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Journal articles on the topic 'Chromatin sequencing'
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1
Gather, Fabian. "Chromatin Immunoprecipitation Sequencing (ChIPseq)." Annals of Anatomy - Anatomischer Anzeiger 260 (June 2025): 152421. https://doi.org/10.1016/j.aanat.2025.152421.
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Soleimani, Vahab D., Gareth A. Palidwor, Parameswaran Ramachandran, Theodore J. Perkins, and Michael A. Rudnicki. "Chromatin tandem affinity purification sequencing." Nature Protocols 8, no. 8 (2013): 1525–34. http://dx.doi.org/10.1038/nprot.2013.088.
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Jukam, David, Charles Limouse, Owen K. Smith, Viviana I. Risca, Jason C. Bell, and Aaron F. Straight. "Chromatin‐Associated RNA Sequencing (ChAR‐seq)." Current Protocols in Molecular Biology 126, no. 1 (2019): e87. http://dx.doi.org/10.1002/cpmb.87.
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Stergachis, Andrew B., Brian M. Debo, Eric Haugen, L. Stirling Churchman, and John A. Stamatoyannopoulos. "Single-molecule regulatory architectures captured by chromatin fiber sequencing." Science 368, no. 6498 (2020): 1449–54. http://dx.doi.org/10.1126/science.aaz1646.
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Gene regulation is chiefly determined at the level of individual linear chromatin molecules, yet our current understanding of cis-regulatory architectures derives from fragmented sampling of large numbers of disparate molecules. We developed an approach for precisely stenciling the structure of individual chromatin fibers onto their composite DNA templates using nonspecific DNA N6-adenine methyltransferases. Single-molecule long-read sequencing of chromatin stencils enabled nucleotide-resolution readout of the primary architecture of multikilobase chromatin fibers (Fiber-seq). Fiber-seq exposed widespread plasticity in the linear organization of individual chromatin fibers and illuminated principles guiding regulatory DNA actuation, the coordinated actuation of neighboring regulatory elements, single-molecule nucleosome positioning, and single-molecule transcription factor occupancy. Our approach and results open new vistas on the primary architecture of gene regulation.
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Xie, Wenhui, Yilang Ke, Qinyi You, et al. "Single-Cell RNA Sequencing and Assay for Transposase-Accessible Chromatin Using Sequencing Reveals Cellular and Molecular Dynamics of Aortic Aging in Mice." Arteriosclerosis, Thrombosis, and Vascular Biology 42, no. 2 (2022): 156–71. http://dx.doi.org/10.1161/atvbaha.121.316883.
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Objective: The impact of vascular aging on cardiovascular diseases has been extensively studied; however, little is known regarding the cellular and molecular mechanisms underlying age-related vascular aging in aortic cellular subpopulations. Approach and Results: Transcriptomes and transposase-accessible chromatin profiles from the aortas of 4-, 26-, and 86-week-old C57/BL6J mice were analyzed using single-cell RNA sequencing and assay for transposase-accessible chromatin sequencing. By integrating the heterogeneous transcriptome and chromatin accessibility data, we identified cell-specific TF (transcription factor) regulatory networks and open chromatin states. We also determined that aortic aging affects cell interactions, inflammation, cell type composition, dysregulation of transcriptional control, and chromatin accessibility. Endothelial cells 1 have higher gene set activity related to cellular senescence and aging than do endothelial cells 2. Moreover, construction of senescence trajectories shows that endothelial cell 1 and fibroblast senescence is associated with distinct TF open chromatin states and an mRNA expression model. Conclusions: Our data provide a system-wide model for transcriptional and epigenetic regulation during aortic aging at single-cell resolution.
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Gorkin, David U., Iros Barozzi, Yuan Zhao, et al. "An atlas of dynamic chromatin landscapes in mouse fetal development." Nature 583, no. 7818 (2020): 744–51. http://dx.doi.org/10.1038/s41586-020-2093-3.
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AbstractThe Encyclopedia of DNA Elements (ENCODE) project has established a genomic resource for mammalian development, profiling a diverse panel of mouse tissues at 8 developmental stages from 10.5 days after conception until birth, including transcriptomes, methylomes and chromatin states. Here we systematically examined the state and accessibility of chromatin in the developing mouse fetus. In total we performed 1,128 chromatin immunoprecipitation with sequencing (ChIP–seq) assays for histone modifications and 132 assay for transposase-accessible chromatin using sequencing (ATAC–seq) assays for chromatin accessibility across 72 distinct tissue-stages. We used integrative analysis to develop a unified set of chromatin state annotations, infer the identities of dynamic enhancers and key transcriptional regulators, and characterize the relationship between chromatin state and accessibility during developmental gene regulation. We also leveraged these data to link enhancers to putative target genes and demonstrate tissue-specific enrichments of sequence variants associated with disease in humans. The mouse ENCODE data sets provide a compendium of resources for biomedical researchers and achieve, to our knowledge, the most comprehensive view of chromatin dynamics during mammalian fetal development to date.
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Wu, Weixin, Zhangming Yan, Tri C. Nguyen, Zhen Bouman Chen, Shu Chien, and Sheng Zhong. "Mapping RNA–chromatin interactions by sequencing with iMARGI." Nature Protocols 14, no. 11 (2019): 3243–72. http://dx.doi.org/10.1038/s41596-019-0229-4.
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Jahan, Sanzida, Tasnim H. Beacon, Wayne Xu, and James R. Davie. "Atypical chromatin structure of immune-related genes expressed in chicken erythrocytes." Biochemistry and Cell Biology 98, no. 2 (2020): 171–77. http://dx.doi.org/10.1139/bcb-2019-0107.
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The major biological role of red blood cells is to carry oxygen to the tissues in the body. However, another role of the erythroid cell is to participate in the immune response. Mature erythrocytes from chickens express Toll-like receptors and several cytokines in response to stimulation of the immune system. We previously reported the application of a biochemical fractionation protocol to isolate highly enriched transcribed DNA from polychromatic erythrocytes from chickens. In conjunction with next-generation DNA, RNA sequencing, chromatin immunoprecipitation-DNA sequencing, and formaldehyde-assisted isolation of regulatory elements (FAIRE) sequencing, we identified the active chromosomal compartments and determined their structural signatures in relation to expression levels. Here, we present the detailed chromatin characteristics of erythroid genes participating in the innate immune response. Our studies revealed an atypical chromatin structure for several genes coding for Toll-like receptors, interleukins, and interferon regulatory factors. The body of these genes had nucleosome-free regions intermingled with nucleosomes modified with H3K4me3 and H3K27ac, suggesting a dynamic unstable chromatin structure. We further show that human genes involved in cell identity have gene bodies with the same chromatin-instability features as the chicken polychromatic erythrocyte genes participating in the innate immune response.
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Haghani, Viktoria, Aditi Goyal, Alan Zhang, et al. "Improving rigor and reproducibility in chromatin immunoprecipitation assay data analysis workflows with Rocketchip." F1000Research 14 (June 25, 2025): 625. https://doi.org/10.12688/f1000research.164319.1.
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Background As genome sequencing technologies advance, the growing accumulation of sequencing data in public databases highlights the need for more robust and adaptable analysis workflows for chromatin immunoprecipitation assays. These workflows must promote reproducibility and replicability of results while effectively leveraging publicly available data to enhance biological insights. Methods Here, we present Rocketchip, a Python-based command-line tool that integrates existing software through Snakemake, enabling efficient, automated, and reproducible analysis of both newly generated chromatin immunoprecipitation data, including chromatin immunoprecipitation followed by sequencing (ChIP-seq), cleavage under targets and release using nuclease (CUT&RUN), and cleavage under targets and tagmentation (CUT&Tag) as well as existing datasets. Results Rocketchip can analyze chromatin immunoprecipitation data using any available software combination for any reference genome. It enhances the utilization of existing datasets, promotes replicability in analyses, and serves as a platform for benchmarking algorithms. Conclusions By facilitating the reanalysis of existing data and allowing for flexible analysis of newly generated data, Rocketchip allows researchers to identify and apply the most accurate and efficient analytical approaches to their data.
