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Results for “chromatin architecture”

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At least 19 recordsLinked to original sources

PubMed2006

Hypophosphorylation of the architectural chromatin protein DEK in death-receptor-induced apoptosis revealed by the isotope coded protein label proteomic platform.

During apoptosis nuclear morphology changes dramatically due to alterations of chromatin architecture and cleavage of structural nuclear proteins. To characterize early events in apoptotic nuclear dismantling we have performed a proteomic study of apoptotic nuclei. To this end we have combined a cell-free apoptosis system with a proteomic platform based on the differential isotopic labeling of primary amines with N-nicotinoyloxy-succinimide. We exploited the ability of this system to produce nuclei arrested at different stages of apoptosis to analyze proteome alterations which occur prior to or at a low level of caspase activation. We show that the majority of proteins affected at the onset of apoptosis are involved in chromatin architecture and RNA metabolism. Among them is DEK, an architectural chromatin protein which is linked to autoimmune disorders. The proteomic analysis points to the occurrence of multiple PTMs in early apoptotic nuclei. This is confirmed by showing that the level of phosphorylation of DEK is decreased following apoptosis induction. These results suggest the unexpected existence of an early crosstalk between cytoplasm and nucleus during apoptosis. They further establish a previously unrecognized link between DEK and cell death, which will prove useful in the elucidation of the physiological function of this protein.

Animals↗

PubMed2004

Common chromatin architecture, common chromatin remodeling, and common transcription kinetics of Adr1-dependent genes in Saccharomyces cerevisiae.

The chromatin structure of several Saccharomyces cerevisiae ADR1-dependent genes was comparatively analyzed in vivo in order to evaluate the role of promoter architecture in transcriptional control. In repressing conditions (high glucose) a nucleosome particle always obstructs the TATA box, immediately adjacent to an upstream-located nucleosome-free region containing a cluster of Adr1 binding sites. Upon derepression the TATA box-containing nucleosome is destabilized according to a mechanism shared by all of the genes studied. The transcription factor Adr1 is always required for the observed chromatin remodeling. mRNA accumulation of all of the genes analyzed is strongly delayed in the absence of the acetyltransferase Gcn5 and is decreased in the presence of a temperature-sensitive Esa1 mutant. The results suggest that a defined promoter chromatin structure, controlled by DNA conformational features, is relevant for the activation of coregulated genes.

Chromatin↗

PubMed2026

dcHiChIP: a comprehensive Nextflow-based pipeline for multiscale analysis of chromatin architecture from HiChIP data.

MOTIVATION: Despite the growing use of HiChIP to investigate protein-directed chromatin architecture, a comprehensive and reproducible pipeline for analysing these datasets-from raw reads to multiscale 3D genome features-remains lacking. Existing tools often focus on isolated components, such as loop calling or matrix generation, but fall short in integrating structural annotation, functional enrichment, and spatial modeling within a unified framework. To address this gap, we developed dcHiChIP, a modular, scalable Nextflow-based workflow that streamlines the analysis of HiChIP data, enabling both routine processing and in-depth exploration of chromatin organization and regulatory interactions. RESULTS: dcHiChIP enables robust and reproducible analysis of HiChIP datasets across multiple scales of chromatin architecture. It accepts raw sequencing data as input and generates high-quality loop calls, domain annotations, and 3D genome models. It also performs functional annotation and motif enrichment analyses. Applied to benchmark CTCF HiChIP datasets, dcHiChIP identifies major chromatin architectural features such as TADs/CCDs, A/B compartments, and chromatin stripes, and offers efficient, end-to-end execution with support for batch processing and workflow resumability. AVAILABILITY: dcHiChIP is publicly available on GitHub at https://github.com/SFGLab/dcHiChIP, with documentation at https://sfglab.github.io/dcHiChIP/. The software version used in this study is archived at Zenodo: https://doi.org/10.5281/zenodo.22030542.

Chromatin↗

PubMed2006

Chromatin architectural proteins.

