Nucleosome Positioning Patterns Research Articles

Chromatin- and nucleosome-associated features in liquid biopsy: implications for cancer biomarker discovery.

Cell-free DNA (cfDNA) from the bloodstream has been studied for cancer biomarker discovery, and chromatin-derived epigenetic features have come into the spotlight for their potential to expand clinical applications. Methylation, fragmentation, and nucleosome positioning patterns of cfDNA have previously been shown to reveal epigenomic and inferred transcriptomic information. More recently, histone modifications have emerged as a tool to further identify tumor-specific chromatin variants in plasma. A number of sequencing methods have been developed to analyze these epigenetic markers, offering new insights into tumor biology. Features within cfDNA allow for cancer detection, subtype and tissue of origin classification, and inference of gene expression. These methods provide a window into the complexity of cancer and the dynamic nature of its progression. In this review, we highlight the array of epigenetic features in cfDNA that can be extracted from chromatin- and nucleosome-associated organization and outline potential use cases in cancer management.

Epigenetic phenotype of plasma cell-free DNA in the prediction of early-onset preeclampsia

Background In the current study, we sought to characterise the methylation haplotypes and nucleosome positioning patterns of placental DNA and plasma cell-free DNA of pregnant women with early-onset preeclampsia using whole genome bisulphite sequencing (WGBS) and methylation capture bisulphite sequencing (MCBS) and further develop and examine the diagnostic performance of a generalised linear model (GLM) by incorporating the epigenetic features for early-onset preeclampsia. Methods This case-control study recruited pregnant women aged at least 18 years who delivered their babies at our Hospital. In addition, non-pregnant women with no previous history of diseases were included. Placental samples of the villous parenchyma were taken at the time of delivery and venous blood was drawn from pregnant women during non-invasive prenatal testing at 12–15 weeks of pregnancy and nonpregnant women during the physical check-up. WGBS and MCBS were carried out of extracted genomic DNA. Then, we established the GLM by incorporating preeclampsia-specific methylation haplotypes and nucleosome positioning patterns and examined the diagnostic performance of the model by receiver operating characteristic (ROC) curve analysis. Results The study included 135 pregnant women and 50 non-pregnant women. Our high-depth MCBS revealed notably different DNA methylation and nucleosome positioning patterns between women with and without preeclampsia. Preeclampsia-specific hypermethylated sites were found predominantly in the promoter regions and particularly enriched in CTCF on the X chromosome. Totally, 2379 preeclampsia-specific methylation haplotypes were found across the entire genome. ROC analysis showed that the area under the ROC curve (AUC) was 0.938 (95%CI 0.877, 1.000). At a GLM cut-off of 0.341, the AUC was the maximum, with a sensitivity of 95.6% and a specificity of 89.7%. Conclusion Pregnant women with early-onset preeclampsia exhibit DNA methylation and nucleosome positioning patterns in placental and plasma DNA.

Nucleosome Patterns in Circulating Tumor DNA Reveal Transcriptional Regulation of Advanced Prostate Cancer Phenotypes.

This study provides insights into the dynamics of nucleosome positioning and gene regulation associated with cancer phenotypes that can be ascertained from ctDNA. New methods for classification in phenotype mixtures extend the utility of ctDNA beyond assessments of somatic DNA alterations with important implications for molecular classification and precision oncology. This article is highlighted in the In This Issue feature, p. 517.

Distinct chromosomal "niches" in the genome of Saccharomyces cerevisiae provide the background for genomic innovation and shape the fate of gene duplicates.

Nearly one third of Saccharomyces cerevisiae protein coding sequences correspond to duplicate genes, equally split between small-scale duplicates (SSD) and whole-genome duplicates (WGD). While duplicate genes have distinct properties compared to singletons, to date, there has been no systematic analysis of their positional preferences. In this work, we show that SSD and WGD genes are organized in distinct gene clusters that occupy different genomic regions, with SSD being more peripheral and WGD more centrally positioned close to centromeric chromatin. Duplicate gene clusters differ from the rest of the genome in terms of gene size and spacing, gene expression variability and regulatory complexity, properties that are also shared by singleton genes residing within them. Singletons within duplicate gene clusters have longer promoters, more complex structure and a higher number of protein-protein interactions. Particular chromatin architectures appear to be important for gene evolution, as we find SSD gene-pair co-expression to be strongly associated with the similarity of nucleosome positioning patterns. We propose that specific regions of the yeast genome provide a favourable environment for the generation and maintenance of small-scale gene duplicates, segregating them from WGD-enriched genomic domains. Our findings provide a valuable framework linking genomic innovation with positional genomic preferences.