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Marr, Luke T., Prasoon Jaya, Laxmi N. Mishra, and Jeffrey J. Hayes. "Whole-genome methods to define DNA and histone accessibility and long-range interactions in chromatin." Biochemical Society Transactions 50, no. 1 (2022): 199–212. http://dx.doi.org/10.1042/bst20210959.
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Defining the genome-wide chromatin landscape has been a goal of experimentalists for decades. Here we review highlights of these efforts, from seminal experiments showing discontinuities in chromatin structure related to gene activation to extensions of these methods elucidating general features of chromatin related to gene states by exploiting deep sequencing methods. We also review chromatin conformational capture methods to identify patterns in long-range interactions between genomic loci.
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Li, Niannian, Kairang Jin, Yanmin Bai, Haifeng Fu, Lin Liu, and Bin Liu. "Tn5 Transposase Applied in Genomics Research." International Journal of Molecular Sciences 21, no. 21 (2020): 8329. http://dx.doi.org/10.3390/ijms21218329.
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The development of high-throughput sequencing (next-generation sequencing technology (NGS)) and the continuous increase in experimental throughput require the upstream sample processing steps of NGS to be as simple as possible to improve the efficiency of the entire NGS process. The transposition system has fast “cut and paste” and “copy and paste” functions, and has been innovatively applied to the NGS field. For example, the Assay for Transposase-Accessible Chromatin with high throughput sequencing (ATAC-Seq) uses high-throughput sequencing to detect chromatin regions accessible by Tn5 transposase. Linear Amplification via Transposon Insertion (LIANTI) uses Tn5 transposase for linear amplification, haploid typing, and structural variation detection. Not only is it efficient and simple, it effectively shortens the time for NGS sample library construction, realizes large-scale and rapid sequencing, improves sequencing resolution, and can be flexibly modified for more technological innovation.
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Fittipaldi, Raffaella, and Giuseppina Caretti. "Tackling Skeletal Muscle Cells Epigenome in the Next-Generation Sequencing Era." Comparative and Functional Genomics 2012 (2012): 1–8. http://dx.doi.org/10.1155/2012/979168.
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Recent advances in high-throughput technologies have transformed methodologies employed to study cell-specific epigenomes and the approaches to investigate complex cellular phenotypes. Application of next-generation sequencing technology in the skeletal muscle differentiation field is rapidly extending our knowledge on how chromatin modifications, transcription factors and chromatin regulators orchestrate gene expression pathways guiding myogenesis. Here, we review recent biological insights gained by the application of next-generation sequencing techniques to decode the epigenetic profile and gene regulatory networks underlying skeletal muscle differentiation.
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Murdoch, Brenda M., Kimberly M. Davenport, Shangqian Xie, et al. "378 Characterizing Functional Genetic Regulatory Elements in Sheep Reference Genome." Journal of Animal Science 100, Supplement_3 (2022): 185. http://dx.doi.org/10.1093/jas/skac247.340.
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Abstract Characterizing the locations of genetic regulatory elements is critical for understanding the regulatory mechanisms of complex phenotypic traits related to production traits and health in livestock species. The Ovine Functional Annotation of Animal Genomes (FAANG) Project aims to characterize transcriptional regulatory elements across the sheep genome to facilitate a better understanding of the biological mechanisms influencing phenotypic traits in sheep. Assays including sequencing of messenger RNA (mRNA-seq), cap analysis of gene expression (CAGE), chromatin immunoprecipitation of histones (ChIP-seq), assay for transposase-accessible chromatin (ATAC-seq), whole genome bisulfite sequencing (WGBS) and reduced representation bisulfite sequencing (RRBS) were performed on tissues collected from the Rambouillet ewe used to assemble the reference genome ARS-UI_Ramb_v2.0. Histone modifications were used to define nine chromatin states for tissues across the genome depicting promoters and enhancers (active, poised, and repressed) using ChromHMM. Chromatin states were overlayed with RNA-seq, ATAC-seq and DNA methylation. These data suggest that active promoter and enhancer states reside in open chromatin regions with a greater transcriptional activity and hypomethylated regions than other states. Further, poised and repressed enhancers did not primarily reside in open chromatin and had less transcriptional activity and more hypermethylated sites compared with active states. Collectively these data define transcriptional regulatory regions throughout the ovine genome which provides a valuable resource to better understand regulatory regions in the genome and how these influence economically important traits in sheep and other livestock species.
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Vasilev, V. A., D. M. Ryabov, A. K. Shaytan, and G. A. Armeev. "Updating nucleosome positions within individual genes using molecular modeling methods and mnase sequencing data." Биофизика 68, no. 5 (2023): 911–19. http://dx.doi.org/10.31857/s0006302923050101.
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Organization of chromatin plays an important role in regulating the genetic machinery of the cell. The basic unit of chromatin packaging is a nucleosome, which harbors DNA of about 145 base pairs in length. The packaging of genetic material and its accessibility to transcription enzymes and other regulatory chromatin proteins depends on the positions of nucleosomes. MNase sequencing is used to examine nucleosome positions in a genome. MNase sequencing data are sufficient for detecting the presence of nucleosomes on the sequence, but a determination of the precise locations of nucleosomes can be problematic. Accurate determination of nucleosome positions requires additional data filtering and processing. In this study, using MNase sequencing data, a combined method based on geometric analysis of nucleosome chain molecular models is proposed for selecting possible nucleosome positions. The developed algorithm efficiently eliminates inaccessible nucleosome chain combinations and conformationally prohibited nucleosome positions.
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Romanowska, Julia, and Anagha Joshi. "From Genotype to Phenotype: Through Chromatin." Genes 10, no. 2 (2019): 76. http://dx.doi.org/10.3390/genes10020076.
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Advances in sequencing technologies have enabled the exploration of the genetic basis for several clinical disorders by allowing identification of causal mutations in rare genetic diseases. Sequencing technology has also facilitated genome-wide association studies to gather single nucleotide polymorphisms in common diseases including cancer and diabetes. Sequencing has therefore become common in the clinic for both prognostics and diagnostics. The success in follow-up steps, i.e., mapping mutations to causal genes and therapeutic targets to further the development of novel therapies, has nevertheless been very limited. This is because most mutations associated with diseases lie in inter-genic regions including the so-called regulatory genome. Additionally, no genetic causes are apparent for many diseases including neurodegenerative disorders. A complementary approach is therefore gaining interest, namely to focus on epigenetic control of the disease to generate more complete functional genomic maps. To this end, several recent studies have generated large-scale epigenetic datasets in a disease context to form a link between genotype and phenotype. We focus DNA methylation and important histone marks, where recent advances have been made thanks to technology improvements, cost effectiveness, and large meta-scale epigenome consortia efforts. We summarize recent studies unravelling the mechanistic understanding of epigenetic processes in disease development and progression. Moreover, we show how methodology advancements enable causal relationships to be established, and we pinpoint the most important issues to be addressed by future research.
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Guo, Ziwei, Xinhong Liu, and Mo Chen. "Defining pervasive transcription units using chromatin RNA-sequencing data." STAR Protocols 3, no. 2 (2022): 101442. http://dx.doi.org/10.1016/j.xpro.2022.101442.
