Chromatin Architecture Research Articles

Epigenetic modulation via the C-terminal tail of H2A.Z

H2A.Z-nucleosomes are present in both euchromatin and heterochromatin and it has proven difficult to interpret their disparate roles in the context of their stability features. Using an in situ assay of nucleosome stability and DT40 cells expressing engineered forms of the histone variant we show that native H2A.Z, but not C-terminally truncated H2A.Z (H2A.Z∆C), is released from nucleosomes of peripheral heterochromatin at unusually high salt concentrations. H2A.Z and H3K9me3 landscapes are reorganized in H2A.Z∆C-nuclei and overall sensitivity of chromatin to nucleases is increased. These tail-dependent differences are recapitulated upon treatment of HeLa nuclei with the H2A.Z-tail-peptide (C9), with MNase sensitivity being increased genome-wide. Fluorescence correlation spectroscopy revealed C9 binding to reconstituted nucleosomes. When introduced into live cells, C9 elicited chromatin reorganization, overall nucleosome destabilization and changes in gene expression. Thus, H2A.Z-nucleosomes influence global chromatin architecture in a tail-dependent manner, what can be modulated by introducing the tail-peptide into live cells.

Acetylation-Dependent Compaction of the Histone H4 Tail Ensemble.

Acetylation of the histone H4 tail (H4Kac) has been established as a significant regulator of chromatin architecture and accessibility; however, the molecular mechanisms that underlie these observations remain elusive. Here, we characterize the ensemble features of the histone H4 tail and determine how they change following acetylation on specific sets of lysine residues. Our comprehensive account is enabled by a robust combination of experimental and computational biophysical methods that converge on molecular details including conformer size, intramolecular contacts, and secondary structure propensity. We find that acetylation significantly alters the chemical environment of basic patch residues (16-20) and leads to tail compaction that is partially mediated by transient intramolecular contacts established between the basic patch and N-terminal amino acids. Beyond acetylation, we identify that the protonation state of H18, which is affected by the acetylation state, is a critical regulator of ensemble characteristics, highlighting the potential for interplay between the sequence context and post-translational modifications to define the ensemble features of intrinsically disordered regions. This study elucidates molecular details that could link H4Kac with the regulation of chromatin architecture, illuminating a small piece of the complex network of molecular mechanisms underlying the histone code hypothesis.

Droplet Hi-C enables scalable, single-cell profiling of chromatin architecture in heterogeneous tissues.

Current methods for analyzing chromatin architecture are not readily scalable to heterogeneous tissues. Here we introduce Droplet Hi-C, which uses a commercial microfluidic device for high-throughput, single-cell chromatin conformation profiling in droplets. Using Droplet Hi-C, we mapped the chromatin architecture of the mouse cortex and analyzed gene regulatory programs in major cortical cell types. In addition, we used this technique to detect copy number variations, structural variations and extrachromosomal DNA in human glioblastoma, colorectal and blood cancer cells, revealing clonal dynamics and other oncogenic events during treatment. We refined the technique to allow joint profiling of chromatin architecture and transcriptome in single cells, facilitating exploration of the links between chromatin architecture and gene expression in both normal tissues and tumors. Thus, Droplet Hi-C both addresses critical gaps in chromatin analysis of heterogeneous tissues and enhances understanding of gene regulation.

Interaction of CTCF and CTCFL in genome regulation through chromatin architecture during the spermatogenesis and carcinogenesis.

The zinc finger protein CTCF is ubiquitously expressed and is integral to the regulation of chromatin architecture through its interaction with cohesin. Conversely, CTCFL expression is predominantly restricted to the adult male testis but is aberrantly expressed in certain cancers. Despite their distinct expression patterns, the cooperative and competitive mechanisms by which CTCF and CTCFL regulate target gene expression in spermatocytes and cancer cells remain inadequately understood. In this review, we comprehensively examine the literature on the divergent amino acid sequences, target sites, expression profiles and functions of CTCF and CTCFL in normal tissues and cancers. We further elucidate the mechanisms by which CTCFL competitively or cooperatively binds to CTCF target sites during spermatogenesis and carcinogenesis to modulate chromatin architecture. We mainly focus on the role of CTCFL in testicular and cancer development, highlighting its interaction with CTCF at CTCF binding sites to regulate target genes. In the testis, CTCF and CTCFL cooperate to regulate the expression of testis-specific genes, essential for proper germ cell progression. In cancers, CTCFL overexpression competes with CTCF for DNA binding, leading to aberrant gene expression, a more relaxed chromatin state, and altered chromatin loops. By uncovering the roles of CTCF and CTCFL in spermatogenesis and carcinogenesis, we can better understand the implications of aberrant CTCFL expression in altering chromatin loops and its contribution to disease pathogenesis.

