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05 Jun 2023 at 01:46
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Bibliography on: Topologically Associating Domains


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RJR: Recommended Bibliography 05 Jun 2023 at 01:46 Created: 

Topologically Associating Domains

"Recent studies have shown that chromosomes in a range of organisms are compartmentalized in different types of chromatin domains. In mammals, chromosomes form compartments that are composed of smaller Topologically Associating Domains (TADs). TADs are thought to represent functional domains of gene regulation but much is still unknown about the mechanisms of their formation and how they exert their regulatory effect on embedded genes. Further, similar domains have been detected in other organisms, including flies, worms, fungi and bacteria. Although in all these cases these domains appear similar as detected by 3C-based methods, their biology appears to be quite distinct with differences in the protein complexes involved in their formation and differences in their internal organization." QUOTE FROM: Dekker Job and Heard Edith (2015), Structural and functional diversity of Topologically Associating Domains, FEBS Letters, 589, doi: 10.1016/j.febslet.2015.08.044

Created with PubMed® Query: ( "Topologically Associating Domains" OR "Topologically Associating Domain" ) NOT pmcbook NOT ispreviousversion

Citations The Papers (from PubMed®)


RevDate: 2023-06-01

Gu J, Li S, Zhu B, et al (2023)

Genetic variation and domestication of horses revealed by 10 chromosome-level genomes and whole-genome resequencing.

Molecular ecology resources [Epub ahead of print].

Understanding the genetic variations of the horse (Equus caballus) genome will improve breeding conservation and welfare. However, genetic variations in long segments, such as structural variants (SVs), remain understudied. We de novo assembled 10 chromosome-level three-dimensional horse genomes, each representing a distinct breed, and analysed horse SVs using a multi-assembly approach. Our findings suggest that SVs with the accumulation of mammalian-wide interspersed repeats related to long interspersed nuclear elements might be a horse-specific mechanism to modulate genome-wide gene regulatory networks. We found that olfactory receptors were commonly loss and accumulated deleterious mutations, but no purge of deleterious mutations occurred during horse domestication. We examined the potential effects of SVs on the spatial structure of chromatin via topologically associating domains (TADs). Breed-specific TADs were significantly enriched by breed-specific SVs. We identified 4199 unique breakpoint-resolved novel insertions across all chromosomes that account for 2.84 Mb sequences missing from the reference genome. Several novel insertions might have potential functional consequences, as 519 appeared to reside within 449 gene bodies. These genes are primarily involved in pathogen recognition, innate immune responses and drug metabolism. Moreover, 37 diverse horses were resequenced. Combining this with public data, we analysed 97 horses through a comparative population genomics approach to identify the genetic basis underlying breed characteristics using Thoroughbreds as a case study. We provide new scientific evidence for horse domestication, an understanding of the genetic mechanism underlying the phenotypic evolution of horses, and a comprehensive genetic variation resource for further genetic studies of horses.

RevDate: 2023-05-31

Li D, Zhao XY, Zhou S, et al (2023)

Multidimensional profiling reveals GATA1-modulated stage-specific chromatin states and functional associations during human erythropoiesis.

Nucleic acids research pii:7186996 [Epub ahead of print].

Mammalian erythroid development can be divided into three stages: hematopoietic stem and progenitor cell (HSPC), erythroid progenitor (Ery-Pro), and erythroid precursor (Ery-Pre). However, the mechanisms by which the 3D genome changes to establish the stage-specific transcription programs that are critical for erythropoiesis remain unclear. Here, we analyze the chromatin landscape at multiple levels in defined populations from primary human erythroid culture. While compartments and topologically associating domains remain largely unchanged, ∼50% of H3K27Ac-marked enhancers are dynamic in HSPC versus Ery-Pre. The enhancer anchors of enhancer-promoter loops are enriched for occupancy of respective stage-specific transcription factors (TFs), indicating these TFs orchestrate the enhancer connectome rewiring. The master TF of erythropoiesis, GATA1, is found to occupy most erythroid gene promoters at the Ery-Pro stage, and mediate conspicuous local rewiring through acquiring binding at the distal regions in Ery-Pre, promoting productive erythroid transcription output. Knocking out GATA1 binding sites precisely abrogates local rewiring and corresponding gene expression. Interestingly, knocking down GATA1 can transiently revert the cell state to an earlier stage and prolong the window of progenitor state. This study reveals mechanistic insights underlying chromatin rearrangements during development by integrating multidimensional chromatin landscape analyses to associate with transcription output and cellular states.

RevDate: 2023-05-30

Tolokh IS, Kinney NA, Sharakhov IV, et al (2023)

Strong interactions between highly dynamic lamina-associated domains and the nuclear envelope stabilize the 3D architecture of Drosophila interphase chromatin.

Epigenetics & chromatin, 16(1):21.

BACKGROUND: Interactions among topologically associating domains (TADs), and between the nuclear envelope (NE) and lamina-associated domains (LADs) are expected to shape various aspects of three-dimensional (3D) chromatin structure and dynamics; however, relevant genome-wide experiments that may provide statistically significant conclusions remain difficult.

RESULTS: We have developed a coarse-grained dynamical model of D. melanogaster nuclei at TAD resolution that explicitly accounts for four distinct epigenetic classes of TADs and LAD-NE interactions. The model is parameterized to reproduce the experimental Hi-C map of the wild type (WT) nuclei; it describes time evolution of the chromatin over the G1 phase of the interphase. The simulations include an ensemble of nuclei, corresponding to the experimentally observed set of several possible mutual arrangements of chromosomal arms. The model is validated against multiple structural features of chromatin from several different experiments not used in model development. Predicted positioning of all LADs at the NE is highly dynamic-the same LAD can attach, detach and move far away from the NE multiple times during interphase. The probabilities of LADs to be in contact with the NE vary by an order of magnitude, despite all having the same affinity to the NE in the model. These probabilities are mostly determined by a highly variable local linear density of LADs along the genome, which also has the same strong effect on the predicted positioning of individual TADs -- higher probability of a TAD to be near NE is largely determined by a higher linear density of LADs surrounding this TAD. The distribution of LADs along the chromosome chains plays a notable role in maintaining a non-random average global structure of chromatin. Relatively high affinity of LADs to the NE in the WT nuclei substantially reduces sensitivity of the global radial chromatin distribution to variations in the strength of TAD-TAD interactions compared to the lamin depleted nuclei, where a small (0.5 kT) increase of cross-type TAD-TAD interactions doubles the chromatin density in the central nucleus region.

CONCLUSIONS: A dynamical model of the entire fruit fly genome makes multiple genome-wide predictions of biological interest. The distribution of LADs along the chromatin chains affects their probabilities to be in contact with the NE and radial positioning of highly mobile TADs, playing a notable role in creating a non-random average global structure of the chromatin. We conjecture that an important role of attractive LAD-NE interactions is to stabilize global chromatin structure against inevitable cell-to-cell variations in TAD-TAD interactions.

RevDate: 2023-05-30

Hamba Y, Kamatani T, Miya F, et al (2023)

Topologically associating domain underlies tissue specific expression of long intergenic non-coding RNAs.

iScience, 26(5):106640.

Accumulating evidence indicates that long intergenic non-coding RNAs (lincRNAs) show more tissue-specific expression patterns than protein-coding genes (PCGs). However, although lincRNAs are subject to canonical transcriptional regulation like PCGs, the molecular basis for the specificity of their expression patterns remains unclear. Here, using expression data and coordinates of topologically associating domains (TADs) in human tissues, we show that lincRNA loci are significantly enriched in the more internal region of TADs compared to PCGs and that lincRNAs within TADs have higher tissue specificity than those outside TADs. Based on these, we propose an analytical framework to interpret transcriptional status using lincRNA as an indicator. We applied it to hypertrophic cardiomyopathy data and found disease-specific transcriptional regulation: ectopic expression of keratin at the TAD level and derepression of myocyte differentiation-related genes by E2F1 with down-regulation of LINC00881. Our results provide understanding of the function and regulation of lincRNAs according to genomic structure.

RevDate: 2023-05-16

Zhang X, Yu G, Dai Y, et al (2023)

High-resolution Hi-C maps highlight multiscale chromatin architecture reorganization during cold stress in Brachypodium distachyon.

BMC plant biology, 23(1):260.

BACKGROUND: The adaptation of plants to cold stress involves changes in gene expression profiles that are associated with epigenetic regulation. Although the three-dimensional (3D) genome architecture is considered an important epigenetic regulator, the role of 3D genome organization in the cold stress response remains unclear.

RESULTS: In this study, we developed high-resolution 3D genomic maps using control and cold-treated leaf tissue of the model plant Brachypodium distachyon using Hi-C to determine how cold stress affects the 3D genome architecture. We generated ~ 1.5 kb resolution chromatin interaction maps and showed that cold stress disrupts different levels of chromosome organization, including A/B compartment transition, a reduction in chromatin compartmentalization and the size of topologically associating domains (TADs), and loss of long-range chromatin loops. Integrating RNA-seq information, we identified cold-response genes and revealed that transcription was largely unaffected by the A/B compartment transition. The cold-response genes were predominantly localized in compartment A. In contrast, transcriptional changes are required for TAD reorganization. We demonstrated that dynamic TAD events were associated with H3K27me3 and H3K27ac state alterations. Moreover, a loss of chromatin looping, rather than a gain of looping, coincides with alterations in gene expression, indicating that chromatin loop disruption may play a more important role than loop formation in the cold-stress response.

CONCLUSIONS: Our study highlights the multiscale 3D genome reprogramming that occurs during cold stress and expands our knowledge of the mechanisms underlying transcriptional regulation in response to cold stress in plants.

RevDate: 2023-05-10

Parodi L, Comeau ME, Georgakis MK, et al (2023)

Deep resequencing of the 1q22 locus in non-lobar intracerebral hemorrhage.

medRxiv : the preprint server for health sciences pii:2023.04.18.23288754.

OBJECTIVE: Genome-wide association studies have identified 1q22 as a susceptibility locus for cerebral small vessel diseases (CSVDs), including non-lobar intracerebral hemorrhage (ICH) and lacunar stroke. In the present study we performed targeted high-depth sequencing of 1q22 in ICH cases and controls to further characterize this locus and prioritize potential causal mechanisms, which remain unknown.

METHODS: 95,000 base pairs spanning 1q22 , including SEMA4A, SLC25A44 and PMF1 / PMF1-BGLAP were sequenced in 1,055 spontaneous ICH cases (534 lobar and 521 non-lobar) and 1,078 controls. Firth regression and RIFT analysis were used to analyze common and rare variants, respectively. Chromatin interaction analyses were performed using Hi-C, ChIP-Seq and ChIA-PET databases. Multivariable Mendelian randomization (MVMR) assessed whether alterations in gene-specific expression relative to regionally co-expressed genes at 1q22 could be causally related to ICH risk.

