone acylations in diabetes and obesity. These histone marks were detected without prior affinity enrichment with an antibody, demonstrating that the histone acylation marks are present at reasonably high stoichiometry. Fifteen histone marks differed in abundance in DIO mouse liver compared with liver from chow-fed mice in label-free quantification, and six histone marks in stable isotope labeling quantification. Analysis of hepatic histone modifications from metformin-treated DIO mice revealed that metformin, a drug widely used for T2D, could reverse DIO-stimulated histone H3K36me2 in prediabetes, suggesting that this mark is likely associated with T2D development. Our study thus offers a comprehensive landscape of histone marks in a prediabetic mouse model, provides a resource for studying epigenetic functions of histone modifications in obesity and T2D, and suggest a new epigenetic mechanism for the physiological function of metformin.