Impacts of Hyperglycemia on Epigenetic Modifications in Human Gingival Fibroblasts and Gingiva in Diabetic Rats

Periodontal disease is considered one of the diabetic complications with high morbidity and severity. Recent studies demonstrated the involvement of the epigenome on diabetic complications. Histone modifications change chromatin architecture and gene activation. Histone modifications have been reported to alter chromatin structure and regulate gene transcription. In this study, we investigated the impacts of H3 lysine 4 trimethylation (H3K4me3) and specific histone methyltransferases of H3K4 methylation, su(var)3-9, enhancer-of-zeste, and trithorax domain 1A (SETD1A) on periodontal tissue affected by the diabetic condition. We observed the increase in H3K4me3 and SETD1A in gingival tissue of diabetic rats compared with the normal rats. Cultured human fibroblasts (hGFs) confirmed a high glucose-induced increase in H3K4me3 and SETD1A. We further demonstrated that high glucose increased the gene expression of matrix metalloproteinase (MMP) 1 and MMP13, which were canceled by sinefungin, an SETD1A inhibitor. Our investigation suggests that diabetes triggers histone modifications in the gingival tissue, resulting in gingival inflammation. Histone modifications may play crucial roles in the development of periodontal disease in diabetes.

Acetylation of Histone H3 in Cancer Progression and Prognosis

Cancer is a multifactorial disease resulting from both genetic factors and epigenetic changes. Histone acetylation, a post-translational modification, which alters chromatin architecture and regulates gene expression is associated with cancer initiation, development and progression. Aberrations in global histone acetylation levels are observed in various cancer cells and are also associated with patients’ tumor aggressiveness. Therefore, histone acetylation may have prognostic utility and serve as a potential biomarker of cancer progression and patients’ prognosis. The reversible modification of histones by an acetyl group is versatile. One particular histone can be acetylated on different lysine residues, subsequently resulting in different biological outcomes. Here, we discuss recent findings on the acetylation of the highly conserved histone protein H3 in the context of cancer biology. Specifically, we review the acetylation of particular H3 residues in various cancer types. We further highlight the significance of H3 acetylation levels as a potential cancer biomarker with prognostic implications.

H3K27me3 Loss in Central Nervous System Tumors: Diagnostic, Prognostic, and Therapeutic Implications.

Central nervous system (CNS) tumors represent a formidable clinical challenge due to their molecular complexity and varied prognostic outcomes. This review delves into the pivotal role of the epigenetic marker H3K27me3 in the development and treatment of CNS tumors. H3K27me3, specifically the trimethylation of lysine 27 on the histone H3 protein, plays a crucial role in regulating gene expression and maintaining chromatin architecture (e.g., in X-chromosome inactivation). Notably, a reduction in H3K27me3 levels, frequently tied to mutations in the H3 gene family such as H3F3A and HIST1H3B, is evident in diverse brain tumor variants, including the diffuse midline glioma characterized by the H3K27M mutation and certain pediatric high-grade gliomas. The loss of H3K27me3 has been linked to more aggressive behavior in meningiomas, with the trimethylation loss associated with significantly shorter recurrence-free survival (RFS) among grade 2 meningiomas, albeit not within grade 1 tumors. Pediatric posterior fossa ependymomas characterized by a lowered H3K27me3 and DNA hypomethylation exhibit poor prognosis, underscoring the prognostic significance of these epigenetic alterations in CNS tumors. Comprehending the role of H3K27me3 in CNS tumors is vital for advancing diagnostic tools and therapeutic interventions, with the goal of enhancing patient outcomes and quality of life. This review underscores the importance of ongoing investigations into H3K27me to refine and optimize management strategies for CNS tumors, paving the way for improved personalized medicine practices in oncology.