RESULTS: Common and rare variant analyses prioritized variants in SEMA4A 5'-UTR and PMF1 intronic regions, overlapping with active promoter and enhancer regions based on ENCODE annotation. Hi-C data analysis determined that 1q22 is spatially organized in a single chromatin loop and that the genes therein belong to the same Topologically Associating Domain. ChIP-Seq and ChIA-PET data analysis highlighted the presence of long-range interactions between the SEMA4A -promoter and PMF1 -enhancer regions prioritized by association testing. MVMR analyses demonstrated that PMF1 overexpression could be causally related to non-lobar ICH risk.

INTERPRETATION: Altered promoter-enhancer interactions leading to PMF1 overexpression, potentially dysregulating polyamine catabolism, could explain demonstrated associations with non-lobar ICH risk at 1q22 , offering a potential new target for prevention of ICH and CSVD.

RevDate: 2023-05-10

Li A, Zeng G, Wang H, et al (2023)

DeDoc2 Identifies and Characterizes the Hierarchy and Dynamics of Chromatin TAD-Like Domains in the Single Cells.

Advanced science (Weinheim, Baden-Wurttemberg, Germany) [Epub ahead of print].

Topologically associating domains (TADs) are functional chromatin units with hierarchical structure. However, the existence, prevalence, and dynamics of such hierarchy in single cells remain unexplored. Here, a new generation TAD-like domain (TLD) detection algorithm, named deDoc2, to decode the hierarchy of TLDs in single cells, is reported. With dynamic programming, deDoc2 seeks genome partitions with global minimal structure entropy for both whole and local contact matrix. Notably, deDoc2 outperforms state-of-the-art tools and is one of only two tools able to identify the hierarchy of TLDs in single cells. By applying deDoc2, it is showed that the hierarchy of TLDs in single cells is highly dynamic during cell cycle, as well as among human brain cortex cells, and that it is associated with cellular identity and functions. Thus, the results demonstrate the abundance of information potentially encoded by TLD hierarchy for functional regulation. The deDoc2 can be freely accessed at https://github.com/zengguangjie/deDoc2.

RevDate: 2023-05-04

Liu J, Li P, Sun J, et al (2023)

LPAD: using network construction and label propagation to detect topologically associating domains from Hi-C data.

Briefings in bioinformatics pii:7150739 [Epub ahead of print].

With the development of chromosome conformation capture technique, the study of spatial conformation of a genome based on Hi-C technique has made a quantum leap. Previous studies reveal that genomes are folded into hierarchy of three-dimensional (3D) structures associated with topologically associating domains (TADs), and detecting TAD boundaries is of great significance in the chromosome-level analysis of 3D genome architecture. In this paper, we propose a novel TAD identification method, LPAD, which first extracts node correlations from global interactions of chromosomes based on the random walk with restart and then builds an undirected graph from Hi-C contact matrix. Next, LPAD designs a label propagation-based approach to discover communities and generates TADs. Experimental results verify the effectiveness and quality of TAD detections compared with existing methods. Furthermore, experimental evaluation of chromatin immunoprecipitation sequencing data shows that LPAD performs high enrichment of histone modifications remarkably nearby the TAD boundaries, and these results demonstrate LPAD's advantages on TAD identification accuracy.

RevDate: 2023-04-28

Liu P, Li D, Zhang J, et al (2023)

Comparative three-dimensional genome architectures of adipose tissues provide insight into human-specific regulation of metabolic homeostasis.

The Journal of biological chemistry pii:S0021-9258(23)01785-4 [Epub ahead of print].

Elucidating the regulatory mechanisms of human adipose tissues (ATs) evolution is essential for understanding human-specific metabolic regulation, but the functional importance and evolutionary dynamics of three-dimensional (3D) genome organizations of ATs are not well defined. Here, we compared the 3D genome architectures of anatomically distinct ATs from humans and six representative mammalian models. We recognized evolutionarily conserved and human-specific chromatin conformation in ATs at multiple scales, including compartmentalization, topologically associating domain (TAD), and promoter-enhancer interactions (PEI), which have not been described previously. We found promoter-enhancer interaction (PEI) are much more evolutionarily dynamic with respect to compartmentalization and topologically associating domain (TAD). Compared to conserved PEIs, human-specific PEIs are enriched for human-specific sequence and the binding motifs of their potential mediators (transcription factors) are less conserved. Our data also demonstrated that genes involved in the evolutionarily dynamics of chromatin organization have weaker transcriptional conservation than those associated with conserved chromatin organization. Furthermore, the genes involved in energy metabolism and the maintenance of metabolic homeostasis are enriched in human-specific chromatin organization, while housekeeping genes, health-related genes and genetic variations are enriched in evolutionarily conserved compared to human-specific chromatin organization. Finally, we showed extensively divergent human-specific 3D genome organizations among one subcutaneous and three visceral ATs. Together, these findings provide a global overview of 3D genome architecture dynamics between ATs from human and mammalian models, and new insights into understanding the regulatory evolution of human ATs.

RevDate: 2023-04-27

Keough KC, Whalen S, Inoue F, et al (2023)

Three-dimensional genome rewiring in loci with human accelerated regions.

Science (New York, N.Y.), 380(6643):eabm1696.

Human accelerated regions (HARs) are conserved genomic loci that evolved at an accelerated rate in the human lineage and may underlie human-specific traits. We generated HARs and chimpanzee accelerated regions with an automated pipeline and an alignment of 241 mammalian genomes. Combining deep learning with chromatin capture experiments in human and chimpanzee neural progenitor cells, we discovered a significant enrichment of HARs in topologically associating domains containing human-specific genomic variants that change three-dimensional (3D) genome organization. Differential gene expression between humans and chimpanzees at these loci suggests rewiring of regulatory interactions between HARs and neurodevelopmental genes. Thus, comparative genomics together with models of 3D genome folding revealed enhancer hijacking as an explanation for the rapid evolution of HARs.

RevDate: 2023-04-26

Zhang H, GA Blobel (2023)

Genome folding dynamics during the M-to-G1-phase transition.

Current opinion in genetics & development, 80:102036 pii:S0959-437X(23)00016-3 [Epub ahead of print].

All measurable features of higher-order chromosomal architecture undergo drastic reorganization as cells enter and exit mitosis. During mitosis, gene transcription is temporarily halted, the nuclear envelope is dismantled, and chromosomes undergo condensation. At this time, chromatin compartments, topologically associating domains (TADs), and loops that connect enhancers with promoters as well as CTCF/cohesin loops are dissolved. Upon G1 entry, genome organization is rebuilt in the daughter nuclei to resemble that of the mother nucleus. We survey recent studies that traced these features in relation to gene expression during the mitosis-to-G1-phase transition at high temporal resolution. Dissection of fluctuating architectural features informed the hierarchical relationships of chromosomal organization, the mechanisms by which they are formed, and their mutual (in-) dependence. These studies highlight the importance of considering the cell cycle dynamics for studies of chromosomal organization.

RevDate: 2023-04-21

Okonechnikov K, Camgöz A, Chapman O, et al (2023)

3D genome mapping identifies subgroup-specific chromosome conformations and tumor-dependency genes in ependymoma.

Nature communications, 14(1):2300.

Ependymoma is a tumor of the brain or spinal cord. The two most common and aggressive molecular groups of ependymoma are the supratentorial ZFTA-fusion associated and the posterior fossa ependymoma group A. In both groups, tumors occur mainly in young children and frequently recur after treatment. Although molecular mechanisms underlying these diseases have recently been uncovered, they remain difficult to target and innovative therapeutic approaches are urgently needed. Here, we use genome-wide chromosome conformation capture (Hi-C), complemented with CTCF and H3K27ac ChIP-seq, as well as gene expression and DNA methylation analysis in primary and relapsed ependymoma tumors, to identify chromosomal conformations and regulatory mechanisms associated with aberrant gene expression. In particular, we observe the formation of new topologically associating domains ('neo-TADs') caused by structural variants, group-specific 3D chromatin loops, and the replacement of CTCF insulators by DNA hyper-methylation. Through inhibition experiments, we validate that genes implicated by these 3D genome conformations are essential for the survival of patient-derived ependymoma models in a group-specific manner. Thus, this study extends our ability to reveal tumor-dependency genes by 3D genome conformations even in tumors that lack targetable genetic alterations.

RevDate: 2023-04-21

Wang B, Liu K, Li Y, et al (2023)

DFHiC: A dilated full convolution model to enhance the resolution of Hi-C data.

Bioinformatics (Oxford, England) pii:7135829 [Epub ahead of print].

MOTIVATION: Hi-C technology has been the most widely used chromosome conformation capture(3C) experiment that measures the frequency of all paired interactions in the entire genome, which is a powerful tool for studying the 3D structure of the genome. The fineness of the constructed genome structure depends on the resolution of Hi-C data. However, due to the fact that high-resolution Hi-C data require deep sequencing and thus high experimental cost, most available Hi-C data are in low-resolution. Hence, it is essential to enhance the quality of Hi-C data by developing the effective computational methods.

RESULTS: In this work, we propose a novel method, so-called DFHiC, which generates the high-resolution Hi-C matrix from the low-resolution Hi-C matrix in the framework of the dilated convolutional neural network. The dilated convolution is able to effectively explore the global patterns in the overall Hi-C matrix by taking advantage of the information of the Hi-C matrix in a way of the longer genomic distance. Consequently, DFHiC can improve the resolution of the Hi-C matrix reliably and accurately. More importantly, the super-resolution Hi-C data enhanced by DFHiC is more in line with the real high-resolution Hi-C data than those done by the other existing methods, in terms of both chromatin significant interactions and identifying topologically associating domains (TADs).

AVAILABILITY: https://github.com/BinWangCSU/DFHiC.

SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

RevDate: 2023-04-20

Rajderkar S, Barozzi I, Zhu Y, et al (2023)

Topologically associating domain boundaries are required for normal genome function.

Communications biology, 6(1):435.

