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At least 19 recordsLinked to original sources
PubMed1976

Chromatin organization in nuclei of sea urchin embryos. Comparison with the chromatin organization of the sperm.

The chromatin in sea urchin embryo nuclei and that in sperm heads are both organized in nucleosomes but show marked differences when analyzed by endonuclease digestion. Sperm chromatin DNA appears to be totally organized in nucleosomes that are highly resistant to nuclease hydrolysis. The kinetics of formation of acid-soluble oligonucleotides is slow and concerns only about 50% of the total DNA. In contrast, the DNA of embryo chromatin does not appear to be totally organized in nucleosomes since 5 to 10% is rapidly and preferentially hydrolysed into acid-soluble oligonucleotides without any appreciable fragmentation of the remaining parts. Futher digestion causes the formation of the usual pattern of DNA bands, as detected by gel electrophoresis. The length of the DNA segment associated with the embryo nucleosomes appears to be shorter than that of the DNA segment associated with the sperm nucleosomes. The kinetics of formation of acid-soluble oligonucleotides upon digestion of embryo chromatin is much faster than that of sperm chromatin and concerns almost all the chromatin DNA.

Animals↗
PubMed2025

ChromaFactor: Deconvolution of single-molecule chromatin organization with non-negative matrix factorization.

The investigation of chromatin organization in single cells holds great promise for identifying causal relationships between genome structure and function. However, analysis of single-molecule data is hampered by extreme yet inherent heterogeneity, making it challenging to determine the contributions of individual chromatin fibers to bulk trends. To address this challenge, we propose ChromaFactor, a novel computational approach based on non-negative matrix factorization that deconvolves single-molecule chromatin organization datasets into their most salient primary components. ChromaFactor provides the ability to identify trends accounting for the maximum variance in the dataset while simultaneously describing the contribution of individual molecules to each component. Applying our approach to two single-molecule imaging datasets across different genomic scales, we find that these primary components demonstrate significant correlation with key functional phenotypes, including active transcription, enhancer-promoter distance, and genomic compartment. Also, we find that some bulk trends exist at the single-cell level, but only in a small fraction of cells, suggesting that critical changes in genome organization may be driven by specific rare subpopulations rather than occurring uniformly across all cells. ChromaFactor offers a robust tool for understanding the complex interplay between chromatin structure and function on individual DNA molecules, pinpointing which subpopulations drive functional changes and fostering new insights into cellular heterogeneity and its implications for bulk genomic phenomena.

Animals↗
PubMed1978

Structure of interphase nuclei in relation to the cell cycle. Chromatin organization in mouse L cells temperature-sensitive for DNA replication.

Mutant lines of mouse L cells, TS A1S9, and TS C1, show temperature-sensitive (TS) DNA synthesis and cell division when shifted from 34 degrees to 38.5 degrees C. With TS A1S9 the decline in DNA synthesis begins after 6-8 h at 38.5 degrees C and is most marked at about 24 h. Most cells in S, G2, or M at temperature upshift complete one mitosis and accumulate in the subsequent interphase at G1 or early S as a result of expression of a primary defect, failure of elongation of newly made small DNA fragments. Heat inactivation of TS C1 cells is more rapid; they fail to complete the interphase in progress at temperature upshift and accumulate at late S or G2. Inhibition of both cell types is reversible on return to 34 degrees C. Cell and nuclear growth continues during inhibition of replication. Expression of both TS mutations leads to a marked change in gross organization of chromatin as revealed by electron microscopy. Nuclei of wild-type cells at 34 degrees and 38.5 degrees C and mutant cells at 34 degrees C show a range of aggregation of condensed chromatin from small dispersed bodies to large discrete clumps, with the majority in an intermediate state. In TS cells at 38.5 degrees C, condensed chromatin bodies in the central nuclear region become disaggregated into small clumps dispersed through the nucleus. Morphometric estimation of volume of condensed chromatin indicates that this process is not due to complete decondensation of chromatin fibrils, but rather involves dispersal of large condensed chromatin bodies into finer aggregates and loosening of fibrils within the aggregates. The dispersed condition is reversed in nuclei which resume DNA synthesis when TS cells are downshifted from 38.5 degrees to 34 degrees C. The morphological observations are consistent with the hypothesis that condensed chromatin normally undergoes an ordered cycle of transient, localized disaggregation and reaggregation associated with replication. In temperature-inactivated mutants, normal progressive disaggregation presumably occurs, but subsequent lack of chromatin replication prevents reaggregation.

