Morphological, Gene, and Hormonal Changes in Gonads and In-Creased Micrococcal Nuclease Accessibility of Sperm Chromatin Induced by Mercury.
Ontology highlight
ABSTRACT:
SUBMITTER: Lettieri G
PROVIDER: S-EPMC8773939 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Nucleic prepared from mouse submandibular salivary gland show marked fragility during isolation. Hwever, intact nuclei relatively free from cytoplasmic contamination were obtained by homogenization in buffers containing 0.88 M-sucrose, Ca2+, spermine, spermidine and the proteinase inhibitor aprotinin, followed by centrifugation through 2.2 M-sucrose. The kinetics of digestion by the micrococcal nuclease of chromatin in these nuclei are similar to those of chromatin from mouse liver nuclei. Base-pair size analysis of the solubilized DNA from both organs shows a stable high-molecular weight species of chromatin, which is further digested to mononucleosome and subnucleosome species. With extensive digestion the chromatin becomes insoluble. The mononucleosomes produced from salivary-gland chromatin after the inhibition of endogenous proteinase activity exhibit an s20,w value of 11S and contain histones H1, H2A, H2B, H3 and H4.
| S-EPMC1161800 | biostudies-other
Project description:Histone post-translational modifications (PTMs) such as acetylation and methylation are known to affect chromatin higher order structures. Primary targets of these modifications include basic residues present at N-terminus tail region of core histones. Four histone acetyltransferase (HAT) genes have been identified in trypanosomatids. HAT1, HAT3 and HAT4 of Leishmania donovani have been partially characterized. However, there is no report about HAT2 of Leishmania donovani. Lysine residues present on the N-terminal tail of Leishmania donovani histone H4 are conserved in other trypanosomatids and humans. PTMs of lysines modulate various functions at chromatin level. The four histone acetyltransferases encoded in Leishmania genome were over-expressed to analyse their functional activity. All four HATs were found actively acetylating core histones H3/H4. Similar to L. donovani HAT3 and HAT4, HAT2 was found to be a member of MYST family protein and have SAS2 type domain. Over-expression of HAT2 significantly increases acetylation of H4K4. To analyse the effect of HAT2 over-expression on chromatin accessibility, micrococcal nuclease digestion assay was performed. MNase digestion resulted in a higher proportion of the mononucleosomes and dinucleosomes in HAT2 over-expressing cells as compared to WT L. donovani cells. Acetylation of lysine-4 neutralizes the amino terminal region of histone H4. This weakens its interaction with neighbouring nucleosomes and the linker DNA. HAT2 over-expression in L. donovani resulted in highly accessible chromatin suggesting chromatin decondensation. HAT2 may have an important role to play in global regulation of transcription in L. donovani. Better understanding of these epigenetic determinants of parasite would help in designing novel therapeutic strategies.
| S-EPMC5423686 | biostudies-literature
Project description:The nuclear lamina has been shown to interact with the genome through lamina-associated domains (LADs). LADs have been identified by DamID, a proximity labeling assay, and more recently by chromatin immunoprecipitation-sequencing (ChIP-seq) of A- and B-type lamins. LADs form megabase-size domains at the nuclear periphery, they are gene-poor and mostly heterochromatic. Here, we show that the mode of chromatin fragmentation for ChIP, namely bath sonication or digestion with micrococcal nuclease (MNase), leads to the discovery of common but also distinct sets of lamin-interacting domains, or LiDs. Using ChIP-seq, we show the existence of lamin A/C (LMNA) LiDs with distinct gene contents, histone composition enrichment and relationships to lamin B1-interacting domains. The extent of genome coverage of lamin A/C (LMNA) LiDs in sonicated or MNase-digested chromatin is similar (∼730 megabases); however over half of these domains are uniquely detected in sonicated or MNase-digested chromatin. Sonication-specific LMNA LiDs are gene-poor and devoid of a broad panel of histone modifications, while MNase-specific LMNA LiDs are of higher gene density and are enriched in H3K9me3, H3K27me3 and in histone variant H2A.Z. LMNB1 LiDs are gene-poor and show no or little enrichment in these marks. Comparison of published LMNB1 DamID LADs with LMNB1 and LMNA LiDs identified here by ChIP-seq further shows that LMNA can associate with 'open' chromatin domains displaying euchromatin characteristics, and which are not associated with LMNB1. The differential genomic and epigenetic properties of lamin-interacting domains reflect the existence of distinct LiD populations identifiable in different chromatin contexts, including nuclease-accessible regions presumably localized in the nuclear interior.