Topologically associating domain (TAD) boundaries partition the genome into distinct regulatory territories. Anecdotal evidence suggests that their disruption may interfere with normal gene expression and cause disease phenotypes[1-3], but the overall extent to which this occurs remains unknown. Here we demonstrate that targeted deletions of TAD boundaries cause a range of disruptions to normal in vivo genome function and organismal development. We used CRISPR genome editing in mice to individually delete eight TAD boundaries (11-80 kb in size) from the genome. All deletions examined resulted in detectable molecular or organismal phenotypes, which included altered chromatin interactions or gene expression, reduced viability, and anatomical phenotypes. We observed changes in local 3D chromatin architecture in 7 of 8 (88%) cases, including the merging of TADs and altered contact frequencies within TADs adjacent to the deleted boundary. For 5 of 8 (63%) loci examined, boundary deletions were associated with increased embryonic lethality or other developmental phenotypes. For example, a TAD boundary deletion near Smad3/Smad6 caused complete embryonic lethality, while a deletion near Tbx5/Lhx5 resulted in a severe lung malformation. Our findings demonstrate the importance of TAD boundary sequences for in vivo genome function and reinforce the critical need to carefully consider the potential pathogenicity of noncoding deletions affecting TAD boundaries in clinical genetics screening.

RevDate: 2023-04-19

Davidson IF, Barth R, Zaczek M, et al (2023)

CTCF is a DNA-tension-dependent barrier to cohesin-mediated loop extrusion.

Nature [Epub ahead of print].

In eukaryotes, genomic DNA is extruded into loops by cohesin[1]. By restraining this process, the DNA-binding protein CCCTC-binding factor (CTCF) generates topologically associating domains (TADs)[2,3] that have important roles in gene regulation and recombination during development and disease[1,4-7]. How CTCF establishes TAD boundaries and to what extent these are permeable to cohesin is unclear[8]. Here, to address these questions, we visualize interactions of single CTCF and cohesin molecules on DNA in vitro. We show that CTCF is sufficient to block diffusing cohesin, possibly reflecting how cohesive cohesin accumulates at TAD boundaries, and is also sufficient to block loop-extruding cohesin, reflecting how CTCF establishes TAD boundaries. CTCF functions asymmetrically, as predicted; however, CTCF is dependent on DNA tension. Moreover, CTCF regulates cohesin's loop-extrusion activity by changing its direction and by inducing loop shrinkage. Our data indicate that CTCF is not, as previously assumed, simply a barrier to cohesin-mediated loop extrusion but is an active regulator of this process, whereby the permeability of TAD boundaries can be modulated by DNA tension. These results reveal mechanistic principles of how CTCF controls loop extrusion and genome architecture.

RevDate: 2023-04-18

Yin X, Romero-Campero FJ, Yang M, et al (2023)

Binding by the Polycomb complex component BMI1 and H2A monoubiquitination shape local and long-range interactions in the Arabidopsis genome.

The Plant cell pii:7127682 [Epub ahead of print].

Three-dimensional (3D) chromatin organization is highly dynamic during development and seems to play a crucial role in regulating gene expression. Self-interacting domains, commonly called topologically associating domains (TADs) or compartment domains (CDs), have been proposed as the basic structural units of chromatin organization. Surprisingly, although these units have been found in several plant species, they escaped detection in Arabidopsis (Arabidopsis thaliana). Here, we show that the Arabidopsis genome is partitioned into contiguous CDs with different epigenetic features, which are required to maintain appropriate intra-CD and long-range interactions. Consistent with this notion, the histone-modifying Polycomb group machinery is involved in 3D chromatin organization. Yet, while it is clear that Polycomb Repressive Complex 2 (PRC2)-mediated trimethylation of histone H3 on lysine 27 (H3K27me3) helps establish local and long-range chromatin interactions in plants, the implications of PRC1-mediated histone H2A monoubiquitination on lysine 121 (H2AK121ub) are unclear. We found that PRC1, together with PRC2, maintains intra-CD interactions, but it also hinders the formation of H3K4me3-enriched local chromatin loops when acting independently of PRC2. Moreover, the loss of PRC1 or PRC2 activity differentially affects long-range chromatin interactions, and these 3D changes differentially affect gene expression. Our results suggest that H2AK121ub helps prevent the formation of transposable element/H3K27me1-rich long loops and serves as a docking point for H3K27me3 incorporation.

RevDate: 2023-04-17

Guo M, Yao Z, Jiang C, et al (2023)

Three-dimensional and single-cell sequencing of liver cancer reveals comprehensive host-virus interactions in HBV infection.

Frontiers in immunology, 14:1161522.

BACKGROUNDS: Hepatitis B virus (HBV) infection is a major risk factor for chronic liver diseases and liver cancer (mainly hepatocellular carcinoma, HCC), while the underlying mechanisms and host-virus interactions are still largely elusive.

METHODS: We applied HiC sequencing to HepG2 (HBV-) and HepG2-2.2.15 (HBV+) cell lines and combined them with public HCC single-cell RNA-seq data, HCC bulk RNA-seq data, and both genomic and epigenomic ChIP-seq data to reveal potential disease mechanisms of HBV infection and host-virus interactions reflected by 3D genome organization.

RESULTS: We found that HBV enhanced overall proximal chromatin interactions (CIs) of liver cells and primarily affected regional CIs on chromosomes 13, 14, 17, and 22. Interestingly, HBV altered the boundaries of many topologically associating domains (TADs), and genes nearby these boundaries showed functional enrichment in cell adhesion which may promote cancer metastasis. Moreover, A/B compartment analysis revealed dramatic changes on chromosomes 9, 13 and 21, with more B compartments (inactive or closed) shifting to A compartments (active or open). The A-to-B regions (closing) harbored enhancers enriched in the regulation of inflammatory responses, whereas B-to-A regions (opening) were enriched for transposable elements (TE). Furthermore, we identified large HBV-induced structural variations (SVs) that disrupted tumor suppressors, NLGN4Y and PROS1. Finally, we examined differentially expressed genes and TEs in single hepatocytes with or without HBV infection, by using single-cell RNA-seq data. Consistent with our HiC sequencing findings, two upregulated genes that promote HBV replication, HNF4A and NR5A2, were located in regions with HBV-enhanced CIs, and five TEs were located in HBV-activated regions. Therefore, HBV may promote liver diseases by affecting the human 3D genome structure.

CONCLUSION: Our work promotes mechanistic understanding of HBV infection and host-virus interactions related to liver diseases that affect billions of people worldwide. Our findings may also have implications for novel immunotherapeutic strategies targeting HBV infection.

RevDate: 2023-04-13

Cheng J, Cao X, Wang X, et al (2023)

Dynamic chromatin architectures provide insights into the genetics of cattle myogenesis.

Journal of animal science and biotechnology, 14(1):59.

BACKGROUND: Sharply increased beef consumption is propelling the genetic improvement projects of beef cattle in China. Three-dimensional genome structure is confirmed to be an important layer of transcription regulation. Although genome-wide interaction data of several livestock species have already been produced, the genome structure states and its regulatory rules in cattle muscle are still limited.

RESULTS: Here we present the first 3D genome data in Longissimus dorsi muscle of fetal and adult cattle (Bos taurus). We showed that compartments, topologically associating domains (TADs), and loop undergo re-organization and the structure dynamics were consistent with transcriptomic divergence during muscle development. Furthermore, we annotated cis-regulatory elements in cattle genome during myogenesis and demonstrated the enrichments of promoter and enhancer in selection sweeps. We further validated the regulatory function of one HMGA2 intronic enhancer near a strong sweep region on primary bovine myoblast proliferation.

CONCLUSIONS: Our data provide key insights of the regulatory function of high order chromatin structure and cattle myogenic biology, which will benefit the progress of genetic improvement of beef cattle.

RevDate: 2023-04-13

Mao A, Chen C, Portillo-Ledesma S, et al (2023)

Effect of Single-Residue Mutations on CTCF Binding to DNA: Insights from Molecular Dynamics Simulations.

International journal of molecular sciences, 24(7): pii:ijms24076395.

In humans and other eukaryotes, DNA is condensed into chromatin fibers that are further wound into chromosomes. This organization allows regulatory elements in the genome, often distant from each other in the linear DNA, to interact and facilitate gene expression through regions known as topologically associating domains (TADs). CCCTC-binding factor (CTCF) is one of the major components of TAD formation and is responsible for recruiting a partner protein, cohesin, to perform loop extrusion and facilitate proper gene expression within TADs. Because single-residue CTCF mutations have been linked to the development of a variety of cancers in humans, we aim to better understand how these mutations affect the CTCF structure and its interaction with DNA. To this end, we compare all-atom molecular dynamics simulations of a wildtype CTCF-DNA complex to those of eight different cancer-linked CTCF mutant sequences. We find that most mutants have lower binding energies compared to the wildtype protein, leading to the formation of less stable complexes. Depending on the type and position of the mutation, this loss of stability can be attributed to major changes in the electrostatic potential, loss of hydrogen bonds between the CTCF and DNA, and/or destabilization of specific zinc fingers. Interestingly, certain mutations in specific fingers can affect the interaction with the DNA of other fingers, explaining why mere single mutations can impair CTCF function. Overall, these results shed mechanistic insights into experimental observations and further underscore CTCF's importance in the regulation of chromatin architecture and gene expression.

RevDate: 2023-04-11

Tan ZW, Toong PJ, Guarnera E, et al (2023)

Disrupted chromatin architecture in olfactory sensory neurons: looking for the link from COVID-19 infection to anosmia.

Scientific reports, 13(1):5906.

We tackle here genomic mechanisms of a rapid onset and recovery from anosmia-a potential diagnostic indicator for early-stage COVID-19 infection. Based on previous observations on how olfactory receptor (OR) gene expression is regulated via chromatin structure in mice, we hypothesized that the disruption of the OR gene expression and, respectively, deficiency of the OR function can be caused by chromatin reorganization taking place upon SARS-CoV-2 infection. We obtained chromatin ensemble reconstructions from COVID-19 patients and control samples using our original computational framework for the whole-genome 3D chromatin ensemble reconstruction. Specifically, we used megabase-scale structural units and effective interactions between them obtained in the Markov State modelling of the Hi-C contact network as an unput in the stochastic embedding procedure of the whole-genome 3D chromatin ensemble reconstruction. We have also developed here a new procedure for analyzing fine structural hierarchy with (sub)TAD-size units in local chromatin regions, which we apply here to parts of chromosomes containing OR genes and corresponding regulatory elements. We observed structural modifications in COVID-19 patients on different levels of chromatin organization, from the alteration of whole genome structure and chromosomal intermingling to reorganization of contacts between chromatin loops at the level of topologically associating domains. While complementary data on known regulatory elements point to potential pathology-associated changes within the overall picture of chromatin alterations, further investigation using additional epigenetic factors mapped on 3D reconstructions with improved resolution will be required for better understanding of anosmia caused by SARS-CoV-2 infection.

RevDate: 2023-04-11

Melo US, Jatzlau J, Prada-Medina CA, et al (2023)

Enhancer hijacking at the ARHGAP36 locus is associated with connective tissue to bone transformation.