Cell Count↗
PubMed1978

Action of heparin on mammalian nuclei. II. Cell-cycle-specific changes in chromatin organization correlate temporally with histone H1 phosphorylation.

The interaction of the polyanion heparin with the inner histones of chromatin has been used to detect changes in chromatin organization associated with cell-cycle traverse. Synchronized populations of Chinese hamster cells were obtained either in early G1 or near the G1/S boundary. The rate of interaction of heparin with chromatin-associated inner histones was measured using nuclei isolated from synchronized cell populations in different phases of the cell cycle. A G1-specific decrease in rate of interaction of heparin with inner histones was observed and found to be independent of the presence of hydroxyurea during traverse of G1. A further decrease in heparin-inner histone interaction occurred in late S and G2. These changes correlate temporally with the interphase phosphorylation(s) of histone H1. This correlation is discussed within the framework of current models of higher order chromatin structure (i.e. organization above the nucleosome level). Analysis of the cooperativity of interaction of heparin with inner histones was performed using the kinetic analog of the Hill equation. This analysis suggests that the organization of inner histones on chromatin does not undergo large variations during the cell cycle.

Cell Cycle↗
PubMed2025

RNA Pol I activity is required for meiotic chromatin organization and the H3K4me3 gradient essential for oogenesis, independent of ribosome synthesis.

Oogenesis requires extensive and dynamic chromatin remodeling that primes gene promoters for later transcriptional activation during embryonic development. Here, we uncover a pivotal, non-canonical role for RNA Polymerase I (Pol I) in driving these chromatin state transitions during Caenorhabditis elegans oogenesis. Using the auxin-inducible degron system to selectively deplete either Pol I catalytic subunits or ribosome assembly factors, we disentangle the consequences of impaired nucleolar integrity from reductions in ribosome biogenesis. Strikingly, although disrupting ribosome assembly caused minimal effects on oocyte production, loss of Pol I activity led to widespread changes in chromatin accessibility, a dampening of the distal-proximal H3K4me3 gradient required for oogenesis, reduced synapsis, and elevated ATM/ATR phosphorylation, resulting in fewer but significantly larger oocytes. Despite their promoters becoming more accessible, oogenesis genes did not show large changes in steady-state mRNA, consistent with transcriptional repression prior to fertilization. Instead, Pol I depletion prematurely remodeled oogenic chromatin, through a misdirection of H3K4me3 deposition towards promoters normally primed for zygotic genome activation. These findings reveal an epigenetic gating function for nucleolar integrity in oocyte maturation: Pol I preserves three-dimensional chromatin organization and maintains proper spatiotemporal regulation of histone modifications, independent of ribosome production. Given the evolutionary conservation of nucleolar dynamics and histone modifications during gametogenesis, our work suggests that nucleolar stress, whether from environmental factors, aging, or genetic disorders, could broadly compromise fertility by disrupting oogenic chromatin priming.

Journal Article↗
PubMed1979

Influence of diet on chromatin organization in insect mid-gut cells, and its implications for the understanding of aging in this model system.

Female Sarcophaga bullata were fed on three different dietary combinations: liver/water; sugar/water; and sugar/liver/water. These diets resulted in different survival patterns for the insect, with the longest lifespans being expressed on the last treatment. The Feulgen-DNA values for mid-gut cell nuclei were determined using a scanning and integrating microdensitometer. The cell nuclei of the mid-gut reacted to the presence of nutrients by an increase in the extinction values for Feulgen-DNA when compared to the unfed state. There were also significant differences between the various treatments in the extent of the activation shown by the mid-gut cells. The dietary regimes resulting in the longest lifespan showed a complete and uniform activation of the mid-gut epithelium, whereas other treatments did not. These observations suggest that changes in the chromatin organization occur with diet, and that maximum Feulgen-DNA extinctions occur (i.e. chromatin uncoiled and supporting transcription) on the sugar/liver/water diet. Nuclei showing maximum extinctions do not show an age-related change between 3 and 12 days of life. However, if the nuclear activation is partial, or incomplete, then age-related reversals in chromatin condensation take place.

Aging↗
PubMed1992

Neutral filter elution detects differences in chromatin organization which can influence cellular radiosensitivity.