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Vega, Vinsensius B., Edwin Cheung, Nallasivam Palanisamy, and Wing-Kin Sung. "Inherent Signals in Sequencing-Based Chromatin-ImmunoPrecipitation Control Libraries." PLoS ONE 4, no. 4 (2009): e5241. http://dx.doi.org/10.1371/journal.pone.0005241.
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Bright, Ann Rose, and Gert Jan C. Veenstra. "Assay for Transposase-Accessible Chromatin-Sequencing Using Xenopus Embryos." Cold Spring Harbor Protocols 2019, no. 1 (2018): pdb.prot098327. http://dx.doi.org/10.1101/pdb.prot098327.
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Wang, Xilin. "ATAC-seq: A powerful tool for investigating chromatin accessibility and transcription factor binding." Theoretical and Natural Science 6, no. 1 (2023): 316–22. http://dx.doi.org/10.54254/2753-8818/6/20230267.
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Chromatin Transposase Accessibility Sequencing is a new high-throughput sequencing technique developed by Professor William Greenleaf in 2013 which uses DNA transposase to probe chromatin accessibility with Tn5 transposase. This technique, which is simpler and more sensitive than DNase-seq, MNase-seq and FAIRE-seq and requires fewer cells, has been used to study chromatin accessibility using Tn5 transposase. ATAC-seq is important for the study of epigenetic molecular mechanisms because it can map chromatin accessibility on a genome-wide scale, compare open chromatin regions in different tumour samples, compare differences in transcription factor binding between treatments, reveal nucleosome localisation information and transcription factor binding sites, and can be used to locate specific unknown transcription factors, which can be used in combination with other methods to screen for specific transcription factors of interest. It is possible to combine this approach with others to investigate specific regulatory factors. Herein, ATAC-seq is systemically profiled to present that ATAC-seq has enormous potential to drive future discoveries in the field of genomics and beyond.
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Baumgarten, Sebastian, and Jessica Bryant. "Chromatin structure can introduce systematic biases in genome-wide analyses of Plasmodium falciparum." Open Research Europe 2 (September 15, 2022): 75. http://dx.doi.org/10.12688/openreseurope.14836.2.
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Background: The maintenance, regulation, and dynamics of heterochromatin in the human malaria parasite, Plasmodium falciparum, has drawn increasing attention due to its regulatory role in mutually exclusive virulence gene expression and the silencing of key developmental regulators. The advent of genome-wide analyses such as chromatin-immunoprecipitation followed by sequencing (ChIP-seq) has been instrumental in understanding chromatin composition; however, even in model organisms, ChIP-seq experiments are susceptible to intrinsic experimental biases arising from underlying chromatin structure. Methods: We performed a control ChIP-seq experiment, re-analyzed previously published ChIP-seq datasets and compared different analysis approaches to characterize biases of genome-wide analyses in P. falciparum. Results: We found that heterochromatic regions in input control samples used for ChIP-seq normalization are systematically underrepresented in regard to sequencing coverage across the P. falciparum genome. This underrepresentation, in combination with a non-specific or inefficient immunoprecipitation, can lead to the identification of false enrichment and peaks across these regions. We observed that such biases can also be seen at background levels in specific and efficient ChIP-seq experiments. We further report on how different read mapping approaches can also skew sequencing coverage within highly similar subtelomeric regions and virulence gene families. To ameliorate these issues, we discuss orthogonal methods that can be used to characterize bona fide chromatin-associated proteins. Conclusions: Our results highlight the impact of chromatin structure on genome-wide analyses in the parasite and the need for caution when characterizing chromatin-associated proteins and features.
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Baumgarten, Sebastian, and Jessica Bryant. "Chromatin structure can introduce systematic biases in genome-wide analyses of Plasmodium falciparum." Open Research Europe 2 (June 10, 2022): 75. http://dx.doi.org/10.12688/openreseurope.14836.1.
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Background: The maintenance, regulation, and dynamics of heterochromatin in the human malaria parasite, Plasmodium falciparum, has drawn increasing attention due to its regulatory role in mutually exclusive virulence gene expression and the silencing of key developmental regulators. The advent of genome-wide analyses such as chromatin-immunoprecipitation followed by sequencing (ChIP-seq) has been instrumental in understanding chromatin composition; however, even in model organisms, ChIP-seq experiments are susceptible to intrinsic experimental biases arising from underlying chromatin structure. Methods: We performed a control ChIP-seq experiment, re-analyzed previously published ChIP-seq datasets and compared different analysis approaches to characterize biases of genome-wide analyses in P. falciparum. Results: We found that heterochromatic regions in input control samples used for ChIP-seq normalization are systematically underrepresented in regard to sequencing coverage across the P. falciparum genome. This underrepresentation, in combination with a non-specific or inefficient immunoprecipitation, can lead to the identification of false enrichment and peaks across these regions. We observed that such biases can also be seen at background levels in specific and efficient ChIP-seq experiments. We further report on how different read mapping approaches can also skew sequencing coverage within highly similar subtelomeric regions and virulence gene families. To ameliorate these issues, we discuss orthogonal methods that can be used to characterize bona fide chromatin-associated proteins. Conclusions: Our results highlight the impact of chromatin structure on genome-wide analyses in the parasite and the need for caution when characterizing chromatin-associated proteins and features.
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Stevens, Claire, Leonardo Gonzalez-Smith, Huan Cao, and Suhn K. Rhie. "Abstract 7013: Methyl-Micro-C: simultaneous high-resolution characterization of three-dimensional chromatin structure and the DNA methylome." Cancer Research 84, no. 6_Supplement (2024): 7013. http://dx.doi.org/10.1158/1538-7445.am2024-7013.
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Abstract Understanding cancer biology allows for the determination of patient prognosis through biomarkers and predicting treatment sensitivity. For many cancer types, genetic changes have been largely characterized, but the contributions of epigenetic changes, which are also implicated in tumorigenesis, are not yet well understood. Three-dimensional chromatin architecture changes occur throughout tumor development, changing the states and interactions between cis-regulatory elements, overall inducing gene dysregulation. Chromatin structure changes have previously been interrogated using Hi-C, a technique which utilizes restriction enzyme-mediated digestion of chromatin. Although this method is useful, it cannot detect chromatin states of regulatory elements with high sensitivity and resolution due to the use of restriction enzymes that give multi-nucleosome-sized fragments. Micro-C instead uses MNase that preferentially digests accessible regulatory elements, resulting in mono, di, and tri-nucleosome fragments, providing higher-resolution chromatin interaction data, indirectly determining which regulatory elements are active and in proximity to one another. However, spatial proximity of genomic regions are only a piece of the chromatin structure epigenetic story. Partially methylated domains, which are gained or altered throughout tumor development, are reported to co-localize with topologically associating domains, suggesting that they can contribute to chromatin structure alterations. Whole methylome sequencing techniques can be used to determine methylation states of these domains and regulatory regions. Recently, enzyme-mediated methylation sequencing has been shown to induce less DNA damage than the canonical whole-genome bisulfite sequencing, providing a more accurate and comprehensive picture of the DNA methylome. Although DNA methylome data alone can provide information about epigenetic status within genomic regions, overlaying chromatin structure changes at these locations can further corroborate findings. In this way, chromatin structure and DNA methylome data together can provide a more complete picture of cancer epigenetics. However, it is very expensive to sequence at the depth necessary to gain useful information. To address this issue, we developed a technique called Methyl-Micro-C to simultaneously interrogate the DNA methylome while observing chromatin structure in the same sample. Methyl-Micro-C integrates the higher resolution Micro-C method and more comprehensive enzyme-mediated methylation sequencing method together to investigate both the chromatin interactions as well as DNA methylation patterns for the same sample simultaneously. Here, we use Methyl-Micro-C with prostate cancer cells to characterize 3D epigenomic mechanisms that drive prostate carcinogenesis. Citation Format: Claire Stevens, Leonardo Gonzalez-Smith, Huan Cao, Suhn K. Rhie. Methyl-Micro-C: simultaneous high-resolution characterization of three-dimensional chromatin structure and the DNA methylome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7013.