Nature communications, 14(1):2034.

Heterotopic ossification is a disorder caused by abnormal mineralization of soft tissues in which signaling pathways such as BMP, TGFβ and WNT are known key players in driving ectopic bone formation. Identifying novel genes and pathways related to the mineralization process are important steps for future gene therapy in bone disorders. In this study, we detect an inter-chromosomal insertional duplication in a female proband disrupting a topologically associating domain and causing an ultra-rare progressive form of heterotopic ossification. This structural variant lead to enhancer hijacking and misexpression of ARHGAP36 in fibroblasts, validated here by orthogonal in vitro studies. In addition, ARHGAP36 overexpression inhibits TGFβ, and activates hedgehog signaling and genes/proteins related to extracellular matrix production. Our work on the genetic cause of this heterotopic ossification case has revealed that ARHGAP36 plays a role in bone formation and metabolism, outlining first details of this gene contributing to bone-formation and -disease.

RevDate: 2023-04-05

Fritz AJ, Ghule PN, Toor R, et al (2023)

Spatiotemporal Epigenetic Control of the Histone Gene Chromatin Landscape during the Cell Cycle.

Critical reviews in eukaryotic gene expression, 33(3):85-97.

Higher-order genomic organization supports the activation of histone genes in response to cell cycle regulatory cues that epigenetically mediates stringent control of transcription at the G1/S-phase transition. Histone locus bodies (HLBs) are dynamic, non-membranous, phase-separated nuclear domains where the regulatory machinery for histone gene expression is organized and assembled to support spatiotemporal epigenetic control of histone genes. HLBs provide molecular hubs that support synthesis and processing of DNA replication-dependent histone mRNAs. These regulatory microenvironments support long-range genomic interactions among non-contiguous histone genes within a single topologically associating domain (TAD). HLBs respond to activation of the cyclin E/CDK2/NPAT/HINFP pathway at the G1/S transition. HINFP and its coactivator NPAT form a complex within HLBs that controls histone mRNA transcription to support histone protein synthesis and packaging of newly replicated DNA. Loss of HINFP compromises H4 gene expression and chromatin formation, which may result in DNA damage and impede cell cycle progression. HLBs provide a paradigm for higher-order genomic organization of a subnuclear domain that executes an obligatory cell cycle-controlled function in response to cyclin E/CDK2 signaling. Understanding the coordinately and spatiotemporally organized regulatory programs in focally defined nuclear domains provides insight into molecular infrastructure for responsiveness to cell signaling pathways that mediate biological control of growth, differentiation phenotype, and are compromised in cancer.

RevDate: 2023-04-04

Narang S, Evensen NA, Saliba J, et al (2023)

NSD2 E1099K drives relapse in pediatric acute lymphoblastic leukemia by disrupting 3D chromatin organization.

Genome biology, 24(1):64.

BACKGROUND: The NSD2 p.E1099K (EK) mutation is shown to be enriched in patients with relapsed acute lymphoblastic leukemia (ALL), indicating a role in clonal evolution and drug resistance.

RESULTS: To uncover 3D chromatin architecture-related mechanisms underlying drug resistance, we perform Hi-C on three B-ALL cell lines heterozygous for NSD2 EK. The NSD2 mutation leads to widespread remodeling of the 3D genome, most dramatically in terms of compartment changes with a strong bias towards A compartment shifts. Systematic integration of the Hi-C data with previously published ATAC-seq, RNA-seq, and ChIP-seq data show an expansion in H3K36me2 and a shrinkage in H3K27me3 within A compartments as well as increased gene expression and chromatin accessibility. These results suggest that NSD2 EK plays a prominent role in chromatin decompaction through enrichment of H3K36me2. In contrast, we identify few changes in intra-topologically associating domain activity. While compartment changes vary across cell lines, a common core of decompacting loci are shared, driving the expression of genes/pathways previously implicated in drug resistance. We further perform RNA sequencing on a cohort of matched diagnosis/relapse ALL patients harboring the relapse-specific NSD2 EK mutation. Changes in patient gene expression upon relapse significantly correlate with core compartment changes, further implicating the role of NSD2 EK in genome decompaction.

CONCLUSIONS: In spite of cell-context-dependent changes mediated by EK, there appears to be a shared transcriptional program dependent on compartment shifts which could explain phenotypic differences across EK cell lines. This core program is an attractive target for therapeutic intervention.

RevDate: 2023-04-03

Dozmorov MG, Marshall MA, Rashid NS, et al (2023)

Rewiring of the 3D genome during acquisition of carboplatin resistance in a triple-negative breast cancer patient-derived xenograft.

Scientific reports, 13(1):5420.

Changes in the three-dimensional (3D) structure of the genome are an emerging hallmark of cancer. Cancer-associated copy number variants and single nucleotide polymorphisms promote rewiring of chromatin loops, disruption of topologically associating domains (TADs), active/inactive chromatin state switching, leading to oncogene expression and silencing of tumor suppressors. However, little is known about 3D changes during cancer progression to a chemotherapy-resistant state. We integrated chromatin conformation capture (Hi-C), RNA-seq, and whole-genome sequencing obtained from triple-negative breast cancer patient-derived xenograft primary tumors (UCD52) and carboplatin-resistant samples and found increased short-range (< 2 Mb) interactions, chromatin looping, formation of TAD, chromatin state switching into a more active state, and amplification of ATP-binding cassette transporters. Transcriptome changes suggested the role of long-noncoding RNAs in carboplatin resistance. Rewiring of the 3D genome was associated with TP53, TP63, BATF, FOS-JUN family of transcription factors and led to activation of aggressiveness-, metastasis- and other cancer-related pathways. Integrative analysis highlighted increased ribosome biogenesis and oxidative phosphorylation, suggesting the role of mitochondrial energy metabolism. Our results suggest that 3D genome remodeling may be a key mechanism underlying carboplatin resistance.

RevDate: 2023-03-31

Wang B, Ji L, Q Bian (2023)

SATB1 regulates 3D genome architecture in T cells by constraining chromatin interactions surrounding CTCF-binding sites.

Cell reports, 42(4):112323 pii:S2211-1247(23)00334-0 [Epub ahead of print].

Special AT-rich sequence binding protein 1 (SATB1) has long been proposed to act as a global chromatin loop organizer in T cells. However, the exact functions of SATB1 in spatial genome organization remain elusive. Here we show that the depletion of SATB1 in human and murine T cells leads to transcriptional dysregulation for genes involved in T cell activation, as well as alterations of 3D genome architecture at multiple levels, including compartments, topologically associating domains, and loops. Importantly, SATB1 extensively colocalizes with CTCF throughout the genome. Depletion of SATB1 leads to increased chromatin contacts among and across the SATB1/CTCF co-occupied sites, thereby affecting the transcription of critical regulators of T cell activation. The loss of SATB1 does not affect CTCF occupancy but significantly reduces the retention of CTCF in the nuclear matrix. Collectively, our data show that SATB1 contributes to 3D genome organization by constraining chromatin topology surrounding CTCF-binding sites.

RevDate: 2023-03-30

Chen LF, Long HK, Park M, et al (2023)

Structural elements promote architectural stripe formation and facilitate ultra-long-range gene regulation at a human disease locus.

Molecular cell pii:S1097-2765(23)00166-1 [Epub ahead of print].

Enhancer clusters overlapping disease-associated mutations in Pierre Robin sequence (PRS) patients regulate SOX9 expression at genomic distances over 1.25 Mb. We applied optical reconstruction of chromatin architecture (ORCA) imaging to trace 3D locus topology during PRS-enhancer activation. We observed pronounced changes in locus topology between cell types. Subsequent analysis of single-chromatin fiber traces revealed that these ensemble-average differences arise through changes in the frequency of commonly sampled topologies. We further identified two CTCF-bound elements, internal to the SOX9 topologically associating domain, which promote stripe formation, are positioned near the domain's 3D geometric center, and bridge enhancer-promoter contacts in a series of chromatin loops. Ablation of these elements results in diminished SOX9 expression and altered domain-wide contacts. Polymer models with uniform loading across the domain and frequent cohesin collisions recapitulate this multi-loop, centrally clustered geometry. Together, we provide mechanistic insights into architectural stripe formation and gene regulation over ultra-long genomic ranges.

RevDate: 2023-03-29

Aldersey JE, Liu N, Tearle R, et al (2023)

Topologically associating domains in the POLLED region are the same for Angus- and Brahman-specific Hi-C reads from F1 hybrid fetal tissue.

Animal genetics [Epub ahead of print].

Horns, a form of headgear carried by Bovidae, have ethical and economic implications for ruminant production species such as cattle and goats. Hornless (polled) individuals are preferred. In cattle, four genetic variants (Celtic, Friesian, Mongolian and Guarani) are associated with the polled phenotype, which are clustered in a 300-kb region on chromosome 1. As the variants are intergenic, the functional effect is unknown. The aim of this study was to determine if the POLLED variants affect chromatin structure or disrupt enhancers using publicly available data. Topologically associating domains (TADs) were analyzed using Angus- and Brahman-specific Hi-C reads from lung tissue of an Angus (Celtic allele) cross Brahman (horned) fetus. Predicted bovine enhancers and chromatin immunoprecipitation sequencing peaks for histone modifications associated with enhancers (H3K27ac and H3K4me1) were mapped to the POLLED region. TADs analyzed from Angus- and Brahman-specific Hi-C reads were the same, therefore, the Celtic variant does not appear to affect this level of chromatin structure. The Celtic variant is located in a different TAD from the Friesian, Mongolian, and Guarani variants. Predicted enhancers and histone modifications overlapped with the Guarani and Friesian variants but not the Celtic or Mongolian variants. This study provides insight into the mechanisms of the POLLED variants for disrupting horn development. These results should be validated using data produced from the horn bud region of horned and polled bovine fetuses.

RevDate: 2023-03-21

Li X, Wang J, Yu Y, et al (2023)

Genomic rearrangements and evolutionary changes in 3D chromatin topologies in the cotton tribe (Gossypieae).

BMC biology, 21(1):56.

BACKGROUND: Analysis of the relationship between chromosomal structural variation (synteny breaks) and 3D-chromatin architectural changes among closely related species has the potential to reveal causes and correlates between chromosomal change and chromatin remodeling. Of note, contrary to extensive studies in animal species, the pace and pattern of chromatin architectural changes following the speciation of plants remain unexplored; moreover, there is little exploration of the occurrence of synteny breaks in the context of multiple genome topological hierarchies within the same model species.