Genome-wide DNA methylation and their transgenerational pattern differ in Arabidopsis thaliana populations originated along the elevation of West Himalaya

Methylation at 5' cytosine of DNA molecule is an important epigenetic mark. It is known to play critical role in adaptation of organisms under different biotic and abiotic stressors via modulating gene expression and/or chromatin architecture. Plant populations evolved under variable climatic conditions may have evolved different epigenetic marks including DNA methylation. Here we, describe the genome-wide DNA methylation pattern under native field, F1 and F6 generation followed by their association with phenotypes, climate and global gene expression in the three Arabidopsis thaliana populations originated at different elevation ranges of Indian West Himalaya. We show that the global methyl cytosine (mC) content is more or less similar in the three populations but differ in their distribution across genome. There was an increase in differential methylation between the populations as elevation increased. The methylation divergence was the highest between the low and the high elevation populations. The high elevation populations were hypo-methylated than the low elevation population. The methylation in the genes was associated with population specific phenotypes and climate of the region. The genes which were differentially methylated as well as differentially expressed between the low and high elevation populations were mostly related to abiotic stresses. When grown under controlled condition, there was gain of differential methylation over native condition and the maximum percent changes was observed in CHH-sequence context. Further ~ 99.8% methylated cytosines were stably passed on from F1 to F6 generation. Overall, our data suggest that high elevation population is epigenetically more plastic under changing environmental condition.BackgroundArabidopsis thaliana is the model plant species and has been extensively studied to understand plants life processes. There are numerous reports on its origin, demography, evolution, epigenomes and adaptation etc. however, Indian populations of Arabidopsis thaliana evolved along wide elevation ranging from ~ 700 m amsl to ~ 3400 m amsl not explored yet. Here we, describe the genome-wide DNA methylation pattern under native field, F1 and F6 generation followed by their association with phenotypes, climate and global gene expression in the three Arabidopsis thaliana populations originated at different elevation ranges of Indian West Himalaya.Results In our study we found that total mCs percent was more or less similar in the three populations but differ in their distribution across genome. The proportion of CG-mCs was the highest, followed by CHH-mCs and CHG-mCs in all the three populations. Under native field condition the methylation divergence was more prominent between low and high elevation populations and the high elevation populations were hypo-methylated than the low elevation population. The methylation in the genes was linked to population-specific phenotypes and the regional climate. The genes that showed differential methylation and expression between low and high elevation populations were primarily associated with abiotic stress responses. When grown under controlled condition, there was gain of differential methylation compared to the native condition and the maximum percent changes was observed in CHH-sequence context. Further 99.8% methylated cytosines were stably passed on from F1 to F6 generation.Conclusions The populations of A. thaliana adapted at different climatic conditions were significantly differentially methylated both under native and controlled condition. However, the magnitude and extent of gain or loss of methylation were most significant between the low and the high elevation populations. Overall, our data suggest that high elevation population is epigenetically more plastic under changing environmental condition.

A compendium of genetic variations associated with promoter usage across 49 human tissues

Promoters play a crucial role in regulating gene transcription. However, our understanding of how genetic variants influence alternative promoter selection is still incomplete. In this study, we implement a framework to identify genetic variants that affect the relative usage of alternative promoters, known as promoter usage quantitative trait loci (puQTLs). By constructing an atlas of human puQTLs across 49 different tissues from 838 individuals, we have identified approximately 76,856 independent loci associated with promoter usage, encompassing 602,009 genetic variants. Our study demonstrates that puQTLs represent a distinct type of molecular quantitative trait loci, effectively uncovering regulatory targets and patterns. Furthermore, puQTLs are regulating in a tissue-specific manner and are enriched with binding sites of epigenetic marks and transcription factors, especially those involved in chromatin architecture formation. Notably, we have also found that puQTLs colocalize with complex traits or diseases and contribute to their heritability. Collectively, our findings underscore the significant role of puQTLs in elucidating the molecular mechanisms underlying tissue development and complex diseases.

The Epigenetic Hallmarks of Cancer.