The accessibility of eukaryotic DNA is dependent upon the hierarchical level of chromatin organization. These include (1) intra-nucleosome interactions, (2) inter-nucleosome interactions and (3) the influence of non-histone chromatin architectural proteins. There appears to be interplay between all these levels, in that one level can override another or that two or more can act in concert. In the first level, the stability of the nucleosome itself is dependent on the number and type of contacts between the core histones and the surrounding DNA, as well as protein-protein interactions within the core histone octamer. Core histone variants, post-translational modifications of the histones, and linker histones binding to the DNA all influence the organization and stability of the nucleosome. When nucleosomes are placed end-to-end in linear chromatin arrays, the second level of organization is revealed. The amino terminal tails of the histone proteins make contacts with adjacent and distant nucleosomes, both within the fiber and between different fibers. The third level of organization is imposed upon these 'intrinsic' constraints, and is due to the influence of chromatin binding proteins that alter the architecture of the underlying fiber. These chromatin architectural proteins can, in some cases, bypass intrinsic constraints and impart their own topological affects, resulting in truly unique, supra-molecular assemblages that undoubtedly influence the accessibility of the underlying DNA. In this review we will provide a brief summary of what has been learned about the intrinsic dynamics of chromatin fibers, and survey the biology and architectural affects of the handful of chromatin architectural proteins that have been identified and characterized. These proteins are likely only a small subset of the architectural proteins encoded within the eukaryotic genome. We hope that an increased understanding and appreciation of the contribution of these proteins to genome accessibility will hasten the identification and characterization of more of these important regulatory factors.

Animals↗

PubMed1989

Chromatin architecture and nuclear RNA.

The maintenance of normal chromatin morphology requires ongoing RNA synthesis. We have examined the role of RNA in chromatin organization, using selective detergent extraction of cells, RNA synthesis inhibitors, and enzymatic digestion of nuclear RNA. Comparison of extracted and unextracted cells showed that the important features of chromatin architecture were largely unchanged by the extraction procedure. Normally, chromatin was distributed in small heterochromatic regions and dispersed euchromatic strands. Ribonucleoprotein granules were dispersed throughout the euchromatic regions. Exposure to actinomycin led to the redistribution of chromatin into large clumps, leaving large empty spaces and a dense clustering of the remaining ribonucleoprotein granules. When the nuclei of extracted cells were digested with RNase A, there was a rearrangement of chromatin similar to but more pronounced than that seen in cells exposed to actinomycin. The inhibitor 5,6-dichloro-1-beta-D-ribofuranosylbenzimidizole also inhibits RNA synthesis but by a different mechanism that leaves no nascent RNA chains. The drug had little effect on chromatin after brief exposure but resembled actinomycin in its effect at longer times. We also examined the structure of the nuclear matrix to which most heteronuclear RNA remains associated. Pretreatment of cells with actinomycin or digestion of the nuclear matrix with RNase A caused the matrix fibers to collapse and aggregate. The experiments show a parallel decay of chromatin and of nuclear matrix organization with the depletion of nuclear RNA and suggest that RNA is a structural component of the nuclear matrix, which in turn may organize the higher order structure of chromatin.

Cell Nucleus↗

PubMed1975

Chromatin architecture: investigation of a subunit of chromatin by dark field electron microscopy.

Dark field scanning electron microscopy of unstained, unfixed samples of chromatin, histone-1-depleted chromatin, and nucleohistone has been used to identify an apparent subunit of chromatin, namely a disk-shaped structure we term the unit particle, which is probably about 135 A wide and 50 A thick in the hydrated state. The unit particles are found at rather uniform intervals along thin DNA-like fibers. Histone 1 depletion leads to a bimodal distribution of these spacings. Our observations suggest that the unit particle consists of a loop of nucleoprotein, perhaps around a histone core.

Animals↗

PubMed2006

The human IL-13 locus in neonatal CD4+ T cells is refractory to the acquisition of a repressive chromatin architecture.

The Th2 cytokine IL-13 is a major effector molecule in human allergic inflammation. Notably, IL-13 expression at birth correlates with subsequent susceptibility to atopic disease. In order to characterize the chromatin-based mechanisms that regulate IL-13 expression in human neonatal CD4(+) T cells, we analyzed patterns of DNase I hypersensitivity and epigenetic modifications within the IL-13 locus in cord blood CD4(+) T cells, naive or differentiated in vitro under Th1- or Th2-polarizing conditions. In naive CD4(+) T cells, hypersensitivity associated with DNA hypomethylation was limited to the distal promoter. Unexpectedly, during both Th1 and Th2 differentiation, the locus was extensively remodeled, as revealed by the formation of numerous HS sites and decreased DNA methylation. Obvious differences in chromatin architecture were limited to the proximal promoter, where strong hypersensitivity, hypomethylation, and permissive histone modifications were found selectively in Th2 cells. In addition to revealing the locations of putative cis-regulatory elements that may be required to control IL-13 expression in neonatal CD4(+) T cells, our results suggest that differential IL-13 expression may depend on the acquisition of a permissive chromatin architecture at the proximal promoter in Th2 cells rather than the formation of locus-wide repressive chromatin in Th1 cells.