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Reeves, G. Adam, Param Priya Singh, and Anne Brunet. "Chromatin Accessibility Profiling and Data Analysis Using ATAC-seq inNothobranchius furzeri." Cold Spring Harbor Protocols 2024, no. 3 (2023): pdb.prot107747. http://dx.doi.org/10.1101/pdb.prot107747.
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The state of genome-wide chromatin accessibility in cells, tissues, or organisms can be investigated with a technique called assay for transposase-accessible chromatin using sequencing (ATAC-seq). ATAC-seq is a powerful approach for profiling the epigenomic landscape of cells using very low input materials. Analysis of chromatin accessibility data allows for prediction of gene expression and identification of regulatory elements such as potential enhancers and specific transcription-factor binding sites. Here, we describe an optimized ATAC-seq protocol for the preparation of isolated nuclei and subsequent next-generation sequencing from whole embryos and tissues of the African turquoise killifish (Nothobranchius furzeri). Importantly, we provide an overview of a pipeline for processing and analyzing ATAC-seq data from killifish.
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Eapen, Amy A., Sreeja Parameswaran, Carmy Forney, et al. "Epigenetic and transcriptional dysregulation in CD4+ T cells in patients with atopic dermatitis." PLOS Genetics 18, no. 5 (2022): e1009973. http://dx.doi.org/10.1371/journal.pgen.1009973.
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Atopic dermatitis (AD) is one of the most common skin disorders among children. Disease etiology involves genetic and environmental factors, with 29 independent AD risk loci enriched for risk allele-dependent gene expression in the skin and CD4+ T cell compartments. We investigated the potential epigenetic mechanisms responsible for the genetic susceptibility of CD4+ T cells. To understand the differences in gene regulatory activity in peripheral blood T cells in AD, we measured chromatin accessibility (an assay based on transposase-accessible chromatin sequencing, ATAC-seq), nuclear factor kappa B subunit 1 (NFKB1) binding (chromatin immunoprecipitation with sequencing, ChIP-seq), and gene expression levels (RNA-seq) in stimulated CD4+ T cells from subjects with active moderate-to-severe AD, as well as in age-matched and non-allergic controls. Open chromatin regions in stimulated CD4+ T cells were highly enriched for AD genetic risk variants, with almost half of the AD risk loci overlapping AD-dependent ATAC-seq peaks. AD-specific open chromatin regions were strongly enriched for NF-κB DNA-binding motifs. ChIP-seq identified hundreds of NFKB1-occupied genomic loci that were AD- or control-specific. As expected, the AD-specific ChIP-seq peaks were strongly enriched for NF-κB DNA-binding motifs. Surprisingly, control-specific NFKB1 ChIP-seq peaks were not enriched for NFKB1 motifs, but instead contained motifs for other classes of human transcription factors, suggesting a mechanism involving altered indirect NFKB1 binding. Using DNA sequencing data, we identified 63 instances of altered genotype-dependent chromatin accessibility at 36 AD risk variant loci (30% of AD risk loci) that might lead to genotype-dependent gene expression. Based on these findings, we propose that CD4+ T cells respond to stimulation in an AD-specific manner, resulting in disease- and genotype-dependent chromatin accessibility alterations involving NFKB1 binding.
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Li, Wangchun, U. Tim Wu, Yu Cheng, et al. "Epigenetic Application of ATAC-Seq Based on Tn5 Transposase Purification Technology." Genetics Research 2022 (August 11, 2022): 1–9. http://dx.doi.org/10.1155/2022/8429207.
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Background. Assays of transposase accessible chromatin sequencing (ATAC-seq) is an efficient assay to investigate chromatin accessibility, which depends on the activity of a robust Tn5 transposase to fragment the genome while cutting in the sequencing adapters. Methods. We propose reliable approaches for purifying hyperactive Tn5 transposase by chitin magnetic bead sorting. Double-stranded DNA of J76 cells and 293T cells were digested and subjected to tagmentation as test samples with Tn5 transposase, and libraries were established and sequenced. Sequencing data was then analyzed for peak calling, GO enrichment, and motif analysis. Results. We report a set of rapid, efficient, and low-cost methods for ATAC-seq library construction and data analysis, through large-scale and rapid sequencing. These methods can provide a reference for the study of epigenetic regulation of gene expression.
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Oh, Kyu Seon, Jisu Ha, Songjoon Baek, and Myong-Hee Sung. "XL-DNase-seq: Improved footprinting of dynamic transcription factors." Journal of Immunology 202, no. 1_Supplement (2019): 125.17. http://dx.doi.org/10.4049/jimmunol.202.supp.125.17.
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Abstract As the cost of high-throughput sequencing technologies decreases, genome-wide chromatin accessibility profiling methods such as the assay of transposase-accessible chromatin using sequencing (ATAC-seq) are employed widely, with data accumulating at an unprecedented rate. However, accurate inference of protein occupancy requires higher resolution footprinting analysis where major hurdles exist, including the sequence bias of nucleases and the short-lived chromatin binding of many transcription factors (TFs) with consequent lack of footprints. Here we introduce an assay termed crosslink (XL)-DNase-seq, designed to capture chromatin interactions of dynamic TFs. Mild crosslinking improved the detection of DNase-based footprints of dynamic TFs but interfered with ATAC-based footprinting of the same TFs. XL-DNase-seq may help extract novel gene regulatory circuits involving previously undetectable TFs. The DNase-seq and ATAC-seq data generated in our systematic comparison of various crosslinking conditions also represent an unprecedented-scale resource derived from activated mouse macrophage-like cells which share many features of inflammatory macrophages.
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Venters, Bryan J., Paul W. Hook, Vishnu S. Kumary, et al. "Abstract 7026: Multi-omic genomic mapping with long read sequencing." Cancer Research 84, no. 6_Supplement (2024): 7026. http://dx.doi.org/10.1158/1538-7445.am2024-7026.