Cancer is a complex disease in which several molecular and cellular pathways converge to foster the tumoral phenotype. Notably, in the latest iteration of the cancer hallmarks, "nonmutational epigenetic reprogramming" was newly added. However, epigenetics, much like genetics, is a broad scientific area that deserves further attention due to its multiple roles in cancer initiation, progression, and adaptive nature. Herein, we present a detailed examination of the epigenetic hallmarks affected in human cancer, elucidating the pathways and genes involved, and dissecting the disrupted landscapes for DNA methylation, histone modifications, and chromatin architecture that define the disease. Significance: Cancer is a disease characterized by constant evolution, spanning from its initial premalignant stages to the advanced invasive and disseminated stages. It is a pathology that is able to adapt and survive amidst hostile cellular microenvironments and diverse treatments implemented by medical professionals. The more fixed setup of the genetic structure cannot fully provide transformed cells with the tools to survive but the rapid and plastic nature of epigenetic changes is ready for the task. This review summarizes the epigenetic hallmarks that define the ecological success of cancer cells in our bodies.

Chromatin condensed domains revealed by AFM, and their transformation in mechanically deformed normal and malignant cell nuclei

It has been generally accepted that heterochromatin is represented by a regular, dense and closed structure, while euchromatin is open and sparse. Recent evidence indicates that chromatin is comprised of irregular nucleosome clutches compacted within the nucleus. Transcriptional events transform the chromatin architecture, resulting in appearance of 100–300 nm nucleosomal aggregates. Meanwhile, the current paradigm of chromatin architecture is largely fragmented. In this communication, we unraveled chromatin ultrastructure of normal and malignant cell nuclei through mechanical deformation of the nuclei and Atomic Force Microscopy (AFM) analysis of the resulting landscape. In human skin fibroblasts cell nuclei, nanodomains of about 16.5–33.5 nm were revealed. Hierarchical folding of the chromatin of normal nuclei was observed: the nanodomains formed irregular fiber-like structures that coalesced into the macroscale chromatin compartments. In fibrosarcoma cell nuclei DNA supercoiling domains (SDs) of about 66.3–113.0 nm, uniformly distributed within the nuclei, were revealed. Transformation of the morphology of the condensed chromatin domains through up- and downregulation of supercoiling was demonstrated.

RNA sequestration in P-bodies sustains myeloid leukaemia.

Post-transcriptional mechanisms are fundamental safeguards of progenitor cell identity and are often dysregulated in cancer. Here, we identified regulators of P-bodies as crucial vulnerabilities in acute myeloid leukaemia (AML) through genome-wide CRISPR screens in normal and malignant haematopoietic progenitors. We found that leukaemia cells harbour aberrantly elevated numbers of P-bodies and show that P-body assembly is crucial for initiation and maintenance of AML. Notably, P-body loss had little effect upon homoeostatic haematopoiesis but impacted regenerative haematopoiesis. Molecular characterization of P-bodies purified from human AML cells unveiled their critical role in sequestering messenger RNAs encoding potent tumour suppressors from the translational machinery. P-body dissolution promoted translation of these mRNAs, which in turn rewired gene expression and chromatin architecture in leukaemia cells. Collectively, our findings highlight the contrasting and unique roles of RNA sequestration in P-bodies during tissue homoeostasis and oncogenesis. These insights open potential avenues for understanding myeloid leukaemia and future therapeutic interventions.

Virus Infection Induces Immune Gene Activation with CTCF Anchored Enhancers and Chromatin Interactions in Pig Genome.

Chromatin organization is important for gene transcription in pig genome. However, its three-dimensional (3D) structure and dynamics are much less investigated than those in human. Here we applied the long-reads chromatin interaction analysis by paired-end tag sequencing (ChIA-PET) method to map the whole-genome chromatin interactions mediated by CCCTC-binding factor (CTCF) and RNA polymerase Ⅱ (RNAPⅡ or POLⅡ) in porcine macrophage cells before and after polyinosinic-polycytidylic acid [Poly(I:C)] induction. Our results revealed that Poly(I:C) induction impacts the 3D genome organization in the 3D4/21 cells at the fine-scale chromatin loop level rather than at the large-scale domain level. Furthermore, our findings underscored the pivotal role of CTCF anchored chromatin interactions in reshaping chromatin architecture during immune responses. Knock-out of the CTCF locus further confirmed that the CTCF anchored enhancers are associated with the activation of immune genes via long-range interactions. Notably, ChIA-PET data also supported the spatial relationship between single nucleotide polymorphisms (SNPs) and the related gene transcription in 3D genome aspect. Our findings in this study provide new clues and potential targets to explore key elements related to diseases in swine and are also likely to shed light on elucidating chromatin organization and dynamics underlying the process of mammalian infectious diseases.