| S-EPMC4615303 | biostudies-literature
Project description:The three-dimensional organization of the enormously long DNA molecules packaged within metaphase chromosomes has been one of the most elusive problems in structural biology. Chromosomal DNA is associated with histones and different structural models consider that the resulting long chromatin fibers are folded forming loops or more irregular three-dimensional networks. Here, we report that fragments of chromatin fibers obtained from human metaphase chromosomes digested with micrococcal nuclease associate spontaneously forming multilaminar platelike structures. These self-assembled structures are identical to the thin plates found previously in partially denatured chromosomes. Under metaphase ionic conditions, the fragments that are initially folded forming the typical 30-nm chromatin fibers are untwisted and incorporated into growing plates. Large plates can be self-assembled from very short chromatin fragments, indicating that metaphase chromatin has a high tendency to generate plates even when there are many discontinuities in the DNA chain. Self-assembly at 37°C favors the formation of thick plates having many layers. All these results demonstrate conclusively that metaphase chromatin has the intrinsic capacity to self-organize as a multilayered planar structure. A chromosome structure consistent of many stacked layers of planar chromatin avoids random entanglement of DNA, and gives compactness and a high physical consistency to chromatids.
| S-EPMC3414884 | biostudies-other
Project description:We have mapped sequence-directed nucleosome positioning on genomic DNA molecules using high-throughput sequencing. Chromatins, prepared by reconstitution with either chicken or frog histones, were separately digested to mononucleosomes using either micrococcal nuclease (MNase) or caspase-activated DNase (CAD). Both enzymes preferentially cleave internucleosomal (linker) DNA, although they do so by markedly different mechanisms. MNase has hitherto been very widely used to map nucleosomes, although concerns have been raised over its potential to introduce bias. Having identified the locations and quantified the strength of both the chicken or frog histone octamer binding sites on each DNA, the results obtained with the two enzymes were compared using a variety of criteria. Both enzymes displayed sequence specificity in their preferred cleavage sites, although the nature of this selectivity was distinct for the two enzymes. In addition, nucleosomes produced by CAD nuclease are 8-10 bp longer than those produced with MNase, with the CAD cleavage sites tending to be 4-5 bp further out from the nucleosomal dyad than the corresponding MNase cleavage sites. Despite these notable differences in cleavage behaviour, the two nucleases identified essentially equivalent patterns of nucleosome positioning sites on each of the DNAs tested, an observation that was independent of the histone type. These results indicate that biases in nucleosome positioning data collected using MNase are, under our conditions, not significant.
| S-EPMC3314939 | biostudies-literature
Project description:The increasing global burden of metabolic disorders including obesity and diabetes necessitates a comprehensive understanding of their etiology, which not only encompasses genetic and environmental factors but also parental influence. Recent evidence has unveiled paternal obesity as a contributing factor to offspring's metabolic health via sperm epigenetic modifications. In this study, we investigated the impact of a Western diet-induced obesity in C57BL/6 male mice on sperm chromatin accessibility and the subsequent metabolic health of their progeny. Utilizing Assay for Transposase-Accessible Chromatin with sequencing, we discovered 450 regions with differential accessibility in sperm from obese fathers, implicating key developmental and metabolic pathways. Contrary to expectations, these epigenetic alterations in sperm were not predictive of long-term metabolic disorders in offspring, who exhibited only mild transient metabolic changes early in life. Both male and female F1 progeny showed no enduring predisposition to obesity or diabetes. These results underscore the biological resilience of offspring to paternal epigenetic inheritance, suggesting a complex interplay between inherited epigenetic modifications and the offspring's own developmental compensatory mechanisms. This study calls for further research into the biological processes that confer this resilience, which could inform interventional strategies to combat the heritability of metabolic diseases.
| S-EPMC11296027 | biostudies-literature
Project description:Nuclear remodeling to a condensed state is a hallmark of spermatogenesis. This is achieved by replacement of histones with protamines. Regions retaining nucleosomes may be of functional significance. To determine their potential roles, sperm from wild type and transgenic mice harboring a single copy insert of the human protamine cluster were subjected to Micrococcal Nuclease-seq. CENTIPEDE, a hierarchical Bayesian model, was used to identify multiple spatial patterns, "footprints", of MNase-seq reads along the sperm genome. Regions predicted by CENTIPEDE analysis to be bound by a regulatory factor in sperm were correlated with genomic landmarks and higher order chromatin structure datasets to identify potential roles for these factors in regulating either prior or post spermatogenic, i.e., early embryonic events. This approach linked robust endogenous protamine transcription and transgene suppression to its chromatin environment within topologically associated domains. Of the candidate enhancer-bound regulatory proteins, Ctcf, was associated with chromatin domain boundaries in testes and embryonic stem cells. The continuity of Ctcf binding through the murine germline may permit rapid reconstitution of chromatin organization following fertilization. This likely reflects its preparation for early zygotic genome activation and comparatively accelerated preimplantation embryonic development program observed in mouse as compared to human and bull.