Dynamic nucleosome organization after fertilization reveals regulatory factors for mouse zygotic genome activation

Chromatin remodeling is essential for epigenome reprogramming after fertilization. However, the underlying mechanisms of chromatin remodeling remain to be explored. Here, we investigated the dynamic changes in nucleosome occupancy and positioning in pronucleus-stage zygotes using ultra low-input MNase-seq. We observed distinct features of inheritance and reconstruction of nucleosome positioning in both paternal and maternal genomes. Genome-wide de novo nucleosome occupancy in the paternal genome was observed as early as 1 h after the injection of sperm into ooplasm. The nucleosome positioning pattern was continually rebuilt to form nucleosome-depleted regions (NDRs) at promoters and transcription factor (TF) binding sites with differential dynamics in paternal and maternal genomes. NDRs formed more quickly on the promoters of genes involved in zygotic genome activation (ZGA), and this formation is closely linked to histone acetylation, but not transcription elongation or DNA replication. Importantly, we found that NDR establishment on the binding motifs of specific TFs might be associated with their potential pioneer functions in ZGA. Further investigations suggested that the predicted factors MLX and RFX1 played important roles in regulating minor and major ZGA, respectively. Our data not only elucidate the nucleosome positioning dynamics in both male and female pronuclei following fertilization, but also provide an efficient method for identifying key transcription regulators during development.

Cell identity bookmarking through heterogeneous chromatin landscape maintenance during the cell cycle.

Genetic and epigenetic information are faithfully duplicated and accurately transmitted to daughter cells to preserve cell identity during the cell cycle. However, how the chromatin-based epigenetic information beyond DNA sequence is stably transmitted along with the disruption and re-establishment of chromatin structure within a cell cycle remains largely unexplored. Through comprehensive analysis DNA methylation and nucleosome positioning patterns of HepG2 cells in G0/G1, early S, late S and G2/M phases, we found that DNA methylation may act as the prime element for epigenetic inheritance after replication, as DNA methylation was extremely stable in each cell cycle phase, while nucleosome occupancy showed notable phase dependent fluctuation. Nucleosome-Secured Regions (NSRs) occupied by polycomb-repressed chromatin played a role in repressing the irrelevant cell type-specific genes and were essential for preventing irrelevant transcription factors binding, while the well-defined Nucleosome-Depleted Regions (NDRs) marked the genes crucial for cell identity maintenance. Chromatin structure at NSRs and NDRs was well maintained throughout the cell cycle, which played crucial roles in steadily preserving the transcriptional identity of the cell to fulfill cell identity maintenance. Collectively, our results demonstrated that while chromatin architecture underwent dynamic changes during cell cycle progression, DNA methylation together with NSRs and NDRs were stable epigenetic elements that were required for faithful transmission to the daughter cell to accurately maintain cell identity during the cell cycle.

Fast, Accurate and Automatic Ancient Nucleosome and Methylation Maps with epiPALEOMIX.

The first epigenomes from archaic hominins (AH) and ancient anatomically modern humans (AMH) have recently been characterized, based, however, on a limited number of samples. The extent to which ancient genome-wide epigenetic landscapes can be reconstructed thus remains contentious. Here, we present epiPALEOMIX, an open-source and user-friendly pipeline that exploits post-mortem DNA degradation patterns to reconstruct ancient methylomes and nucleosome maps from shotgun and/or capture-enrichment data. Applying epiPALEOMIX to the sequence data underlying 35 ancient genomes including AMH, AH, equids and aurochs, we investigate the temporal, geographical and preservation range of ancient epigenetic signatures. We first assess the quality of inferred ancient epigenetic signatures within well-characterized genomic regions. We find that tissue-specific methylation signatures can be obtained across a wider range of DNA preparation types than previously thought, including when no particular experimental procedures have been used to remove deaminated cytosines prior to sequencing. We identify a large subset of samples for which DNA associated with nucleosomes is protected from post-mortem degradation, and nucleosome positioning patterns can be reconstructed. Finally, we describe parameters and conditions such as DNA damage levels and sequencing depth that limit the preservation of epigenetic signatures in ancient samples. When such conditions are met, we propose that epigenetic profiles of CTCF binding regions can be used to help data authentication. Our work, including epiPALEOMIX, opens for further investigations of ancient epigenomes through time especially aimed at tracking possible epigenetic changes during major evolutionary, environmental, socioeconomic, and cultural shifts.