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Abstract Gene transcription is regulated by the complex interplay between histone post-translational modifications (PTMs), chromatin associated proteins (CAPs), and DNA methylation (DNAme). Mapping their genomic locations and examining the relationships between these chromatin elements is a powerful approach to decipher mechanisms of disease, thereby enabling discovery of novel biomarkers and therapeutics. Leading epigenomic mapping technologies (e.g., ChIP-seq, CUT&RUN) rely upon DNA fragmentation to isolate regions of interest for sequencing on short read platforms (e.g., Illumina). This strategy leads to substantial loss of contextual information regarding the surrounding DNA, precluding the identification of multiple co-occurring epigenomic features on a single DNA molecule. By contrast, long-read sequencing (LRS) platforms are capable of sequencing very long reads from a single molecule (typically >10kb), allowing relationships between features on a single molecule to be used to resolve heterogeneity within mixed populations. Here we report a robust multi-omic method that leverages LRS to simultaneously profile histone PTMs (or CAPs), DNAme, and parental haplotype in a single assay. This nondestructive, epigenomic mapping approach leverages a novel DNA methyltransferase fusion protein (pAG-M.EcoGII) to label DNA underneath antibody-targeted chromatin features, thereby marking sites of interest while preserving DNA molecules intact for LRS. Inspired by our work with state-of-the-art immunotethering-based approaches (CUT&RUN/CUT&Tag), nuclei are bound to magnetic beads to streamline and automate sample processing. Next, adenosines nearby antibody-targeted chromatin features are methylated with pAG-M.EcoGII, which are then directly read from genomic DNA using Oxford Nanopore Technologies or Pacific Biosciences LRS platforms. To determine the capabilities and limitations of this assay, we tested multiple chromatin targets in various cell lines. Importantly, this method is highly reproducible across biological replicates, and highly concordant with orthogonal SRS assays (e.g., CUT&RUN). Further, we showed that this method is a true multi-omic approach by simultaneously profiling histone PTMs, native DNAme (5mC), and parental single-nucleotide variants from single DNA molecules within a single reaction. Finally, this workflow preserves chromatin integrity for LRS, revealing heterogeneity (e.g., haplotype or paternal origin) within/between data types and providing access to previously unmappable genomic regions (e.g., centromeres). Citation Format: Bryan J. Venters, Paul W. Hook, Vishnu S. Kumary, Alli R. Hickman, James T. Anderson, Anup Vaidya, Ryan J. Ezell, Jonathan M. Burg, Zu-Wen Sun, Martis W. Cowles, Winston Timp, Michael-Christopher Keogh. Multi-omic genomic mapping with long read sequencing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7026.
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Das, Akash Chandra, Aidin Foroutan, Brian Qian, Nader Hosseini Naghavi, Kayvan Shabani, and Parisa Shooshtari. "Single-Cell Chromatin Accessibility Data Combined with GWAS Improves Detection of Relevant Cell Types in 59 Complex Phenotypes." International Journal of Molecular Sciences 23, no. 19 (2022): 11456. http://dx.doi.org/10.3390/ijms231911456.
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Several disease risk variants reside on non-coding regions of DNA, particularly in open chromatin regions of specific cell types. Identifying the cell types relevant to complex traits through the integration of chromatin accessibility data and genome-wide association studies (GWAS) data can help to elucidate the mechanisms of these traits. In this study, we created a collection of associations between the combinations of chromatin accessibility data (bulk and single-cell) with an array of 201 complex phenotypes. We integrated the GWAS data of these 201 phenotypes with bulk chromatin accessibility data from 137 cell types measured by DNase-I hypersensitive sequencing and found significant results (FDR adjusted p-value ≤ 0.05) for at least one cell type in 21 complex phenotypes, such as atopic dermatitis, Graves’ disease, and body mass index. With the integration of single-cell chromatin accessibility data measured by an assay for transposase-accessible chromatin with high-throughput sequencing (scATAC-seq), taken from 111 adult and 111 fetal cell types, the resolution of association was magnified, enabling the identification of further cell types. This resulted in the identification of significant correlations (FDR adjusted p-value ≤ 0.05) between 15 categories of single-cell subtypes and 59 phenotypes ranging from autoimmune diseases like Graves’ disease to cardiovascular traits like diastolic/systolic blood pressure.
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Eagen, Kyle P., Erez Lieberman Aiden, and Roger D. Kornberg. "Polycomb-mediated chromatin loops revealed by a subkilobase-resolution chromatin interaction map." Proceedings of the National Academy of Sciences 114, no. 33 (2017): 8764–69. http://dx.doi.org/10.1073/pnas.1701291114.
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The locations of chromatin loops in Drosophila were determined by Hi-C (chemical cross-linking, restriction digestion, ligation, and high-throughput DNA sequencing). Whereas most loop boundaries or “anchors” are associated with CTCF protein in mammals, loop anchors in Drosophila were found most often in association with the polycomb group (PcG) protein Polycomb (Pc), a subunit of polycomb repressive complex 1 (PRC1). Loops were frequently located within domains of PcG-repressed chromatin. Promoters located at PRC1 loop anchors regulate some of the most important developmental genes and are less likely to be expressed than those not at PRC1 loop anchors. Although DNA looping has most commonly been associated with enhancer–promoter communication, our results indicate that loops are also associated with gene repression.
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Zhou, Jiawei, Junjing Wu, Tao Yang, et al. "Integration of ATAC-Seq and RNA-Seq Reveals VDR–SELENBP1 Axis Promotes Adipogenesis of Porcine Intramuscular Preadipocytes." International Journal of Molecular Sciences 25, no. 23 (2024): 12528. http://dx.doi.org/10.3390/ijms252312528.
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Intramuscular fat (IMF) content plays a crucial role in determining pork quality. Recent studies have highlighted transcriptional mechanisms controlling adipogenesis in porcine IMF. However, the changes in chromatin accessibility during adipogenic differentiation are still not well understood. In this study, we performed the assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) and transcriptome sequencing (RNA-Seq) analyses on porcine intramuscular preadipocytes to explore their adipogenic differentiation into mature adipocytes. We identified a total of 56,374 differentially accessible chromatin peaks and 4226 differentially expressed genes at day 0 and day 4 during adipogenic differentiation. A combined analysis of the ATAC-seq and RNA-seq data revealed that 1750 genes exhibited both differential chromatin accessibility and differential RNA expression during this process, including selenium-binding protein 1 (SELENBP1), PLIN1, ADIPOQ, and FASN. Furthermore, we found that vitamin D receptor (VDR) could bind to the promoter region of the SELENBP1 gene, activate SELENBP1 transcription, and ultimately promote lipid accumulation during adipogenic differentiation. This study provides a detailed overview of chromatin accessibility and gene expression changes during the adipogenic differentiation of porcine intramuscular preadipocytes. Moreover, we propose a novel regulatory mechanism involving the VDR–SELENBP1 signaling axis in adipogenic differentiation.
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Kubalová, Ivona, Amanda Souza Câmara, Petr Cápal, et al. "Helical coiling of metaphase chromatids." Nucleic Acids Research, March 2, 2023. http://dx.doi.org/10.1093/nar/gkad028.
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Abstract Chromatids of mitotic chromosomes were suggested to coil into a helix in early cytological studies and this assumption was recently supported by chromosome conformation capture (3C) sequencing. Still, direct differential visualization of a condensed chromatin fibre confirming the helical model was lacking. Here, we combined Hi-C analysis of purified metaphase chromosomes, biopolymer modelling and spatial structured illumination microscopy of large fluorescently labeled chromosome segments to reveal the chromonema - a helically-wound, 400 nm thick chromatin thread forming barley mitotic chromatids. Chromatin from adjacent turns of the helix intermingles due to the stochastic positioning of chromatin loops inside the chromonema. Helical turn size varies along chromosome length, correlating with chromatin density. Constraints on the observable dimensions of sister chromatid exchanges further supports the helical chromonema model.
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Ma, Kai, Kaili Yin, Jiong Li, et al. "The Hypothalamic Epigenetic Landscape in Dietary Obesity." Advanced Science, December 20, 2023. http://dx.doi.org/10.1002/advs.202306379.