RESULTS: Here we used Hi-C and epigenomic analyses to characterize and compare the profiles of hierarchical chromatin architectural features in representative species of the cotton tribe (Gossypieae), including Gossypium arboreum, Gossypium raimondii, and Gossypioides kirkii, which differ with respect to chromosome rearrangements. We found that (i) overall chromatin architectural territories were preserved in Gossypioides and Gossypium, which was reflected in their similar intra-chromosomal contact patterns and spatial chromosomal distributions; (ii) the non-random preferential occurrence of synteny breaks in A compartment significantly associate with the B-to-A compartment switch in syntenic blocks flanking synteny breaks; (iii) synteny changes co-localize with open-chromatin boundaries of topologically associating domains, while TAD stabilization has a greater influence on regulating orthologous expression divergence than do rearrangements; and (iv) rearranged chromosome segments largely maintain ancestral in-cis interactions.

CONCLUSIONS: Our findings provide insights into the non-random occurrence of epigenomic remodeling relative to the genomic landscape and its evolutionary and functional connections to alterations of hierarchical chromatin architecture, on a known evolutionary timescale.

RevDate: 2023-03-14

Li D, Wu F, Zhou S, et al (2023)

Heterochromatin rewiring and domain disruption-mediated chromatin compaction during erythropoiesis.

Nature structural & molecular biology [Epub ahead of print].

Mammalian erythropoiesis involves progressive chromatin compaction and subsequent enucleation in terminal differentiation, but the mechanisms underlying the three-dimensional chromatin reorganization remain obscure. Here, we systematically analyze the higher-order chromatin in purified populations of primary human erythroblasts. Our results reveal that heterochromatin regions undergo substantial compression, with H3K9me3 markers relocalizing to the nuclear periphery and forming a significant number of long-range interactions, and that ~58% of the topologically associating domain (TAD) boundaries are disrupted, while certain TADs enriched for markers of the active transcription state and erythroid master regulators, GATA1 and KLF1, are selectively maintained during terminal erythropoiesis. Finally, we demonstrate that GATA1 is involved in safeguarding selected essential chromatin domains during terminal erythropoiesis. Our study therefore delineates the molecular characteristics of a development-driven chromatin compaction process, which reveals transcription competence as a key indicator of the selected domain maintenance to ensure appropriate gene expression during the extreme compaction of chromatin.

RevDate: 2023-03-09

Dang D, Zhang SW, Duan R, et al (2023)

Defining the separation landscape of topological domains for decoding consensus domain organization of 3D genome.

Genome research pii:gr.277187.122 [Epub ahead of print].

Topologically associating domains (TADs) have emerged as basic structural and functional units of genome organization, and have been determined by many computational methods from Hi-C contact maps. However, the TADs obtained by different methods vary greatly, which makes the accurate determination of TADs a challenging issue and hinders subsequent biological analyses about their organization and functions. Obvious inconsistencies among the TADs identified by different methods indeed make the statistical and biological properties of TADs overly depend on the method we chose rather than on the data. To this end, we employ the consensus structural information captured by these methods to define the TAD separation landscape for decoding the consensus domain organization of the 3D genome. We demonstrate that the TAD separation landscape could be used to compare domain boundaries across multiple cell types for discovering conserved and divergent topological structures, decipher three types of boundary regions with diverse biological features, and identify Consensus TADs (ConsTADs). We illustrate that these analyses could deepen our understanding of the relationships between the topological domains and chromatin states, gene expression, and DNA replication timing.

RevDate: 2023-03-03

Richer S, Tian Y, Schoenfelder S, et al (2023)

Widespread allele-specific topological domains in the human genome are not confined to imprinted gene clusters.

Genome biology, 24(1):40.

BACKGROUND: There is widespread interest in the three-dimensional chromatin conformation of the genome and its impact on gene expression. However, these studies frequently do not consider parent-of-origin differences, such as genomic imprinting, which result in monoallelic expression. In addition, genome-wide allele-specific chromatin conformation associations have not been extensively explored. There are few accessible bioinformatic workflows for investigating allelic conformation differences and these require pre-phased haplotypes which are not widely available.

RESULTS: We developed a bioinformatic pipeline, "HiCFlow," that performs haplotype assembly and visualization of parental chromatin architecture. We benchmarked the pipeline using prototype haplotype phased Hi-C data from GM12878 cells at three disease-associated imprinted gene clusters. Using Region Capture Hi-C and Hi-C data from human cell lines (1-7HB2, IMR-90, and H1-hESCs), we can robustly identify the known stable allele-specific interactions at the IGF2-H19 locus. Other imprinted loci (DLK1 and SNRPN) are more variable and there is no "canonical imprinted 3D structure," but we could detect allele-specific differences in A/B compartmentalization. Genome-wide, when topologically associating domains (TADs) are unbiasedly ranked according to their allele-specific contact frequencies, a set of allele-specific TADs could be defined. These occur in genomic regions of high sequence variation. In addition to imprinted genes, allele-specific TADs are also enriched for allele-specific expressed genes. We find loci that have not previously been identified as allele-specific expressed genes such as the bitter taste receptors (TAS2Rs).

CONCLUSIONS: This study highlights the widespread differences in chromatin conformation between heterozygous loci and provides a new framework for understanding allele-specific expressed genes.

RevDate: 2023-02-26

Karpinska MA, AM Oudelaar (2023)

The role of loop extrusion in enhancer-mediated gene activation.

Current opinion in genetics & development, 79:102022 pii:S0959-437X(23)00002-3 [Epub ahead of print].

Gene expression patterns in complex multicellular organisms are regulated by enhancers, which communicate with their target gene promoters in three-dimensional (3D) chromatin structures. Despite advances in our understanding of the mechanisms that organize mammalian genomes into compartments and topologically associating domains (TADs), it is not well understood how specific interactions between enhancers and promoters are controlled in this 3D context. In this review, we give an overview of recent evidence that shows that a process of loop extrusion plays an important role in the regulation of enhancer-promoter communication and discuss recent insights into the molecular mechanism by which loop extrusion contributes to enhancer-mediated gene activation.

RevDate: 2023-02-25

Sabaté T, Lelandais B, Bertrand E, et al (2023)

Polymer simulations guide the detection and quantification of chromatin loop extrusion by imaging.

Nucleic acids research pii:7058223 [Epub ahead of print].

Genome-wide chromosome conformation capture (Hi-C) has revealed the organization of chromatin into topologically associating domains (TADs) and loops, which are thought to help regulate genome functions. TADs and loops are understood as the result of DNA extrusion mediated by the cohesin complex. However, despite recent efforts, direct visualization and quantification of this process in single cells remains an open challenge. Here, we use polymer simulations and dedicated analysis methods to explore if, and under which conditions, DNA loop extrusion can be detected and quantitatively characterized by imaging pairs of fluorescently labeled loci located near loop or TAD anchors in fixed or living cells. We find that under realistic conditions, extrusion can be detected and the frequency of loop formation can be quantified from fixed cell images alone, while the lifetime of loops and the speed of extrusion can be estimated from dynamic live-cell data. Our delineation of appropriate imaging conditions and the proposed analytical methods lay the groundwork for a systematic quantitative characterization of loop extrusion in fixed or living cells.

RevDate: 2023-02-17

Zhao Y, Ding Y, He L, et al (2023)

Multiscale 3D genome reorganization during skeletal muscle stem cell lineage progression and aging.

Science advances, 9(7):eabo1360.

Little is known about three-dimensional (3D) genome organization in skeletal muscle stem cells [also called satellite cells (SCs)]. Here, we comprehensively map the 3D genome topology reorganization during mouse SC lineage progression. Specifically, rewiring at the compartment level is most pronounced when SCs become activated. Marked loss in topologically associating domain (TAD) border insulation and chromatin looping also occurs during early activation process. Meanwhile, TADs can form TAD clusters and super-enhancer-containing TAD clusters orchestrate stage-specific gene expression. Furthermore, we uncover that transcription factor PAX7 is pivotal in enhancer-promoter (E-P) loop formation. We also identify cis-regulatory elements that are crucial for local chromatin organization at Pax7 locus and Pax7 expression. Lastly, we unveil that geriatric SC displays a prominent gain in long-range contacts and loss of TAD border insulation. Together, our results uncover that 3D chromatin extensively reorganizes at multiple architectural levels and underpins the transcriptome remodeling during SC lineage development and SC aging.

RevDate: 2023-02-09

Kobets VA, Ulianov SV, Galitsyna AA, et al (2023)

HiConfidence: a novel approach uncovering the biological signal in Hi-C data affected by technical biases.

Briefings in bioinformatics pii:7033301 [Epub ahead of print].

The chromatin interaction assays, particularly Hi-C, enable detailed studies of genome architecture in multiple organisms and model systems, resulting in a deeper understanding of gene expression regulation mechanisms mediated by epigenetics. However, the analysis and interpretation of Hi-C data remain challenging due to technical biases, limiting direct comparisons of datasets obtained in different experiments and laboratories. As a result, removing biases from Hi-C-generated chromatin contact matrices is a critical data analysis step. Our novel approach, HiConfidence, eliminates biases from the Hi-C data by weighing chromatin contacts according to their consistency between replicates so that low-quality replicates do not substantially influence the result. The algorithm is effective for the analysis of global changes in chromatin structures such as compartments and topologically associating domains. We apply the HiConfidence approach to several Hi-C datasets with significant technical biases, that could not be analyzed effectively using existing methods, and obtain meaningful biological conclusions. In particular, HiConfidence aids in the study of how changes in histone acetylation pattern affect chromatin organization in Drosophila melanogaster S2 cells. The method is freely available at GitHub: https://github.com/victorykobets/HiConfidence.

RevDate: 2023-02-09
CmpDate: 2023-02-09

Maslova A, Plotnikov V, Nuriddinov M, et al (2023)

Hi-C analysis of genomic contacts revealed karyotype abnormalities in chicken HD3 cell line.

BMC genomics, 24(1):66.

BACKGROUND: Karyotype abnormalities are frequent in immortalized continuous cell lines either transformed or derived from primary tumors. Chromosomal rearrangements can cause dramatic changes in gene expression and affect cellular phenotype and behavior during in vitro culture. Structural variations of chromosomes in many continuous mammalian cell lines are well documented, but chromosome aberrations in cell lines from other vertebrate models often remain understudied. The chicken LSCC-HD3 cell line (HD3), generated from erythroid precursors, was used as an avian model for erythroid differentiation and lineage-specific gene expression. However, karyotype abnormalities in the HD3 cell line were not assessed. In the present study, we applied high-throughput chromosome conformation capture to analyze 3D genome organization and to detect chromosome rearrangements in the HD3 cell line.