Exploration of nucleosome positioning patterns in transcription factor function.

The binding of transcription factors (TFs) triggers activation of specific chromatin regions through the recruitment and activation of RNA polymerase. Unique nucleosome positioning (NP) occurs during gene expression and has been suggested to be involved in various other chromatin functions. However, the diversity of NP that can occur for each function has not been clarified. Here we used MNase-Seq data to evaluate NP around 258 cis-regulatory elements in the mouse genome. Principal component analysis of the 258 elements revealed that NP consisted of five major patterns. Furthermore, the five NP patterns had predictive power for the level of gene expression. We also demonstrated that selective NP patterns appeared around TF binding sites. These results suggest that the NP patterns are correlated to specific functions on chromatin.

Review fifteen years of search for strong nucleosomes

Don Crothers, Mikael Kubista, Jon Widom, and their teams have been first to look for strong nucleosomes, in a bid to reveal the nucleosome positioning pattern(s) carried by the nucleosome DNA sequences. They were first to demonstrate that the nucleosome stability correlates with 10-11 base sequence periodicity, and that the strong nucleosomes localize preferentially in centromeres. This review describes these findings and their connection to recent discovery of the strong nucleosomes (SNs) with visibly periodic nucleosome DNA sequences.

Genome-wide mapping of DNase I hypersensitive sites in plants.

Genomic regions associated with regulatory proteins are known to be highly sensitive to DNase I digestion and are termed DNase I hypersensitive sites (DHSs). DHSs can be identified by DNase I digestion followed by high-throughput DNA sequencing (DNase-seq). DNase-seq has become a powerful technique for genome-wide mapping of chromatin accessibility in eukaryotes with a sequenced genome. We have developed a DNase-seq procedure in plants. This procedure was adapted from the protocol originally developed for mammalian cell lines. It includes plant nuclei isolation, digestion of purified nuclei with DNase I, recovery of DNase-trimmed DNA fragments, DNase-seq library development, Illumina sequencing and data analysis. We also introduce a barcoding system for library preparation. We have conducted DNase-seq in both Arabidopsis thaliana and rice, and developed genome-wide open chromatin maps in both species. These DHS datasets have been used to detect footprints from regulatory protein binding and to reveal genome-wide nucleosome positioning patterns.

The chickpea genomic web resource: visualization and analysis of the desi-type Cicer arietinum nuclear genome for comparative exploration of legumes.

BackgroundAvailability of the draft nuclear genome sequences of small-seeded desi-type legume crop Cicer arietinum has provided an opportunity for investigating unique chickpea genomic features and evaluation of their biological significance. The increasing number of legume genome sequences also presents a challenge for developing reliable and information-driven bioinformatics applications suitable for comparative exploration of this important class of crop plants.ResultsThe Chickpea Genomic Web Resource (CGWR) is an implementation of a suite of web-based applications dedicated to chickpea genome visualization and comparative analysis, based on next generation sequencing and assembly of Cicer arietinum desi-type genotype ICC4958. CGWR has been designed and configured for mapping, scanning and browsing the significant chickpea genomic features in view of the important existing and potential roles played by the various legume genome projects in mutant mapping and cloning. It also enables comparative informatics of ICC4958 DNA sequence analysis with other wild and cultivated genotypes of chickpea, various other leguminous species as well as several non-leguminous model plants, to enable investigations into evolutionary processes that shape legume genomes.ConclusionsCGWR is an online database offering a comprehensive visual and functional genomic analysis of the chickpea genome, along with customized maps and gene-clustering options. It is also the only plant based web resource supporting display and analysis of nucleosome positioning patterns in the genome. The usefulness of CGWR has been demonstrated with discoveries of biological significance made using this server. The CGWR is compatible with all available operating systems and browsers, and is available freely under the open source license at http://www.nipgr.res.in/CGWR/home.php.