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AbstractThe hypothalamus in the brain plays a pivotal role in controlling energy balance in vertebrates. Nutritional excess through high‐fat diet (HFD) feeding can dysregulate hypothalamic signaling at multiple levels. Yet, it remains largely unknown in what magnitude HFD feeding may impact epigenetics in this brain region. Here, it is shown that HFD feeding can significantly alter hypothalamic epigenetic events, including posttranslational histone modifications, DNA methylation, and chromatin accessibility. The authors comprehensively analyze the chromatin immunoprecipitation‐sequencing (ChIP‐seq), methylated DNA immunoprecipitation‐sequencing (MeDIP‐seq), single nucleus assay for transposase‐accessible chromatin using sequencing (snATAC‐seq), and RNA‐seq data of the hypothalamus of C57 BL/6 mice fed with a chow or HFD for 1 to 6 months. The chromatins are categorized into 6 states using the obtained ChIP‐seq data for H3K4me3, H3K27ac, H3K9me3, H3K27me3, and H3K36me3. A 1‐month HFD feeding dysregulates histone modifications and DNA methylation more pronouncedly than that of 3‐ or 6‐month. Besides, HFD feeding differentially impacts chromatin accessibility in hypothalamic cells. Thus, the epigenetic landscape is dysregulated in the hypothalamus of dietary obesity mice.
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Elise, van Bree, Haring Nina, and Jacobs Frank. "Snakemake pipeline for Chromatin Immunoprecipitation sequencing." March 1, 2019. https://doi.org/10.5281/zenodo.2581325.
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Subramanian, Vijayalakshmi V. "Preprint Highlight: Cohesin mediates DNA loop extrusion and sister chromatid cohesion by distinct mechanisms." Molecular Biology of the Cell 34, no. 5 (2023). http://dx.doi.org/10.1091/mbc.p23-03-0010.
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Cohesins organize chromatin within the nucleus by promoting loop extrusion via cis contacts on chromatin and sister chromatid cohesion via trans contacts between chromatids. Whether cohesins employ a distinct mechanism for these two functions is not known. The authors use a carefully designed conditional expression of cohesin, mutated at its hinge domain, and assess its function with a variety of in vitro and in vivo experiments including live TIRF imaging, loop extrusion assay, Hi-C, and calibrated ChIP sequencing. A mutation in the hinge domain separates the two functions of cohesin, as the mutant supports loop extrusion but not sister chromatid cohesion. A unique mechanism of cohesins in promoting loop extrusion has implications for their distinct role in genome organization and for transcriptional regulation and cell fate determination. This study will be of broad interest to genome biologists with respect to these phenomena. This preprint has been assigned the following badges: New Hypothesis, New Materials. Read the preprint on bioRxiv ( Nagasaka et al., 2022 ): https://doi.org/10.1101/2022.09.23.509019 .
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Ma, Shaoqian, and Yongyou Zhang. "Profiling chromatin regulatory landscape: insights into the development of ChIP-seq and ATAC-seq." Molecular Biomedicine 1, no. 1 (2020). http://dx.doi.org/10.1186/s43556-020-00009-w.
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Abstract Chromatin regulatory landscape plays a critical role in many disease processes and embryo development. Epigenome sequencing technologies such as chromatin immunoprecipitation sequencing (ChIP-seq) and assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) have enabled us to dissect the pan-genomic regulatory landscape of cells and tissues in both time and space dimensions by detecting specific chromatin state and its corresponding transcription factors. Pioneered by the advancement of chromatin immunoprecipitation-chip (ChIP-chip) technology, abundant epigenome profiling technologies have become available such as ChIP-seq, DNase I hypersensitive site sequencing (DNase-seq), ATAC-seq and so on. The advent of single-cell sequencing has revolutionized the next-generation sequencing, applications in single-cell epigenetics are enriched rapidly. Epigenome sequencing technologies have evolved from low-throughput to high-throughput and from bulk sample to the single-cell scope, which unprecedentedly benefits scientists to interpret life from different angles. In this review, after briefly introducing the background knowledge of epigenome biology, we discuss the development of epigenome sequencing technologies, especially ChIP-seq & ATAC-seq and their current applications in scientific research. Finally, we provide insights into future applications and challenges.
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Lee, Isac, Roham Razaghi, Timothy Gilpatrick, et al. "Methylation and accessibility profiling of GM12878, MCF-10A, MCF-7, and MDA-MB-231 using nanopore sequencing." August 5, 2020. https://doi.org/10.5281/zenodo.3969567.
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Probing epigenetic features on long molecules of DNA has tremendous potential to advance our understanding of the phased epigenome. In this study, we evaluate CpG methylation and chromatin accessibility simultaneously on long strands of DNA using GpC methyltransferase to exogenously label open chromatin, coupled with nanopore sequencing technology. We performed nanopore sequencing of Nucleosome Occupancy and Methylome (nanoNOMe) on four human cell lines (GM12878, MCF-10A, MCF-7, MDA-MB-231), and demonstrate the ability to directly measure methylation and chromatin accessibility in genomic features such as structural variations and repetitive elements. The long single-molecule resolution allows footprinting of protein and nucleosome binding and determining the combinatorial promoter epigenetic state on individual molecules. Long-read sequencing makes it possible to robustly assign reads to haplotypes, enabling allele-specific epigenetic analysis across the genome. We use existing SNV data on GM12878 to present the first fully phased human Probing epigenetic features on long molecules of DNA has tremendous potential to advance our understanding of the phased epigenome. We evaluate CpG methylation and chromatin accessibility simultaneously on long strands of DNA using GpC methyltransferase to exogenously label open chromatin, coupled with nanopore sequencing technology. We performed nanopore sequencing of Nucleosome Occupancy and Methylome (nanoNOMe) on four human cell lines (GM12878, MCF-10A, MCF-7, MDA-MB-231), and demonstrate the ability to directly measure methylation and chromatin accessibility in genomic features such as structural variations and repetitive elements. The long single-molecule resolution allows footprinting of protein and nucleosome binding and determining the combinatorial promoter epigenetic state on individual molecules. Long-read sequencing makes it possible to robustly assign reads to haplotypes, enabling allele-specific epigenetic analysis across the genome. We use existing SNV data on GM12878 to present the first fully phased human epigenome, consisting of chromosome-level allele-specific profiles of CpG methylation and chromatin accessibility.mosome-level allele-specific profiles of CpG methylation and chromatin accessibility.
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Ren, Zongna, Wanqing Zhao, Dandan Li, et al. "INO80-Dependent Remodeling of Transcriptional Regulatory Network Underlies the Progression of Heart Failure." Circulation, December 28, 2023. http://dx.doi.org/10.1161/circulationaha.123.065440.
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BACKGROUND: Progressive remodeling of cardiac gene expression underlies decline in cardiac function, eventually leading to heart failure. However, the major determinants of transcriptional network switching from normal to failed hearts remain to be determined. METHODS: In this study, we integrated human samples, genetic mouse models, and genomic approaches, including bulk RNA sequencing, single-cell RNA sequencing, chromatin immunoprecipitation followed by high-throughput sequencing, and assay for transposase-accessible chromatin with high-throughput sequencing, to identify the role of chromatin remodeling complex INO80 in heart homeostasis and dysfunction. RESULTS: The INO80 chromatin remodeling complex was abundantly expressed in mature cardiomyocytes, and its expression further increased in mouse and human heart failure. Cardiomyocyte-specific overexpression of Ino80 , its core catalytic subunit, induced heart failure within 4 days. Combining RNA sequencing, high-throughput sequencing, and assay for transposase-accessible chromatin with high-throughput sequencing, we revealed INO80 overexpression-dependent reshaping of the nucleosomal landscape that remodeled a core set of transcription factors, most notably the MEF2 family, whose target genes were closely associated with cardiac function. Conditional cardiomyocyte-specific deletion of Ino80 in an established mouse model of heart failure demonstrated remarkable preservation of cardiac function. CONCLUSIONS: In summary, our findings shed light on the INO80-dependent remodeling of the chromatin landscape and transcriptional networks as a major mechanism underlying cardiac dysfunction in heart failure, and suggest INO80 as a potential preventative or interventional target.