CD4-Positive T-Lymphocytes↗

PubMed2003

A conserved chromatin architecture marks and maintains the restricted germ cell lineage in worms and flies.

In C. elegans, mRNA production is initially repressed in the embryonic germline by a protein unique to C. elegans germ cells, PIE-1. PIE-1 is degraded upon the birth of the germ cell precursors, Z2 and Z3. We have identified a chromatin-based mechanism that succeeds PIE-1 repression in these cells. A subset of nucleosomal histone modifications, methylated lysine 4 on histone H3 (H3meK4) and acetylated lysine 8 on histone H4 (H4acetylK8), are globally lost and the DNA appears more condensed. This coincides with PIE-1 degradation and requires that germline identity is not disrupted. Drosophila pole cell chromatin also lacks H3meK4, indicating that a unique chromatin architecture is a conserved feature of embryonic germ cells. Regulation of the germline-specific chromatin architecture requires functional nanos activity in both organisms. These results indicate that genome-wide repression via a nanos-regulated, germ cell-specific chromatin organization is a conserved feature of germline maintenance during embryogenesis.

Animals↗

PubMed2026

Allele-specific chromatin architecture shapes imprinted domains and coordinates a distal enhancer and antisense transcription at the mouse Mest-Copg2 domain.

Genomic imprinting results in parent-of-origin-dependent gene expression, but how three-dimensional genome organization contributes to imprinted gene regulation remains unclear. Using Capture Hi-C in mouse cortex and primary cortical neurons, we identified parental allele-specific chromatin architectures across multiple imprinted domains. These architectures largely originate from imprinting control regions and correlate with DNA methylation-sensitive CTCF binding. Active and inactive alleles of imprinted genes show distinct promoter interaction profiles and differential engagement with distal regulatory elements in both contact frequency and the epigenetic state of distal regions. A CRISPR interference screen identified a distal enhancer that regulates Mest-Copg2 imprinted expression through allele-specific chromatin interactions. In neurons, this enhancer activates Copg2 on the maternal allele, whereas on the paternal allele it drives Mest isoforms transcribed antisense to Copg2 and contributes to Copg2 repression. In summary, we show that allele-specific chromatin architecture coordinates maternal enhancer activity and paternal antisense transcription to control imprinted expression in neurons.

Animals↗

PubMed2006

Chromatin architecture.

A complete understanding of the structure-function relationships of chromatin requires extending primarily one dimensional information, obtained from molecular genetic techniques and based on the underlying linear DNA sequence, to the three dimensional conformation. Recent progress in this endeavor has included the examination of fully defined nucleosomes and nucleosomal arrays assembled in vitro using X-ray diffraction, NMR spectroscopy, electron microscopy and atomic force microscopy. These studies have provided valuable insights into the structural roles of histone variants, the impact of histone mutations and the compaction of nucleosomal arrays. In addition, the diverse structural consequences of the binding of specific chromatin 'architectural' proteins are becoming apparent. These approaches provide an essential basis for understanding the conformation of the 'epigenome'.

Animals↗

PubMed1996

Chromatin architectural proteins and transcription factors: a structural connection.

It has long been assumed that the architectural proteins of chromatin (the histones, for example) are unrelated to their functional proteins (transcription factors, polymerases, etc). New studies drastically change this perspective. It appears that a portion of the general transcription initiation complex TFIID is made up of proteins that not only carry marked sequence and structural resemblances to the core histones of the nucleosome, but also form an octameric complex similar to the histone octamer. This can now be seen as part of a general pattern of continuity among structural and functional chromatin proteins.

Animals↗

PubMed2006

Changes in intranuclear chromatin architecture induce bipolar nuclear localization of histone variant H1T2 in male haploid spermatids.