We have shown previously that the neutral filter elution assay is dependent not only on the number of DNA double-strand breaks present in a mammalian cell but also on the way in which DNA expands on the filter following lysis. Results in this study indicate that the rate of DNA elution appears to be dependent upon the proximity of the DNA in relation to the replication complex. The rate of elution for DNA analyzed immediately after a 30-min labeling period with [14C]thymidine was about five times slower than the rate of elution for bulk-labeled DNA. However, the rate was increased a few hours later when the recently replicated DNA had matured and was likely to be farther from replication-associated attachment sites on the nuclear protein matrix. About one cell cycle after pulse labeling, when the labeled DNA was replicated again, DNA underwent similar changes in elution rate. For the four cell lines examined here, the elution rate 3-4 h after pulse labeling correlated with cellular radiosensitivity. Changes in rate of elution caused by altering EDTA concentration or pH may also be explained by DNA structural changes which occur during lysis. We conclude that the neutral filter elution method is sensitive to differences in chromatin organization which may also play a role in cell sensitivity to ionizing radiation.

Animals↗
PubMed1979

Chromatin organization in the oomycete Achlya ambisexualis.

Nuclei from the Oömycete Achlya ambisexualis and rabbit kidney nuclei were digested with micrococcal nuclease and the resultant DNA fragments analyzed on slab gels. The average DNA repeat size was found to be 159 +/- 1.2 base pairs for Achlya and 199.8 +/- 3.7 base pairs for rabbit kidney. The presence of a DNA repeat size of 159 base pairs for Achlya extends the characterization of eukaryotic chromatins to this most primitive and perhaps unique microbe.

Biological Evolution↗
PubMed2025

Capturing Chromatin Organization by MNase-seq and ATAC-seq.

Hox genes play a pivotal role during development. Their expression is tightly controlled in a spatiotemporal manner, ensuring that specific body structures develop at the correct locations and times during development. Various genomics approaches have been used to capture temporal and dynamic regulation of Hox gene expression at the nucleosome/chromatin level. This chapter focuses on the utilization of capture MNase-seq and Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq), two advanced techniques that enable the exploration of chromatin accessibility and nucleosome positioning within these critical genomic regions.

Chromatin↗
PubMed2025

Pre-established ATF4 occupancy and chromatin organization instruct selective transcription activation during integrated stress response.

Cells rapidly and extensively remodel their transcriptome in response to stress to restore homeostasis, but the underlying mechanisms are not fully understood. Here, we characterize the dynamic changes in transcriptome, epigenetics, and 3D genome organization during the integrated stress response (ISR). ISR induction triggers widespread transcriptional changes within 6 h, coinciding with increased binding of ATF4, a key transcriptional effector. Notably, ATF4 binds to hundreds of genes even under non-stress conditions, priming them for stronger activation upon stress. The transcriptional changes at ATF4-bound sites during ISR do not rely on increased H3K27 acetylation, chromatin accessibility, or rewired enhancer-promoter looping. Instead, ATF4-mediated gene activation is linked to the redistribution of CEBPγ from non-ATF4 sites to a subset of ATF4-bound regions, likely by forming an ATF4/CEBPγ heterodimer. CEBPγ preferentially targets the sites pre-occupied by ATF4, as well as genomic regions exhibiting a unique higher-order chromatin structure signature. Thus, the transcriptional responses during ISR are largely pre-wired by intrinsic chromatin properties. These findings provide critical insights into transcriptional remodeling during ISR with broader implications for other stress responses.

Activating Transcription Factor 4↗
PubMed1992

Cytoplasmic microtubular dynamics and chromatin organization during mammalian oogenesis and oocyte maturation.

A chronological series of coordinated alterations in oocyte chromosome and microtubule disposition occur during oogenesis and oocyte maturation in the mammal. Timely transitions in meiotic spindle and cytoplasmic microtubules, due to modifications in both the assembly competence of the tubulin pool and nucleation capacity of centrosomes, underscore key nuclear events during the progressive stages of meiosis I and II. The regulation of these transitional states during meiosis is discussed with respect to hormonal influences imparted to the oocyte within the follicular microenvironment, and the possible ways in which environmental perturbations may result in defective chromosomal partitioning during meiosis.

Animals↗
PubMed2026

Foundation model reveals the shared organization of transcription and topologically associating domains.

The three-dimensional organization of chromatin into topologically associating domains (TADs) may impact gene regulation by bringing distant genes into contact. However, studies of TADs' function and their influence on transcription have been constrained by ambiguities in TAD boundary definitions and challenges in directly measuring their regulatory effects. We overcome these limitations by developing species-level consensus TAD maps for human and mouse by using a bag-of-genes approach that exposes an emergent regulatory structure. To quantify TAD-mediated relationships, we use a foundation model trained on 33 million transcriptomes to define a contextual similarity metric that captures higher-order relationships missed by co-expression. We find that TADs are regions of elevated co-regulation, with our framework yielding testable hypotheses about chromatin organization across cellular states. This TAD-linked enhancement is strongest during early development and declines with aging, while cancer cells show distinct TAD usage that shifts with chemotherapy. Together, these findings suggest that chromatin organization acts through probabilistic rather than deterministic mechanisms.