RESULTS: We obtained Hi-C maps of genomic interactions for the HD3 cell line and compared A/B compartments and topologically associating domains between HD3 and several other cell types. By analysis of contact patterns in the Hi-C maps of HD3 cells, we identified more than 25 interchromosomal translocations of regions ≥ 200 kb on both micro- and macrochromosomes. We classified most of the observed translocations as unbalanced, leading to the formation of heteromorphic chromosomes. In many cases of microchromosome rearrangements, an entire microchromosome together with other macro- and microchromosomes participated in the emergence of a derivative chromosome, resembling "chromosomal fusions'' between acrocentric microchromosomes. Intrachromosomal inversions, deletions and duplications were also detected in HD3 cells. Several of the identified simple and complex chromosomal rearrangements, such as between GGA2 and GGA1qter; GGA5, GGA4p and GGA7p; GGA4q, GGA6 and GGA19; and duplication of the sex chromosome GGAW, were confirmed by FISH.

CONCLUSIONS: In the erythroid progenitor HD3 cell line, in contrast to mature and immature erythrocytes, the genome is organized into distinct topologically associating domains. The HD3 cell line has a severely rearranged karyotype with most of the chromosomes engaged in translocations and can be used in studies of genome structure-function relationships. Hi-C proved to be a reliable tool for simultaneous assessment of the spatial genome organization and chromosomal aberrations in karyotypes of birds with a large number of microchromosomes.

RevDate: 2023-02-07
CmpDate: 2023-02-07

Yang L, Akgol Oksuz B, Dekker J, et al (2023)

Capturing Chromosome Conformation Across Length Scales.

Journal of visualized experiments : JoVE.

Chromosome conformation capture (3C) is used to detect three-dimensional chromatin interactions. Typically, chemical crosslinking with formaldehyde (FA) is used to fix chromatin interactions. Then, chromatin digestion with a restriction enzyme and subsequent religation of fragment ends converts three-dimensional (3D) proximity into unique ligation products. Finally, after reversal of crosslinks, protein removal, and DNA isolation, DNA is sheared and prepared for high-throughput sequencing. The frequency of proximity ligation of pairs of loci is a measure of the frequency of their colocalization in three-dimensional space in a cell population. A sequenced Hi-C library provides genome-wide information on interaction frequencies between all pairs of loci. The resolution and precision of Hi-C relies on efficient crosslinking that maintains chromatin contacts and frequent and uniform fragmentation of the chromatin. This paper describes an improved in situ Hi-C protocol, Hi-C 3.0, that increases the efficiency of crosslinking by combining two crosslinkers (formaldehyde [FA] and disuccinimidyl glutarate [DSG]), followed by finer digestion using two restriction enzymes (DpnII and DdeI). Hi-C 3.0 is a single protocol for the accurate quantification of genome folding features at smaller scales such as loops and topologically associating domains (TADs), as well as features at larger nucleus-wide scales such as compartments.

RevDate: 2023-02-05

Selcen I, Prentice E, P Casaccia (2023)

The epigenetic landscape of oligodendrocyte lineage cells.

Annals of the New York Academy of Sciences [Epub ahead of print].

The epigenetic landscape of oligodendrocyte lineage cells refers to the cell-specific modifications of DNA, chromatin, and RNA that define a unique gene expression pattern of functionally specialized cells. Here, we focus on the epigenetic changes occurring as progenitors differentiate into myelin-forming cells and respond to the local environment. First, modifications of DNA, RNA, nucleosomal histones, key principles of chromatin organization, topologically associating domains, and local remodeling will be reviewed. Then, the relationship between epigenetic modulators and RNA processing will be explored. Finally, the reciprocal relationship between the epigenome as a determinant of the mechanical properties of cell nuclei and the target of mechanotransduction will be discussed. The overall goal is to provide an interpretative key on how epigenetic changes may account for the heterogeneity of the transcriptional profiles identified in this lineage.

RevDate: 2023-02-03

Costea J, Schoeberl UE, Malzl D, et al (2023)

A de novo transcription-dependent TAD boundary underpins critical multiway interactions during antibody class switch recombination.

Molecular cell pii:S1097-2765(23)00037-0 [Epub ahead of print].

Interactions between transcription and cohesin-mediated loop extrusion can influence 3D chromatin architecture. However, their relevance in biology is unclear. Here, we report a direct role for such interactions in the mechanism of antibody class switch recombination (CSR) at the murine immunoglobulin heavy chain locus (Igh). Using Tri-C to measure higher-order multiway interactions on single alleles, we find that the juxtaposition (synapsis) of transcriptionally active donor and acceptor Igh switch (S) sequences, an essential step in CSR, occurs via the interaction of loop extrusion complexes with a de novo topologically associating domain (TAD) boundary formed via transcriptional activity across S regions. Surprisingly, synapsis occurs predominantly in proximity to the 3' CTCF-binding element (3'CBE) rather than the Igh super-enhancer, suggesting a two-step mechanism whereby transcription of S regions is not topologically coupled to synapsis, as has been previously proposed. Altogether, these insights advance our understanding of how 3D chromatin architecture regulates CSR.

RevDate: 2023-02-09
CmpDate: 2023-02-07

Cavalheiro GR, Girardot C, Viales RR, et al (2023)

CTCF, BEAF-32, and CP190 are not required for the establishment of TADs in early Drosophila embryos but have locus-specific roles.

Science advances, 9(5):eade1085.

The boundaries of topologically associating domains (TADs) are delimited by insulators and/or active promoters; however, how they are initially established during embryogenesis remains unclear. Here, we examined this during the first hours of Drosophila embryogenesis. DNA-FISH confirms that intra-TAD pairwise proximity is established during zygotic genome activation (ZGA) but with extensive cell-to-cell heterogeneity. Most newly formed boundaries are occupied by combinations of CTCF, BEAF-32, and/or CP190. Depleting each insulator individually from chromatin revealed that TADs can still establish, although with lower insulation, with a subset of boundaries (~10%) being more dependent on specific insulators. Some weakened boundaries have aberrant gene expression due to unconstrained enhancer activity. However, the majority of misexpressed genes have no obvious direct relationship to changes in domain-boundary insulation. Deletion of an active promoter (thereby blocking transcription) at one boundary had a greater impact than deleting the insulator-bound region itself. This suggests that cross-talk between insulators and active promoters and/or transcription might reinforce domain boundary insulation during embryogenesis.

RevDate: 2023-02-02
CmpDate: 2023-02-02

Landshammer A, Bolondi A, Kretzmer H, et al (2023)

T-REX17 is a transiently expressed non-coding RNA essential for human endoderm formation.

eLife, 12:.

Long non-coding RNAs (lncRNAs) have emerged as fundamental regulators in various biological processes, including embryonic development and cellular differentiation. Despite much progress over the past decade, the genome-wide annotation of lncRNAs remains incomplete and many known non-coding loci are still poorly characterized. Here, we report the discovery of a previously unannotated lncRNA that is transcribed 230 kb upstream of the SOX17 gene and located within the same topologically associating domain. We termed it T-REX17 (Transcript Regulating Endoderm and activated by soX17) and show that it is induced following SOX17 activation but its expression is more tightly restricted to early definitive endoderm. Loss of T-REX17 affects crucial functions independent of SOX17 and leads to an aberrant endodermal transcriptome, signaling pathway deregulation and epithelial to mesenchymal transition defects. Consequently, cells lacking the lncRNA cannot further differentiate into more mature endodermal cell types. Taken together, our study identified and characterized T-REX17 as a transiently expressed and essential non-coding regulator in early human endoderm differentiation.

RevDate: 2023-02-03
CmpDate: 2023-02-03

Barajas-Mora EM, Lee L, Lu H, et al (2023)

Enhancer-instructed epigenetic landscape and chromatin compartmentalization dictate a primary antibody repertoire protective against specific bacterial pathogens.

Nature immunology, 24(2):320-336.

Antigen receptor loci are organized into variable (V), diversity (D) and joining (J) gene segments that rearrange to generate antigen receptor repertoires. Here, we identified an enhancer (E34) in the murine immunoglobulin kappa (Igk) locus that instructed rearrangement of Vκ genes located in a sub-topologically associating domain, including a Vκ gene encoding for antibodies targeting bacterial phosphorylcholine. We show that E34 instructs the nuclear repositioning of the E34 sub-topologically associating domain from a recombination-repressive compartment to a recombination-permissive compartment that is marked by equivalent activating histone modifications. Finally, we found that E34-instructed Vκ-Jκ rearrangement was essential to combat Streptococcus pneumoniae but not methicillin-resistant Staphylococcus aureus or influenza infections. We propose that the merging of Vκ genes with Jκ elements is instructed by one-dimensional epigenetic information imposed by enhancers across Vκ and Jκ genomic regions. The data also reveal how enhancers generate distinct antibody repertoires that provide protection against lethal bacterial infection.

RevDate: 2023-01-28

Nayak S, Jiang K, Hope E, et al (2023)

Chromatin landscape governing murine epidermal differentiation.

The Journal of investigative dermatology pii:S0022-202X(23)00017-9 [Epub ahead of print].

Chromatin landscape and regulatory networks are determinant in lineage specification and differentiation. To define the temporospatial differentiation axis in murine epidermal cells in vivo, we generated datasets profiling expression dynamics (RNA-Seq), chromatin accessibility (ATAC-Seq), architecture (Hi-C), and histone modifications (ChIP-Seq) in the epidermis. We show that many differentially regulated genes are suppressed during the differentiation process, with super-enhancers (SEs) controlling differentiation-specific epigenomic changes. Our data shows the relevance of the Dlx/Klf/Grhl combinatorial regulatory network in maintaining correct temporospatial gene expression during epidermal differentiation. We determined differential open compartments, topologically associating domain (TAD) score and looping in the Basal cell (B) and Suprabasal cell (SB) epidermal fractions, with the evolutionarily conserved Epidermal Differentiation Complex (EDC) region showing distinct SB-specific TAD and loop formation that coincided with SE sites. Overall, our study provides a global genome-wide resource of chromatin dynamics that define unrecognized regulatory networks and the epigenetic control of Dlx3-bound SE elements during epidermal differentiation.

RevDate: 2023-01-28

Fan S, Dang D, Ye Y, et al (2023)

scHi-CSim: a flexible simulator that generates high-fidelity single-cell Hi-C data for benchmarking.

Journal of molecular cell biology pii:7008500 [Epub ahead of print].