Spermiogenesis entails a major biochemical and morphological restructuring of the germ cell packing the DNA into the condensed spermatid nucleus. H1T2 is a histone H1 variant selectively and transiently expressed in male haploid germ cells during spermiogenesis that specifically localizes to a chromatin domain at the apical pole under the acrosome. We explored the mechanisms determining polar localization of H1T2 in spermatids. In acrosome-deficient round spermatids of hrb -/- and gopc -/- mice, H1T2 localization is not altered, indicating that proper acrosome development is not required for specifying nuclear polarity. In contrast, in late round spermatids from trf2 -/- or hmgb2 -/- mice, a bipolar H1T2 localization was observed revealing that polarity is modified by loss of proteins specifying chromatin architecture. Our results show that intranuclear chromatin organization is critical for correct polar localization of H1T2 and that H1T2 can be a useful molecular marker revealing chromatin disorganization in spermatids.

Active Transport, Cell Nucleus↗

PubMed2002

Spatial preservation of nuclear chromatin architecture during three-dimensional fluorescence in situ hybridization (3D-FISH).

3D-FISH has become a major tool for studying the higher order chromatin organization in the cell nucleus. It is not clear, however, to what extent chromatin arrangement in the nucleus after fixation and 3D-FISH still reflects the order in living cells. To study this question, we compared higher order chromatin arrangements in living cells with those found after the 3D-FISH procedure. For in vivo studies we employed replication labeling of DNA with Cy3-conjugated nucleotides and/or chromatin labeling by GFP-tagged histone 2B. At the light microscope level, we compared the intranuclear distribution of H2B-GFP-tagged chromatin and the positions of replication-labeled chromatin domains in the same individual cells in vivo, after fixation with 4% paraformaldehyde, and after 3D-FISH. Light microscope data demonstrate a high degree of preservation of the spatial arrangement of approximately 1-Mb chromatin domains. Subsequent electron microscope investigations of chromatin structure showed strong alterations in the ultrastructure of the nucleus caused mainly by the heat denaturation step. Through this step chromatin acquires the appearance of a net with mesh size of 50-200 nm roughly corresponding to the average displacement of the chromatin domains observed at light microscope level. We conclude that 3D-FISH is a useful tool to study chromosome territory structure and arrangements down to the level of approximately 1-Mb chromatin domain positions. However, important ultrastructural details of the chromatin architecture are destroyed by the heat denaturation step, thus putting a limit to the usefulness of 3D-FISH analyses at nanometer scales.

Carbocyanines↗

PubMed2005

Synergistic and additive properties of the beta-globin locus control region (LCR) revealed by 5'HS3 deletion mutations: implication for LCR chromatin architecture.

Deletion of the 234-bp core element of the DNase I hypersensitive site 3 (5'HS3) of the locus control region (LCR) in the context of a human beta-globin locus yeast artificial chromosome (beta-YAC) results in profound effects on globin gene expression in transgenic mice. In contrast, deletion of a 2.3-kb 5'HS3 region, which includes the 234-bp core sequence, has a much milder phenotype. Here we report the effects of these deletions on chromatin structure in the beta-globin locus of adult erythroblasts. The 234-bp 5'HS3 deletion abolished histone acetylation throughout the beta-globin locus; recruitment of RNA polymerase II (pol II) to the LCR and beta-globin gene promoter was reduced to a basal level; and formation of all the 5' DNase I hypersensitive sites of the LCR was disrupted. The 2.3-kb 5'HS3 deletion mildly reduced the level of histone acetylation but did not change the profile across the whole locus; the 5' DNase I hypersensitive sites of the LCR were formed, but to a lesser extent; and recruitment of pol II was reduced, but only marginally. These data support the hypothesis that the LCR forms a specific chromatin structure and acts as a single entity. Based on these results we elaborate on a model of LCR chromatin architecture which accommodates the distinct phenotypes of the 5'HS3 and HS3 core deletions.

Acetylation↗

PubMed2001

Methylation-mediated silencing of TMS1/ASC is accompanied by histone hypoacetylation and CpG island-localized changes in chromatin architecture.