Dynamic chromatin architecture identifies new autoimmune-associated enhancers for IL2 and novel genes regulating CD4+ T cell activation.

Genome-wide association studies (GWAS) have identified hundreds of genetic signals associated with autoimmune disease. The majority of these signals are located in non-coding regions and likely impact cis-regulatory elements (cRE). Because cRE function is dynamic across cell types and states, profiling the epigenetic status of cRE across physiological processes is necessary to characterize the molecular mechanisms by which autoimmune variants contribute to disease risk. We localized risk variants from 15 autoimmune GWAS to cRE active during TCR-CD28 co-stimulation of naïve human CD4+ T cells. To characterize how dynamic changes in gene expression correlate with cRE activity, we measured transcript levels, chromatin accessibility, and promoter-cRE contacts across three phases of naive CD4+ T cell activation using RNA-seq, ATAC-seq, and HiC. We identified ~1200 protein-coding genes physically connected to accessible disease-associated variants at 423 GWAS signals, at least one-third of which are dynamically regulated by activation. From these maps, we functionally validated a novel stretch of evolutionarily conserved intergenic enhancers whose activity is required for activation-induced IL2 gene expression in human and mouse, and is influenced by autoimmune-associated genetic variation. The set of genes implicated by this approach are enriched for genes controlling CD4+ T cell function and genes involved in human inborn errors of immunity, and we pharmacologically validated eight implicated genes as novel regulators of T cell activation. These studies directly show how autoimmune variants and the genes they regulate influence processes involved in CD4+ T cell proliferation and activation.

Dynamic chromatin architecture identifies new autoimmune-associated enhancers for IL2 and novel genes regulating CD4+ T cell activation

Dynamic chromatin architecture identifies new autoimmune-associated enhancers for IL2 and novel genes regulating CD4+ T cell activation

Fasting shapes chromatin architecture through an mTOR/RNA Pol I axis.

Chromatin architecture is a fundamental mediator of genome function. Fasting is a major environmental cue across the animal kingdom, yet how it impacts three-dimensional (3D) genome organization is unknown. Here we show that fasting induces an intestine-specific, reversible and large-scale spatial reorganization of chromatin in Caenorhabditis elegans. This fasting-induced 3D genome reorganization requires inhibition of the nutrient-sensing mTOR pathway, acting through the regulation of RNA Pol I, but not Pol II nor Pol III, and is accompanied by remodelling of the nucleolus. By uncoupling the 3D genome configuration from the animal's nutritional status, we find that the expression of metabolic and stress-related genes increases when the spatial reorganization of chromatin occurs, showing that the 3D genome might support the transcriptional response in fasted animals. Our work documents a large-scale chromatin reorganization triggered by fasting and reveals that mTOR and RNA Pol I shape genome architecture in response to nutrients.

The emerging role of SARS-CoV-2 nonstructural protein 1 (nsp1) in epigenetic regulation of host gene expression.

Infection with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes widespread changes in epigenetic modifications and chromatin architecture in the host cell. Recent evidence suggests that SARS-CoV-2 nonstructural protein 1 (nsp1) plays an important role in driving these changes. Previously thought to be primarily involved in host translation shutoff and cellular mRNA degradation, nsp1 has now been shown to be a truly multifunctional protein that affects host gene expression at multiple levels. The functions of nsp1 are surprisingly diverse and include not only the downregulation of cellular mRNA translation and stability, but also the inhibition of mRNA export from the nucleus, the suppression of host immune signaling, and, most recently, the epigenetic regulation of host gene expression. In this review, we first summarize the current knowledge on SARS-CoV-2-induced changes in epigenetic modifications and chromatin structure. We then focus on the role of nsp1 in epigenetic reprogramming, with a particular emphasis on the silencing of immune-related genes. Finally, we discuss potential molecular mechanisms underlying the epigenetic functions of nsp1 based on evidence from SARS-CoV-2 interactome studies.