Single-cell Hi-C technology provides an unprecedented opportunity to reveal chromatin structure in individual cells. However, high sequencing cost impedes the generation of biological Hi-C data with high sequencing depths and multiple replicates for downstream analysis. Here we developed a single-cell Hi-C simulator (scHi-CSim) that generates high-fidelity data for benchmarking. scHi-CSim merges neighboring cells to overcome the sparseness of data, samples interactions in distance-stratified chromosomes to maintain the heterogeneity of single cells, and estimates the empirical distribution of restriction fragments to generate simulated data. We demonstrated that scHi-CSim can generate high-fidelity data by comparing the performance of single-cell clustering and detection of chromosomal high-order structures with raw data. Furthermore, scHi-CSim is flexible to change sequencing depths and the number of simulated replicates. We showed that increasing sequencing depths could improve the accuracy of detecting topologically associating domains. We also used scHi-CSim to generate a series of simulated datasets with different sequencing depths to benchmark single-cell Hi-C clustering methods.

RevDate: 2023-02-07
CmpDate: 2023-02-07

Kim S, J Wysocka (2023)

Deciphering the multi-scale, quantitative cis-regulatory code.

Molecular cell, 83(3):373-392.

Uncovering the cis-regulatory code that governs when and how much each gene is transcribed in a given genome and cellular state remains a central goal of biology. Here, we discuss major layers of regulation that influence how transcriptional outputs are encoded by DNA sequence and cellular context. We first discuss how transcription factors bind specific DNA sequences in a dosage-dependent and cooperative manner and then proceed to the cofactors that facilitate transcription factor function and mediate the activity of modular cis-regulatory elements such as enhancers, silencers, and promoters. We then consider the complex and poorly understood interplay of these diverse elements within regulatory landscapes and its relationships with chromatin states and nuclear organization. We propose that a mechanistically informed, quantitative model of transcriptional regulation that integrates these multiple regulatory layers will be the key to ultimately cracking the cis-regulatory code.

RevDate: 2023-02-06
CmpDate: 2023-01-23

Ni L, Liu Y, Ma X, et al (2023)

Pan-3D genome analysis reveals structural and functional differentiation of soybean genomes.

Genome biology, 24(1):12.

BACKGROUND: High-order chromatin structure plays important roles in gene regulation. However, the diversity of the three-dimensional (3D) genome across plant accessions are seldom reported.

RESULTS: Here, we perform the pan-3D genome analysis using Hi-C sequencing data from 27 soybean accessions and comprehensively investigate the relationships between 3D genomic variations and structural variations (SVs) as well as gene expression. We find that intersection regions between A/B compartments largely contribute to compartment divergence. Topologically associating domain (TAD) boundaries in A compartments exhibit significantly higher density compared to those in B compartments. Pan-3D genome analysis shows that core TAD boundaries have the highest transcription start site (TSS) density and lowest GC content and repeat percentage. Further investigation shows that non-long terminal repeat (non-LTR) retrotransposons play important roles in maintaining TAD boundaries, while Gypsy elements and satellite repeats are associated with private TAD boundaries. Moreover, presence and absence variation (PAV) is found to be the major contributor to 3D genome variations. Nevertheless, approximately 55% of 3D genome variations are not associated with obvious genetic variations, and half of them affect the flanking gene expression. In addition, we find that the 3D genome may also undergo selection during soybean domestication.

CONCLUSION: Our study sheds light on the role of 3D genomes in plant genetic diversity and provides a valuable resource for studying gene regulation and genome evolution.

RevDate: 2023-02-08
CmpDate: 2023-02-03

Islam Z, Saravanan B, Walavalkar K, et al (2023)

Active enhancers strengthen insulation by RNA-mediated CTCF binding at chromatin domain boundaries.

Genome research, 33(1):1-17.

Vertebrate genomes are partitioned into chromatin domains or topologically associating domains (TADs), which are typically bound by head-to-head pairs of CTCF binding sites. Transcription at domain boundaries correlates with better insulation; however, it is not known whether the boundary transcripts themselves contribute to boundary function. Here we characterize boundary-associated RNAs genome-wide, focusing on the disease-relevant INK4a/ARF and MYC TAD. Using CTCF site deletions and boundary-associated RNA knockdowns, we observe that boundary-associated RNAs facilitate recruitment and clustering of CTCF at TAD borders. The resulting CTCF enrichment enhances TAD insulation, enhancer-promoter interactions, and TAD gene expression. Importantly, knockdown of boundary-associated RNAs results in loss of boundary insulation function. Using enhancer deletions and CRISPRi of promoters, we show that active TAD enhancers, but not promoters, induce boundary-associated RNA transcription, thus defining a novel class of regulatory enhancer RNAs.

RevDate: 2023-01-15

Yeo SJ, Ying C, Fullwood MJ, et al (2023)

Emerging regulatory mechanisms of noncoding RNAs in topologically associating domains.

Trends in genetics : TIG pii:S0168-9525(22)00312-2 [Epub ahead of print].

Topologically associating domains (TADs) are integral to spatial genome organization, instructing gene expression, and cell fate. Recently, several advances have uncovered roles for noncoding RNAs (ncRNAs) in the regulation of the form and function of mammalian TADs. Phase separation has also emerged as a potential arbiter of ncRNAs in the regulation of TADs. In this review we discuss the implications of these novel findings in relation to how ncRNAs might structurally and functionally regulate TADs from two perspectives: moderating loop extrusion through interactions with architectural proteins, and facilitating TAD phase separation. Additionally, we propose future studies and directions to investigate these phenomena.

RevDate: 2023-02-08
CmpDate: 2023-02-03

Mulhair PO, Crowley L, Boyes DH, et al (2023)

Diversity, duplication, and genomic organization of homeobox genes in Lepidoptera.

Genome research, 33(1):32-44.

Homeobox genes encode transcription factors with essential roles in patterning and cell fate in developing animal embryos. Many homeobox genes, including Hox and NK genes, are arranged in gene clusters, a feature likely related to transcriptional control. Sparse taxon sampling and fragmentary genome assemblies mean that little is known about the dynamics of homeobox gene evolution across Lepidoptera or about how changes in homeobox gene number and organization relate to diversity in this large order of insects. Here we analyze an extensive data set of high-quality genomes to characterize the number and organization of all homeobox genes in 123 species of Lepidoptera from 23 taxonomic families. We find most Lepidoptera have around 100 homeobox loci, including an unusual Hox gene cluster in which the lab gene is repositioned and the ro gene is next to pb A topologically associating domain spans much of the gene cluster, suggesting deep regulatory conservation of the Hox cluster arrangement in this insect order. Most Lepidoptera have four Shx genes, divergent zen-derived loci, but these loci underwent dramatic duplication in several lineages, with some moths having over 165 homeobox loci in the Hox gene cluster; this expansion is associated with local LINE element density. In contrast, the NK gene cluster content is more stable, although there are differences in organization compared with other insects, as well as major rearrangements within butterflies. Our analysis represents the first description of homeobox gene content across the order Lepidoptera, exemplifying the potential of newly generated genome assemblies for understanding genome and gene family evolution.

RevDate: 2023-01-17
CmpDate: 2023-01-17

Zhang L, Xu M, Zhang W, et al (2023)

Three-dimensional genome landscape comprehensively reveals patterns of spatial gene regulation in papillary and anaplastic thyroid cancers: a study using representative cell lines for each cancer type.

Cellular & molecular biology letters, 28(1):1.

BACKGROUND: Spatial chromatin structure is intricately linked with somatic aberrations, and somatic mutations of various cancer-related genes, termed co-mutations (CoMuts), occur in certain patterns during cancer initiation and progression. The functional mechanisms underlying these genetic events remain largely unclear in thyroid cancer (TC). With discrepant differentiation, papillary thyroid cancer (PTC) and anaplastic thyroid cancer (ATC) differ greatly in characteristics and prognosis. We aimed to reveal the spatial gene alterations and regulations between the two TC subtypes.

METHODS: We systematically investigated and compared the spatial co-mutations between ATC (8305C), PTC (BCPAP and TPC-1), and normal thyroid cells (Nthy-ori-3-1). We constructed a framework integrating whole-genome sequencing (WGS), high-throughput chromosome conformation capture (Hi-C), and transcriptome sequencing, to systematically detect the associations between the somatic co-mutations of cancer-related genes, structural variations (SVs), copy number variations (CNVs), and high-order chromatin conformation.

RESULTS: Spatial co-mutation hotspots were enriched around topologically associating domains (TADs) in TC. A common set of 227 boundaries were identified in both ATC and PTC, with significant overlaps between them. The spatial proximities of the co-mutated gene pairs in the two TC types were significantly greater than in the gene-level and overall backgrounds, and ATC cells had higher TAD contact frequency with CoMuts > 10 compared with PTC cells. Compared with normal thyroid cells, in ATC the number of the created novel three-dimensional chromatin structural domains increased by 10%, and the number of shifted TADs decreased by 7%. We found five TAD blocks with CoMut genes/events specific to ATC with certain mutations in genes including MAST-NSUN4, AM129B/TRUB2, COL5A1/PPP1R26, PPP1R26/GPSM1/CCDC183, and PRAC2/DLX4. For the majority of ATC and PTC cells, the HOXA10 and HIF2α signals close to the transcription start sites of CoMut genes within TADs were significantly stronger than those at the background. CNV breakpoints significantly overlapped with TAD boundaries in both TC subtypes. ATCs had more CNV losses overlapping with TAD boundaries, and noncoding SVs involved in intrachromosomal SVs, amplified inversions, and tandem duplication differed between ATC and PTC. TADs with short range were more abundant in ATC than PTC. More switches of A/B compartment types existed in ATC cells compared with PTC. Gene expression was significantly synchronized, and orchestrated by complex epigenetics and regulatory elements.

CONCLUSION: Chromatin interactions and gene alterations and regulations are largely heterogeneous in TC. CNVs and complex SVs may function in the TC genome by interplaying with TADs, and are largely different between ATC and PTC. Complexity of TC genomes, which are highly organized by 3D genome-wide interactions mediating mutational and structural variations and gene activation, may have been largely underappreciated. Our comprehensive analysis may provide key evidence and targets for more customized diagnosis and treatment of TC.

RevDate: 2023-02-02
CmpDate: 2023-01-26

van Mierlo G, Pushkarev O, Kribelbauer JF, et al (2023)

Chromatin modules and their implication in genomic organization and gene regulation.

Trends in genetics : TIG, 39(2):140-153.