Aberrant methylation of CpG-dense islands in the promoter regions of genes is an acquired epigenetic alteration associated with the silencing of tumor suppressor genes in human cancers. In a screen for endogenous targets of methylation-mediated gene silencing, we identified a novel CpG island-associated gene, TMS1, which is aberrantly methylated and silenced in response to the ectopic expression of DNA methyltransferase-1. TMS1 functions in the regulation of apoptosis and is frequently methylated and silenced in human breast cancers. In this study, we characterized the methylation pattern and chromatin architecture of the TMS1 locus in normal fibroblasts and determined the changes associated with its progressive methylation. In normal fibroblasts expressing TMS1, the CpG island is defined by an unmethylated domain that is separated from densely methylated flanking DNA by distinct 5' and 3' boundaries. Analysis of the nucleoprotein architecture of the locus in intact nuclei revealed three DNase I-hypersensitive sites that map within the CpG island. Strikingly, two of these sites coincided with the 5'- and 3'-methylation boundaries. Methylation of the TMS1 CpG island was accompanied by loss of hypersensitive site formation, hypoacetylation of histones H3 and H4, and gene silencing. This altered chromatin structure was confined to the CpG island and occurred without significant changes in methylation, histone acetylation, or hypersensitive site formation at a fourth DNase I-hypersensitive site 2 kb downstream of the TMS1 CpG island. The data indicate that there are sites of protein binding and/or structural transitions that define the boundaries of the unmethylated CpG island in normal cells and that aberrant methylation overcomes these boundaries to direct a local change in chromatin structure, resulting in gene silencing.

Acetylation↗

PubMed2001

Methyl CpG binding proteins: coupling chromatin architecture to gene regulation.

A correlation between DNA methylation and transcriptional silencing has existed for many years. Recently, substantial progress has been reported in the search for proteins that interpret the regulatory information inherent in DNA methylation and translate this information into functional states, resulting in the identification of a family of highly conserved proteins, the MBD family. Direct connections between these proteins and histone modification enzymes have emerged as a common theme, implying that DNA methylation exerts its effects primarily through repressive chromatin architecture. Recent structural determinations of the DNA binding domain of two MBD family members, MeCP2 and MBD1, provide a framework to model the interactions of this family with DNA. Comparative sequence analysis and experimental DNA binding data can be interpreted using this structural framework allowing one to contrast the members of the MBD family with each other and to predict the properties of new family members. The identification of mutations in MeCP2, the founding member of this family, as causal for the neurological developmental disorder Rett Syndrome provides additional information regarding amino acid residues crucial to the functions of this interesting protein family.

Amino Acid Sequence↗

PubMed2004

Nascent RNA synthesis in the context of chromatin architecture.

Based on the idea that chromatin domains provide physical barriers for large molecules and multi-enzyme complexes, including the components of the transcription machinery, it has been proposed that transcription should be confined to the surfaces of chromatin domains. As a consequence nascent RNA should accumulate in the interchromatin space, which is thought to provide a special nuclear compartment involved in transcription, as well as in the processing and export of RNA (Cremer et al. 1993, Cremer & Cremer 2001). To further address the relationships between chromatin organization and RNA synthesis, we investigated the localization of BrUTP-labelled nascent RNA in HeLa cells stably expressing green fluorescent protein (GFP)-tagged histone H2B, which highlights the chromatin structure. Our results showed that nascent RNA does not preferentially localize within the interchromatin space. The findings do not support the idea that the interchromatin space provides a nuclear compartment playing an essential role in nascent RNA synthesis. However, the results are in agreement with the emerging view that even condensed chromatin domains display a highly dynamic organization and are not a physical barrier for transcription factors.

Chromatin↗

PubMed2001

Genetic lesions and perturbation of chromatin architecture: a road to cell transformation.

Differential gene expression is a rigorously precise procedure that defines the developmental program of cells, tissues, organs, and of the entire organism. The correct execution of this program requires the participation of multiple and complex groups of regulators. In addition to transcription factors, which are key tools in ontogenesis by providing sequential switch of different genes, the structure of the chromatin is a dominant determinant leading to gene expression. Through the novel and insightful work of several investigators, it appears that the architecture of the chromatin spanning the genes can and does influence the efficiency of RNA transcription, and therefore of gene expression. Several new enzymatic complexes have been identified that reversibly modify the chromatin architecture by methylation, phosphorylation, and acetylation of the nucleosomal core proteins. These enzymes are crucial for the proper balance and maintenance of gene expression, and are often the target of mutations and alterations in human cancer. Here, we review briefly the current models proposing how some of these enzymes normally modify the chromatin structure and how their functional disruption leads to inappropriate gene expression and cell transformation.

Acetylation↗