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Lee, Beoung Hun, Zexun Wu, and Suhn K. Rhie. "Characterizing chromatin interactions of regulatory elements and nucleosome positions, using Hi-C, Micro-C, and promoter capture Micro-C." Epigenetics & Chromatin 15, no. 1 (2022). http://dx.doi.org/10.1186/s13072-022-00473-4.
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Abstract Background Regulatory elements such as promoters, enhancers, and insulators interact each other to mediate molecular processes. To capture chromatin interactions of regulatory elements, 3C-derived methods such as Hi-C and Micro-C are developed. Here, we generated and analyzed Hi-C, Micro-C, and promoter capture Micro-C datasets with different sequencing depths to study chromatin interactions of regulatory elements and nucleosome positions in human prostate cancer cells. Results Compared to Hi-C, Micro-C identifies more high-resolution loops, including ones around structural variants. By evaluating the effect of sequencing depth, we revealed that more than 2 billion reads of Micro-C are needed to detect chromatin interactions at 1 kb resolution. Moreover, we found that deep-sequencing identifies additional long-range loops that are longer than 1 Mb in distance. Furthermore, we found that more than 50% of the loops are involved in insulators while less than 10% of the loops are promoter–enhancer loops. To comprehensively capture chromatin interactions that promoters are involved in, we performed promoter capture Micro-C. Promoter capture Micro-C identifies loops near promoters with a lower amount of sequencing reads. Sequencing of 160 million reads of promoter capture Micro-C resulted in reaching a plateau of identifying loops. However, there was still a subset of promoters that are not involved in loops even after deep-sequencing. By integrating Micro-C with NOMe-seq and ChIP-seq, we found that active promoters involved in loops have a more accessible region with lower levels of DNA methylation and more highly phased nucleosomes, compared to active promoters that are not involved in loops. Conclusion We determined the required sequencing depth for Micro-C and promoter capture Micro-C to generate high-resolution chromatin interaction maps and loops. We also investigated the effect of sequencing coverage of Hi-C, Micro-C, and promoter capture Micro-C on detecting chromatin loops. Our analyses suggest the presence of distinct regulatory element groups, which are differently involved in nucleosome positions and chromatin interactions. This study does not only provide valuable insights on understanding chromatin interactions of regulatory elements, but also present guidelines for designing research projects on chromatin interactions among regulatory elements.
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Holzmann, Johann, Antonio Z. Politi, Kota Nagasaka, et al. "Absolute quantification of cohesin, CTCF and their regulators in human cells." eLife 8 (June 17, 2019). http://dx.doi.org/10.7554/elife.46269.
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The organisation of mammalian genomes into loops and topologically associating domains (TADs) contributes to chromatin structure, gene expression and recombination. TADs and many loops are formed by cohesin and positioned by CTCF. In proliferating cells, cohesin also mediates sister chromatid cohesion, which is essential for chromosome segregation. Current models of chromatin folding and cohesion are based on assumptions of how many cohesin and CTCF molecules organise the genome. Here we have measured absolute copy numbers and dynamics of cohesin, CTCF, NIPBL, WAPL and sororin by mass spectrometry, fluorescence-correlation spectroscopy and fluorescence recovery after photobleaching in HeLa cells. In G1-phase, there are ~250,000 nuclear cohesin complexes, of which ~ 160,000 are chromatin-bound. Comparison with chromatin immunoprecipitation-sequencing data implies that some genomic cohesin and CTCF enrichment sites are unoccupied in single cells at any one time. We discuss the implications of these findings for how cohesin can contribute to genome organisation and cohesion.
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Bell, Jason C., David Jukam, Nicole A. Teran, et al. "Chromatin-associated RNA sequencing (ChAR-seq) maps genome-wide RNA-to-DNA contacts." eLife 7 (April 12, 2018). http://dx.doi.org/10.7554/elife.27024.
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RNA is a critical component of chromatin in eukaryotes, both as a product of transcription, and as an essential constituent of ribonucleoprotein complexes that regulate both local and global chromatin states. Here, we present a proximity ligation and sequencing method called Chromatin-Associated RNA sequencing (ChAR-seq) that maps all RNA-to-DNA contacts across the genome. Using Drosophila cells, we show that ChAR-seq provides unbiased, de novo identification of targets of chromatin-bound RNAs including nascent transcripts, chromosome-specific dosage compensation ncRNAs, and genome-wide trans-associated RNAs involved in co-transcriptional RNA processing.
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Li, Mengyao, Zhenhuan Jiang, Xueqiang Xu, et al. "Chromatin accessibility landscape of mouse early embryos revealed by single-cell NanoATAC-seq2." Science 387, no. 6741 (2025). https://doi.org/10.1126/science.adp4319.
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In mammals, fertilized eggs undergo genome-wide epigenetic reprogramming to generate the organism. However, our understanding of epigenetic dynamics during preimplantation development at single-cell resolution remains incomplete. Here, we developed scNanoATAC-seq2, a single-cell assay for transposase-accessible chromatin using long-read sequencing for scarce samples. We present a detailed chromatin accessibility landscape of mouse preimplantation development, revealing distinct chromatin signatures in the epiblast, primitive endoderm, and trophectoderm during lineage segregation. Differences between zygotes and two-cell embryos highlight reprogramming in chromatin accessibility during the maternal-to-zygotic transition. Single-cell long-read sequencing enables in-depth analysis of chromatin accessibility in noncanonical imprinting, imprinted X chromosome inactivation, and low-mappability genomic regions, such as repetitive elements and paralogs. Our data provide insights into chromatin dynamics during mammalian preimplantation development and lineage differentiation.
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Kumar, Banushree, Carmen Navarro, Philip Yuk Kwong Yung, et al. "Multiplexed chromatin immunoprecipitation sequencing for quantitative study of histone modifications and chromatin factors." Nature Protocols, October 3, 2024. http://dx.doi.org/10.1038/s41596-024-01058-z.
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Xi, Xiao-Li, Yi-Dong Yang, Hui-Ling Liu, Jie Jiang, and Bin Wu. "Chromatin accessibility module identified by single-cell sequencing underlies the diagnosis and prognosis of hepatocellular carcinoma." World Journal of Hepatology 17, no. 6 (2025). https://doi.org/10.4254/wjh.v17.i6.107329.
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BACKGROUND Hepatocellular carcinoma (HCC) is notorious for its aggressive progression and dismal prognosis, with chromatin accessibility dynamics emerging as pivotal yet poorly understood drivers. AIM To dissect how multilayered chromatin regulation sustains oncogenic transcription and tumor-stroma crosstalk in HCC, we combined multiomics single cell analysis. METHODS We integrated single-cell RNA sequencing and paired single-cell assay for transposase-accessible chromatin with sequencing data of HCC samples, complemented by bulk RNA sequencing validation across The Cancer Genome Atlas, Liver Cancer Institute, and GSE25907 cohorts. Cell type-specific chromatin architectures were resolved via ArchR, with regulatory hubs identified through peak-to-gene linkages and coaccessibility networks. Functional validation employed A485-mediated histone 3 lysine 27 acetylation suppression and small interfering RNA targeting DGAT1 . RESULTS Malignant hepatocytes exhibited expanded chromatin accessibility profiles, characterized by increased numbers of accessible peaks and larger physical regions despite reduced peak intensity. Enhancer-like peaks enriched in malignant regulation, forming long-range hubs. Eighteen enhancer-like peak-related genes showed tumor-specific overexpression and diagnostic accuracy, correlating with poor prognosis. Intercellular coaccessibility analysis revealed tumor-stroma symbiosis via shared chromatin states. Pharmacological histone 3 lysine 27 acetylation inhibition paradoxically downregulated DGAT1 , the hub gene most strongly regulated by chromatin accessibility. DGAT1 knockdown suppressed cell proliferation. CONCLUSION Multilayered chromatin reprogramming sustains HCC progression through tumor-stroma crosstalk and DGAT1 -related oncogenic transcription, defining targetable epigenetic vulnerabilities.