Innate Immune Response and Epigenetic Regulation: A Closely Intertwined Tale in Inflammation.

Maintenance of delicate homeostasis is very important in various diseases because it ensures appropriate immune surveillance against pathogens and prevents excessive inflammation. In a disturbed homeostatic condition, hyperactivation of immune cells takes place and interplay between these cells triggers a plethora of signaling pathways, releasing various pro-inflammatory cytokines such as Tumor necrosis factor alpha (TNFα), Interferon-gamma (IFNƴ), Interleukin-6 (IL-6), and Interleukin-1 beta (IL-1β), which marks cytokine storm formation. To be precise, dysregulated balance can impede or increase susceptibility to various pathogens. Pathogens have the ability to hijack the host immune system by interfering with the host's chromatin architecture for their survival and replication in the host cell. Cytokines, particularly IL-6, Interleukin-17 (IL-17), and Interleukin-23 (IL-23), play a key role in orchestrating innate immune responses and shaping adaptive immunity. Understanding the interplay between immune response and the role of epigenetic modification to maintain immune homeostasis and the structural aspects of IL-6, IL-17, and IL-23 can be illuminating for a novel therapeutic regimen to treat various infectious diseases. In this review, the light is shed on how the orchestration of epigenetic regulation facilitates immune homeostasis.

Three-dimensional chromatin mapping of sensory neurons reveals that promoter–enhancer looping is required for axonal regeneration

The in vivo three-dimensional genomic architecture of adult mature neurons at homeostasis and after medically relevant perturbations such as axonal injury remains elusive. Here, we address this knowledge gap by mapping the three-dimensional chromatin architecture and gene expression program at homeostasis and after sciatic nerve injury in wild-type and cohesin-deficient mouse sensory dorsal root ganglia neurons via combinatorial Hi-C, promoter-capture Hi-C, CUT&Tag for H3K27ac and RNA-seq. We find that genes involved in axonal regeneration form long-range, complex chromatin loops, and that cohesin is required for the full induction of the regenerative transcriptional program. Importantly, loss of cohesin results in disruption of chromatin architecture and severely impaired nerve regeneration. Complex enhancer-promoter loops are also enriched in the human fetal cortical plate, where the axonal growth potential is highest, and are lost in mature adult neurons. Together, these data provide an original three-dimensional chromatin map of adult sensory neurons in vivo and demonstrate a role for cohesin-dependent long-range promoter interactions in nerve regeneration.

Unraveling the key role of chromatin structure in cancer development through epigenetic landscape characterization of oral cancer

Epigenetic alterations, such as those in chromatin structure and DNA methylation, have been extensively studied in a number of tumor types. But oral cancer, particularly oral adenocarcinoma, has received far less attention. Here, we combined laser-capture microdissection and muti-omics mini-bulk sequencing to systematically characterize the epigenetic landscape of oral cancer, including chromatin architecture, DNA methylation, H3K27me3 modification, and gene expression. In carcinogenesis, tumor cells exhibit reorganized chromatin spatial structures, including compromised compartment structures and altered gene-gene interaction networks. Notably, some structural alterations are observed in phenotypically non-malignant paracancerous but not in normal cells. We developed transformer models to identify the cancer propensity of individual genome loci, thereby determining the carcinogenic status of each sample. Insights into cancer epigenetic landscapes provide evidence that chromatin reorganization is an important hallmark of oral cancer progression, which is also linked with genomic alterations and DNA methylation reprogramming. In particular, regions of frequent copy number alternations in cancer cells are associated with strong spatial insulation in both cancer and normal samples. Aberrant methylation reprogramming in oral squamous cell carcinomas is closely related to chromatin structure and H3K27me3 signals, which are further influenced by intrinsic sequence properties. Our findings indicate that structural changes are both significant and conserved in two distinct types of oral cancer, closely linked to transcriptomic alterations and cancer development. Notably, the structural changes remain markedly evident in oral adenocarcinoma despite the considerably lower incidence of genomic copy number alterations and lesser extent of methylation alterations compared to squamous cell carcinoma. We expect that the comprehensive analysis of epigenetic reprogramming of different types and subtypes of primary oral tumors can provide additional guidance to the design of novel detection and therapy for oral cancer.