DNA Methylation Effects on Tetra-Nucleosome Compaction and Aggregation

DNA CpG methylation has been associated with chromatin compaction and gene silencing. Whether DNA methylation directly contributes to chromatin compaction remains an open question. In this study, we used fluorescence fluctuation spectroscopy (FFS) to evaluate the compaction and aggregation of tetra-nucleosomes containing specific CpG patterns and methylation levels. The compactness of both unmethylated and methylated tetra-nucleosomes is dependent on DNA sequences. Specifically, methylation of the CpG sites located in the central dyad and the major grooves of DNA seem to have opposite effects on modulating the compactness of tetra-nucleosomes. The interactions among tetra-nucleosomes, however, seem to be enhanced because of DNA methylation independent of sequence contexts. Our finding can shed light on understanding the role of DNA methylation in determining nucleosome positioning pattern and chromatin compactness.

Probabilistic partitioning methods to find significant patterns in ChIP-Seq data.

We have witnessed an enormous increase in ChIP-Seq data for histone modifications in the past few years. Discovering significant patterns in these data is an important problem for understanding biological mechanisms. We propose probabilistic partitioning methods to discover significant patterns in ChIP-Seq data. Our methods take into account signal magnitude, shape, strand orientation and shifts. We compare our methods with some current methods and demonstrate significant improvements, especially with sparse data. Besides pattern discovery and classification, probabilistic partitioning can serve other purposes in ChIP-Seq data analysis. Specifically, we exemplify its merits in the context of peak finding and partitioning of nucleosome positioning patterns in human promoters. The software and code are available in the supplementary material. Supplementary data are available at Bioinformatics online.

DNA Methylation Regulated Nucleosome Dynamics

A strong correlation between nucleosome positioning and DNA methylation patterns has been reported in literature. However, the mechanistic model accounting for the correlation remains elusive. In this study, we evaluated the effects of specific DNA methylation patterns on modulating nucleosome conformation and stability using FRET and SAXS. CpG dinucleotide repeats at 10 bp intervals were found to play different roles in nucleosome stability dependent on their methylation states and their relative nucleosomal locations. An additional (CpG)5 stretch located in the nucleosomal central dyad does not alter the nucleosome conformation, but significant conformational differences were observed between the unmethylated and methylated nucleosomes. These findings suggest that the correlation between nucleosome positioning and DNA methylation patterns can arise from the variations in nucleosome stability dependent on their sequence and epigenetic content. This knowledge will help to reveal the detailed role of DNA methylation in regulating chromatin packaging and gene transcription.

Determinants of nucleosome positioning

Nucleosome positioning is critical for gene expression and most DNA-related processes. Here we review the dominant patterns of nucleosome positioning that have been observed and summarize the current understanding of their underlying determinants. The genome-wide pattern of nucleosome positioning is determined by the combination of DNA sequence, ATP-dependent nucleosome remodeling enzymes and transcription factors that include activators, components of the preinitiation complex and elongating RNA polymerase II. These determinants influence each other such that the resulting nucleosome positioning patterns are likely to differ among genes and among cells in a population, with consequent effects on gene expression.

Dynamic Regulation of Nucleosome for Detection of CD80 Gene Expression in Murine

The basic building blocks of chromatin are referred to as the nucleosome. Nucleosomes play major role in transcriptional regulation, ranging from repression through occlusion of binding sites for transcription factors, to activation through spatial juxtaposition of transcription factor-binding sites. Examining the position of nucleosome in CD80 promoter is essential to detect the mechanism of tissue specific transcriptional regulation at chromatin level. The CD80 is the most potent of all of the co-stimulatory molecule, expressed on the surface of B cells upon antigen stimulation. The expression of CD80 is tightly regulated, normally limited to only a few cell types including B cells. Therefore this study is important, how nucleosomes become positioned during gene regulation in diverse murine tissues and cell lines. The upstream region of CD80 promoter was examined for its nucleosomal organization by low resolution analysis of MNase-southern blot assay. Four nucleosomes were detected in all murine tissues and cells screened for this analysis. Curiously, similar pattern of nucleosome positioning was detected in all the tissues and cell lines screened. This results conclude that there is no regulation at the level of nucleosome positioning in murine CD80 promoter. However, there must be a regulatory mechanism involved in these tissues either at the molecular level or chromatin level or both as the CD80 gene expression was detected in organs rich in antigen presenting cells, spleen, thymus and lung. This was addressed by possible remodeling mechanism through covalent modification of histone proteins which mark for active transcription and repression of gene regulation. Journal of Food and Agriculture 2010 Vol. 03, No. 1 & 2, Page 14-23 DOI: http://dx.doi.org/10.4038/jfa.v3i1-2.5166