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Leduque, Basile, Alejandro Edera, Clémentine Vitte, and Leandro Quadrana. "Simultaneous profiling of chromatin accessibility and DNA methylation in complete plant genomes using long-read sequencing." Nucleic Acids Research, April 27, 2024. http://dx.doi.org/10.1093/nar/gkae306.
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Abstract Epigenetic regulations, including chromatin accessibility, nucleosome positioning and DNA methylation intricately shape genome function. However, current chromatin profiling techniques relying on short-read sequencing technologies fail to characterise highly repetitive genomic regions and cannot detect multiple chromatin features simultaneously. Here, we performed Simultaneous Accessibility and DNA Methylation Sequencing (SAM-seq) of purified plant nuclei. Thanks to the use of long-read nanopore sequencing, SAM-seq enables high-resolution profiling of m6A-tagged chromatin accessibility together with endogenous cytosine methylation in plants. Analysis of naked genomic DNA revealed significant sequence preference biases of m6A-MTases, controllable through a normalisation step. By applying SAM-seq to Arabidopsis and maize nuclei we obtained fine-grained accessibility and DNA methylation landscapes genome-wide. We uncovered crosstalk between chromatin accessibility and DNA methylation within nucleosomes of genes, TEs, and centromeric repeats. SAM-seq also detects DNA footprints over cis-regulatory regions. Furthermore, using the single-molecule information provided by SAM-seq we identified extensive cellular heterogeneity at chromatin domains with antagonistic chromatin marks, suggesting that bivalency reflects cell-specific regulations. SAM-seq is a powerful approach to simultaneously study multiple epigenetic features over unique and repetitive sequences, opening new opportunities for the investigation of epigenetic mechanisms.
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Zhang, Haowen, Li Song, Xiaotao Wang, et al. "Fast alignment and preprocessing of chromatin profiles with Chromap." Nature Communications 12, no. 1 (2021). http://dx.doi.org/10.1038/s41467-021-26865-w.
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AbstractAs sequencing depth of chromatin studies continually grows deeper for sensitive profiling of regulatory elements or chromatin spatial structures, aligning and preprocessing of these sequencing data have become the bottleneck for analysis. Here we present Chromap, an ultrafast method for aligning and preprocessing high throughput chromatin profiles. Chromap is comparable to BWA-MEM and Bowtie2 in alignment accuracy and is over 10 times faster than traditional workflows on bulk ChIP-seq/Hi-C profiles and than 10x Genomics’ CellRanger v2.0.0 pipeline on single-cell ATAC-seq profiles.
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Nakamura, Kosuke, Kosuke Nakamura, Keji Zhao, et al. "A single-cell chromatin immunocleavage sequencing (scChIC-seq)." Protocol Exchange, February 5, 2019. http://dx.doi.org/10.1038/protex.2019.011.
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Fullwood, Melissa J., Yuyuan Han, Chia‐Lin Wei, Xiaoan Ruan, and Yijun Ruan. "Chromatin Interaction Analysis Using Paired‐End Tag Sequencing." Current Protocols in Molecular Biology 89, no. 1 (2010). http://dx.doi.org/10.1002/0471142727.mb2115s89.
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Long, Keren, Duo Su, Xiaokai Li, et al. "Identification of enhancers responsible for the coordinated expression of myosin heavy chain isoforms in skeletal muscle." BMC Genomics 23, no. 1 (2022). http://dx.doi.org/10.1186/s12864-022-08737-9.
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Abstract Background Skeletal muscles consist of fibers of differing contractility and metabolic properties, which are primarily determined by the content of myosin heavy chain (MYH) isoforms (MYH7, MYH2, MYH1, and MYH4). The regulation of Myh genes transcription depends on three-dimensional chromatin conformation interaction, but the mechanistic details remain to be determined. Results In this study, we characterized the interaction profiles of Myh genes using 4C-seq (circular chromosome conformation capture coupled to high-throughput sequencing). The interaction profile of Myh genes changed between fast quadriceps and slow soleus muscles. Combining chromatin immunoprecipitation-sequencing (ChIP-seq) and transposase accessible chromatin with high-throughput sequencing (ATAC-seq), we found that a 38 kb intergenic region interacting simultaneously with fast Myh genes promoters controlled the coordinated expression of fast Myh genes. We also identified four active enhancers of Myh7, and revealed that binding of MYOG and MYOD increased the activity of Myh7 enhancers. Conclusions This study provides new insight into the chromatin interactions that regulate Myh genes expression.
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Zhang, Qiqi, Fengli Zhao, Zhe Wu, and Danling Zhu. "A simple and robust method for isolating and analyzing chromatin-bound RNAs in Arabidopsis." Plant Methods 18, no. 1 (2022). http://dx.doi.org/10.1186/s13007-022-00967-y.
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Abstract Background Chromatin-bound RNAs are the primary product of transcription that undergo on-chromatin processing such as capping, splicing, and polyadenylation. These processing steps then determine the fate of RNAs. Albeit its vital importance, a simple and robust method for isolating different fractions of chromatin-bound RNAs is missing in plants. Result Here, we describe our updated method and the associated step-by-step protocol for chromatin-bound RNAs isolation in A. thaliana. The chromatin-bound RNAs isolation is based on the 1 M UREA wash that removes the majority of non-chromatin-associated proteins from the nucleus, as previously developed in mammalian cells. On-demand, the isolated chromatin-bound RNAs can be either used directly for gene-specific analysis or subject to further rRNA removal and also the optional polyadenylated RNA removal, followed by high-throughput sequencing. Detailed protocols for these procedures are also provided. Comparison of sequencing results of chromatin-bound RNAs with and without polyadenylated RNA removal revealed that a small fraction of CB-RNAs is polyadenylated but not yet fully spliced, representing RNA-processing intermediate on-chromatin. Conclusion This optimized chromatin-bound RNAs purification method is simple and robust and can be used to study transcription and its-coupled RNA processing in plants.
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Shin, Hyun Mu, Gwanghun Kim, Sangjib Kim, et al. "Chromatin accessibility of circulating CD8+ T cells predicts treatment response to PD-1 blockade in patients with gastric cancer." Nature Communications 12, no. 1 (2021). http://dx.doi.org/10.1038/s41467-021-21299-w.
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AbstractAlthough tumor genomic profiling has identified small subsets of gastric cancer (GC) patients with clinical benefit from anti-PD-1 treatment, not all responses can be explained by tumor sequencing alone. We investigate epigenetic elements responsible for the differential response to anti-PD-1 therapy by quantitatively assessing the genome-wide chromatin accessibility of circulating CD8+ T cells in patients’ peripheral blood. Using an assay for transposase-accessible chromatin using sequencing (ATAC-seq), we identify unique open regions of chromatin that significantly distinguish anti-PD-1 therapy responders from non-responders. GC patients with high chromatin openness of circulating CD8+ T cells are significantly enriched in the responder group. Concordantly, patients with high chromatin openness at specific genomic positions of their circulating CD8+ T cells demonstrate significantly better survival than those with closed chromatin. Here we reveal that epigenetic characteristics of baseline CD8+ T cells can be used to identify metastatic GC patients who may benefit from anti-PD-1 therapy.