| S-EPMC4869110 | biostudies-literature
Project description:Eukaryotic genomes are packed into chromatin, whose basic repeating unit is the nucleosome. Nucleosome positioning is a widely researched area. A common experimental procedure to determine nucleosome positions involves the use of micrococcal nuclease (MNase). Here, we show that the cutting preference of MNase in combination with size selection generates a sequence-dependent bias in the resulting fragments. This strongly affects nucleosome positioning data and especially sequence-dependent models for nucleosome positioning. As a consequence we see a need to re-evaluate whether the DNA sequence is a major determinant of nucleosome positioning in vivo. More generally, our results show that data generated after MNase digestion of chromatin requires a matched control experiment in order to determine nucleosome positions.
| S-EPMC3012088 | biostudies-literature
Project description:Hormones are dynamic signaling molecules that influence gene activity and phenotype, and they are thus thought to play a central role in phenotypic evolution. In vertebrates, many fitness-related traits are mediated by the hormone testosterone (T), but the mechanisms by which T levels evolve are unclear. Here, we summarize a series of studies that advance our understanding of these mechanisms by comparing males from two subspecies of dark-eyed junco (Junco hyemalis) that differ in aggression, body size, and ornamentation. We first review our research demonstrating population differences in the time-course of T production, as well as findings that point to the gonad as a major source of this variation. In a common garden, the subspecies do not differ in pituitary output of luteinizing hormone, but males from the more androgenized subspecies have greater gonadal gene expression for specific steroidogenic enzymes, and they may be less sensitive to feedback along the hypothalamo-pituitary-gonadal (HPG) axis. Furthermore, we present new data from a common garden study demonstrating that the populations do not differ in gonadal sensitivity to gonadotropin-inhibitory hormone (i.e., GnIH receptor mRNA abundance), but the more androgenized subspecies expresses less gonadal mRNA for glucocorticoid receptor and mineralocorticoid receptor, suggesting altered cross-talk between the hypothalamo-pituitary-gonadal and -adrenal axes as another mechanism by which these subspecies have diverged in T production. These findings highlight the diversity of mechanisms that may generate functional variation in T and influence hormone-mediated phenotypic evolution.
| S-EPMC5922417 | biostudies-literature
Project description:Nucleosome positioning is critical to chromatin accessibility and is associated with gene expression programs in cells1-3. Previous nucleosome mapping methods assemble profiles from cell populations and reveal a cell-averaged pattern: nucleosomes are positioned and form a phased array that surrounds the transcription start sites of active genes3-6 and DNase I hypersensitive sites7. However, even in a homogenous population of cells, cells exhibit heterogeneity in expression in response to active signalling8,9 that may be related to heterogeneity in chromatin accessibility10-12. Here we report a technique, termed single-cell micrococcal nuclease sequencing (scMNase-seq), that can be used to simultaneously measure genome-wide nucleosome positioning and chromatin accessibility in single cells. Application of scMNase-seq to NIH3T3 cells, mouse primary naive CD4 T cells and mouse embryonic stem cells reveals two principles of nucleosome organization: first, nucleosomes in heterochromatin regions, or that surround the transcription start sites of silent genes, show large variation in positioning across different cells but are highly uniformly spaced along the nucleosome array; and second, nucleosomes that surround the transcription start sites of active genes and DNase I hypersensitive sites show little variation in positioning across different cells but are relatively heterogeneously spaced along the nucleosome array. We found a bimodal distribution of nucleosome spacing at DNase I hypersensitive sites, which corresponds to inaccessible and accessible states and is associated with nucleosome variation and variation in accessibility across cells. Nucleosome variation is smaller within single cells than across cells, and smaller within the same cell type than across cell types. A large fraction of naive CD4 T cells and mouse embryonic stem cells shows depleted nucleosome occupancy at the de novo enhancers detected in their respective differentiated lineages, revealing the existence of cells primed for differentiation to specific lineages in undifferentiated cell populations.
| S-EPMC8353605 | biostudies-literature