Humans↗
PubMed1992

[The possible role of endogenous nucleases in the structural organization of chromatin].

Two fractions of rat liver nuclei with different buoyant density have been obtained. The electrophoretic analysis of the oligonucleosome patterns of DNA out of nuclei of these two fractions revealed different levels of activity in endonucleases. In case of inhibition during the extraction of activity in Ca, Mg-dependent endonucleases, the average size of high polymeric DNA is larger for nuclei with bigger buoyant density (fraction I) than for nuclei with smaller ones (fraction II). This finding is evidence of in situ existence of two pools of liver nuclei with different endogenic nuclease activities. In nuclear chromatin fraction I DNA is torsionally stressed; in fraction II it is relaxed that correlates with larger activity of endonucleases and smaller buoyant density of this fraction. A hypothesis on a possible role of endonucleases in chromatin structure organization has been put forward. According to this hypothesis a modulation of activity in nuclear endonucleases can determine different packaging and activity of chromatin from different pools of cellular nuclei.

Animals↗
PubMed2025

Engineered histones reshape chromatin in human cells.

Histone proteins and their variants have been found to play crucial and specialized roles in chromatin organization and the regulation of downstream gene expression; however, the relationship between histone sequence and its effect on chromatin organization remains poorly understood, limiting our functional understanding of sequence variation between distinct subtypes and across evolution and frustrating efforts to rationally design synthetic histones that can be used to engineer specified cell states. Here, we make the first advance towards engineered histone-driven chromatin organization. By expressing libraries of sequence variants of core histones in human cells, we identify variants that dominantly modulate chromatin structure. We further interrogate variants using a combination of imaging, proteomics, and genomics to reveal both cis and trans-acting mechanisms of effect. Functional screening with transcription factor libraries identifies transcriptional programs that are facilitated by engineered histone expression. Double mutation screens combined with protein language models allow us to learn sequence-to-function patterns and design synthetic histone proteins optimized to drive specific chromatin states. This work establishes a foundation for the high-throughput evaluation and engineering of chromatin-associated proteins and positions histones as tunable nodes for understanding and modulating mesoscale chromatin organization.

Journal Article↗
PubMed1979

The structural organization of mouse chromatin as a function of age.

Chromatin is organized into a repeating structure (nucleosome) made up of proteins and DNA. Micrococcal nuclease and DNAase I have been used to probe this structure in nuclear populations from three tissues (liver, brain, and heart) of the inbred mouse strain C57BL at different ages. For those parameters examined, for each tissue, chromatin contained essentially the same features of nucleosomal organization, regardless of the age of the mouse. Thus, the rate and extent of nuclease digestion and the size of the DNA repeat unit and nucleosome core are not significantly different as a function of age. However, the accessibility of internucleosomal DNA to micrococcal nuclease, as determined by measuring the DNA size distribution after nuclease cutting, may be partially limited in chromatin of brain (but not liver or heart) of older animals. These results indicate that there are no gross, age-related changes in the conformational state or organization of chromatin in these tissues. The results do not exclude smaller alterations in chromatin that might occur with age, which the current methodology might not be sensitive enough to detect.

Aging↗
PubMed1978

[Analysis of brain chromatin subunit organization].

Autodigestion of chromosomal DNA does not take place during the brain nuclei incubation in the presence of Ca2+ and Mg2+. The kinetic of chromatin digestion in brain and liver nuclei by staphylococcal nuclease and the formation of DNP-fragments suggest that subnucleosomes are generated in both cases by digesting of monomer specific sites. This monomer contains 185--200 DNA base pairs and the most starting DNA going throughout it. However the quantity of nuclease-resistant DNA in brain chromatin is more and the rate of subnucleosome formation is less than in liver chromatin. Redigestion of isolated monomers of brain chromatin results in the appearance of subnucleosomes similar to those which are formed under limited digestion of nuclear chromatin. The incubation of brain nuclei in the presence of Ca, Mg-dependent endonuclease prepared from liver nuclei results in the appearance of fragment. DNA-spectra of these fragments are similar to those prepared under digestion of liver chromatin in situ. These data suggest definite resemblance of subunit organization in brain and liver chromatin.

Animals↗