A Functional Evolutionary Approach to Identify Determinants of Nucleosome Positioning: A Unifying Model for Establishing the Genome-wide Pattern

Although the genomic pattern of nucleosome positioning is broadly conserved, quantitative aspects vary over evolutionary timescales. We identify the cis and trans determinants of nucleosome positioning using a functional evolutionary approach involving S.cerevisiae strains containing large genomic regions from other yeast species. In a foreign species, nucleosome depletion at promoters is maintained over poly(dA:dT) tracts, whereas internucleosome spacing and all other aspects of nucleosome positioning tested are not. Interestingly, the locations of the+1 nucleosome and RNA start sites shift in concert. Strikingly, in a foreign species, nucleosome-depleted regions occur fortuitously in coding regions, and they often act as promoters that are associated with a positioned nucleosome array linked to the length of the transcription unit. We suggest a three-step model in which nucleosome remodelers, general transcription factors, and the transcriptional elongation machinery are primarily involved in generating the nucleosome positioning pattern invivo.

Nucleosome Positioning Patterns Derived from Human Apoptotic Nucleosomes

This communication reports on the nucleosome positioning patterns (bendability matrices) for the human genome, derived from over 8 million nucleosome DNA sequences obtained from apoptotically digested lymphocytes. This digestion procedure is used here for the first time for the purpose of extraction and sequencing of the nucleosome DNA fragments. The dominant motifs suggested by the matrices of DNA bendability calculated for light and heavy isochores are significantly different. Both, however, are in full agreement with the linear description YRRRRRYYYYYR, and with earlier derivations by N-gram extensions. Thus, the choice of the nucleosome positioning patterns crucially depends on the G + C composition of the analyzed sequences.

Physical properties of naked DNA influence nucleosome positioning and correlate with transcription start and termination sites in yeast

BackgroundIn eukaryotic organisms, DNA is packaged into chromatin structure, where most of DNA is wrapped into nucleosomes. DNA compaction and nucleosome positioning have clear functional implications, since they modulate the accessibility of genomic regions to regulatory proteins. Despite the intensive research effort focused in this area, the rules defining nucleosome positioning and the location of DNA regulatory regions still remain elusive.ResultsNaked (histone-free) and nucleosomal DNA from yeast were digested by microccocal nuclease (MNase) and sequenced genome-wide. MNase cutting preferences were determined for both naked and nucleosomal DNAs. Integration of their sequencing profiles with DNA conformational descriptors derived from atomistic molecular dynamic simulations enabled us to extract the physical properties of DNA on a genomic scale and to correlate them with chromatin structure and gene regulation. The local structure of DNA around regulatory regions was found to be unusually flexible and to display a unique pattern of nucleosome positioning. Ab initio physical descriptors derived from molecular dynamics were used to develop a computational method that accurately predicts nucleosome enriched and depleted regions.ConclusionsOur experimental and computational analyses jointly demonstrate a clear correlation between sequence-dependent physical properties of naked DNA and regulatory signals in the chromatin structure. These results demonstrate that nucleosome positioning around TSS (Transcription Start Site) and TTS (Transcription Termination Site) (at least in yeast) is strongly dependent on DNA physical properties, which can define a basal regulatory mechanism of gene expression.

“Anticipated” nucleosome positioning pattern in prokaryotes

Linguistic (word count) analysis of prokaryotic genome sequences, by Shannon N-gram extension, reveals that the dominant hidden motifs in A + T rich genomes are T(A)(T)A and G(A)(T)C with uncertain number of repeating A and T. Since prokaryotic sequences are largely protein-coding, the motifs would correspond to amphipathic alpha-helices with alternating lysine and phenylalanine as preferential polar and non-polar residues. The motifs are also known in eukaryotes, as nucleosome positioning patterns. Their existence in prokaryotes as well may serve for binding of histone-like proteins to DNA. In this case the above patterns in prokaryotes may be considered as “anticipated” nucleosome positioning patterns which, quite likely, existed in prokaryotic genomes before the evolutionary separation between eukaryotes and prokaryotes.