Regulation of gene expression is a complex but highly guided process. While genomic technologies and computational approaches have allowed high-throughput mapping of cis-regulatory elements (CREs) and their interactions in 3D, their precise role in regulating gene expression remains obscure. Recent complementary observations revealed that interactions between CREs frequently result in the formation of small-scale functional modules within topologically associating domains. Such chromatin modules likely emerge from a complex interplay between regulatory machineries assembled at CREs, including site-specific binding of transcription factors. Here, we review the methods that allow identifying chromatin modules, summarize possible mechanisms that steer CRE interactions within these modules, and discuss outstanding challenges to uncover how chromatin modules fit in our current understanding of the functional 3D genome.

RevDate: 2022-12-26

Kato H, Tateishi K, Iwadate D, et al (2022)

HNF1B-driven three-dimensional chromatin structure for molecular classification in pancreatic cancers.

Cancer science [Epub ahead of print].

The molecular subtypes of pancreatic cancer (PC), either classical/progenitor-like or basal/squamous-like, are currently a major topic of research because of their direct association with clinical outcomes. Some transcription factors (TFs) have been reported to be associated with these subtypes. However, the mechanisms by which these molecular signatures of PCs are established remain unknown. Epigenetic regulatory processes, supported by dynamic changes in the chromatin structure, are essential for transcriptional profiles. Previously, we reported the importance of open chromatin profiles in the biological features and transcriptional status of PCs. Here, we aimed to analyze the relationships between three-dimensional (3D) genome structures and the molecular subtypes of human PCs using Hi-C analysis. We observed a correlation of the specific elements of 3D genome modules, including compartments, topologically associating domains, and enhancer-promoter loops, with the expression of related genes. We focused on HNF1B, a TF that is implicated in the progenitor subtype. Forced expression of HNF1B in squamous-type PC organoids induced the upregulation and downregulation of genes associated with progenitor and squamous subtypes, respectively. Long-range genomic interactions induced by HNF1B were accompanied by compartment modulation and H3K27ac redistribution. We also found that these HNF1B-induced changes in subtype-related gene expression required an intrinsically disordered region, suggesting a possible involvement of phase separation in compartment modulation. Thus, mapping of 3D structural changes induced by TFs, such as HNF1B, may become a useful resource for further understanding the molecular features of PCs.

RevDate: 2023-02-10
CmpDate: 2023-01-13

Alavattam KG, Mitzelfelt KA, Bonora G, et al (2023)

Dynamic chromatin organization and regulatory interactions in human endothelial cell differentiation.

Stem cell reports, 18(1):159-174.

Vascular endothelial cells are a mesoderm-derived lineage with many essential functions, including angiogenesis and coagulation. The gene-regulatory mechanisms underpinning endothelial specialization are largely unknown, as are the roles of chromatin organization in regulating endothelial cell transcription. To investigate the relationships between chromatin organization and gene expression, we induced endothelial cell differentiation from human pluripotent stem cells and performed Hi-C and RNA-sequencing assays at specific time points. Long-range intrachromosomal contacts increase over the course of differentiation, accompanied by widespread heteroeuchromatic compartment transitions that are tightly associated with transcription. Dynamic topologically associating domain boundaries strengthen and converge on an endothelial cell state, and function to regulate gene expression. Chromatin pairwise point interactions (DNA loops) increase in frequency during differentiation and are linked to the expression of genes essential to vascular biology. Chromatin dynamics guide transcription in endothelial cell development and promote the divergence of endothelial cells from cardiomyocytes.

RevDate: 2022-12-15
CmpDate: 2022-12-15

Yang JY, JM Chang (2022)

Pattern recognition of topologically associating domains using deep learning.

BMC bioinformatics, 22(Suppl 10):634.

BACKGROUND: Recent increasing evidence indicates that three-dimensional chromosome structure plays an important role in genomic function. Topologically associating domains (TADs) are self-interacting regions that have been shown to be a chromosomal structural unit. During evolution, these are conserved based on checking synteny block cross species. Are there common TAD patterns across species or cell lines?

RESULTS: To address the above question, we propose a novel task-TAD recognition-as opposed to traditional TAD identification. Specifically, we treat Hi-C maps as images, thus re-casting TAD recognition as image pattern recognition, for which we use a convolutional neural network and a residual neural network. In addition, we propose an elegant way to generate non-TAD data for binary classification. We demonstrate deep learning performance which is quite promising, AUC > 0.80, through cross-species and cell-type validation.

CONCLUSIONS: TADs have been shown to be conserved during evolution. Interestingly, our results confirm that the TAD recognition model is practical across species, which indicates that TADs between human and mouse show common patterns from an image classification point of view. Our approach could be a new way to identify TAD variations or patterns among Hi-C maps. For example, TADs of two Hi-C maps are conserved if the two classification models are exchangeable.

RevDate: 2023-01-12
CmpDate: 2022-12-15

Mach P, Kos PI, Zhan Y, et al (2022)

Cohesin and CTCF control the dynamics of chromosome folding.

Nature genetics, 54(12):1907-1918.

In mammals, interactions between sequences within topologically associating domains enable control of gene expression across large genomic distances. Yet it is unknown how frequently such contacts occur, how long they last and how they depend on the dynamics of chromosome folding and loop extrusion activity of cohesin. By imaging chromosomal locations at high spatial and temporal resolution in living cells, we show that interactions within topologically associating domains are transient and occur frequently during the course of a cell cycle. Interactions become more frequent and longer in the presence of convergent CTCF sites, resulting in suppression of variability in chromosome folding across time. Supported by physical models of chromosome dynamics, our data suggest that CTCF-anchored loops last around 10 min. Our results show that long-range transcriptional regulation might rely on transient physical proximity, and that cohesin and CTCF stabilize highly dynamic chromosome structures, facilitating selected subsets of chromosomal interactions.

RevDate: 2023-01-13
CmpDate: 2022-12-07

Zhao X, Zhu S, Peng W, et al (2022)

The Interplay of Transcription and Genome Topology Programs T Cell Development and Differentiation.

Journal of immunology (Baltimore, Md. : 1950), 209(12):2269-2278.

T cells are essential for mounting defense against various pathogens and malignantly transformed cells. Thymic development and peripheral T cell differentiation are highly orchestrated biological processes that require precise gene regulation. Higher-order genome organization on multiple scales, in the form of chromatin loops, topologically associating domains and compartments, provides pivotal control of T cell gene expression. CTCF and the cohesin machinery are ubiquitously expressed architectural proteins responsible for establishing chromatin structures. Recent studies indicate that transcription factors, such as T lineage-defining Tcf1 and TCR-induced Batf, may have intrinsic ability and/or engage CTCF to shape chromatin architecture. In this article, we summarize current knowledge on the dynamic changes in genome topology that underlie normal or leukemic T cell development, CD4+ helper T cell differentiation, and CD8+ cytotoxic T cell functions. The knowledge lays a solid foundation for elucidating the causative link of spatial chromatin configuration to transcriptional and functional output in T cells.

RevDate: 2022-12-06

Wang X, Yang B, Zhao W, et al (2022)

Capture Hi-C reveals the influence on dynamic three-dimensional chromosome organization perturbed by genetic variation or vanillin stress in Saccharomyces cerevisiae.

Frontiers in microbiology, 13:1012377.

Studying the mechanisms of resistance to vanillin in microorganisms, which is derived from lignin and blocks a major pathway of DNA double-strand break repair in yeast, will benefit the design of robust cell factories that produce biofuels and chemicals using lignocellulosic materials. A high vanillin-tolerant Saccharomyces cerevisiae strain EMV-8 carrying site mutations compared to its parent strain NAN-27 was selected for the analyses. The dynamics of the chromatin structure of eukaryotic cells play a critical role in transcription and the regulation of gene expression and thus the phenotype. Consequently, Hi-C and transcriptome analyses were conducted in EMV-8 and NAN-27 in the log phase with or without vanillin stress to determine the effects of mutations and vanillin disturbance on the dynamics of three-dimensional chromosome organization and the influence of the organization on the transcriptome. The outcomes indicated that the chromosome interaction pattern disturbed by vanillin stress or genetic mutations in the log phase was similar to that in mouse cells. The short chromosomes contact the short chromosomes, and the long chromosomes contact the long chromosomes. In response to vanillin stress, the boundaries of the topologically associating domain (TAD) in the vanillin-tolerant strain EMV-8 were more stable than those in its parent strain NAN-27. The motifs of SFL1, STB3, and NHP6A/B were enriched at TAD boundaries in both EMV-8 and NAN-27 with or without vanillin, indicating that these four genes were probably related to TAD formation. The Indel mutation of YRR1, whose absence was confirmed to benefit vanillin tolerance in EMV-8, caused two new interaction sites that contained three genes, WTM2, PUP1, and ALE1, whose overexpression did not affect vanillin resistance in yeast. Overall, our results revealed that in the log phase, genetic mutations and vanillin disturbance have a negligible effect on three-dimensional chromosome organization, and the reformation or disappearance of TAD boundaries did not show an association with gene expression, which provides an example for studying yeast chromatin structure during stress tolerance using Hi-C technology.

RevDate: 2022-12-06

Bi H, Hou Y, Wang J, et al (2022)

Chromatin reconstruction during mouse terminal erythropoiesis.

iScience, 25(12):105554.

Mammalian terminal erythropoiesis involves chromatin and nuclear condensation followed by enucleation. Late-stage erythroblasts undergo caspase-mediated nuclear opening that is important for nuclear condensation through partial histone release. It remains unknown the dynamic changes of three-dimensional (3D) genomic organization during terminal erythropoiesis. Here, we used Hi-C to determine the chromatin structural change during primary mouse erythroblast terminal differentiation. We also performed RNA-sequencing and ATAC-sequencing under the same experimental setting to further reveal the genome accessibility and gene expression changes during this process. We found that late-stage terminal erythropoiesis involves global loss of topologically associating domains and establishment of inter-chromosomal interactions of the heterochromatin regions, which are associated with globally increased chromatin accessibility and upregulation of erythroid-related genes.

RevDate: 2022-12-13
CmpDate: 2022-12-06

Zhang Y, M Blanchette (2022)

Reference panel guided topological structure annotation of Hi-C data.

Nature communications, 13(1):7426.

Accurately annotating topological structures (e.g., loops and topologically associating domains) from Hi-C data is critical for understanding the role of 3D genome organization in gene regulation. This is a challenging task, especially at high resolution, in part due to the limited sequencing coverage of Hi-C data. Current approaches focus on the analysis of individual Hi-C data sets of interest, without taking advantage of the facts that (i) several hundred Hi-C contact maps are publicly available, and (ii) the vast majority of topological structures are conserved across multiple cell types. Here, we present RefHiC, an attention-based deep learning framework that uses a reference panel of Hi-C datasets to facilitate topological structure annotation from a given study sample. We compare RefHiC against tools that do not use reference samples and find that RefHiC outperforms other programs at both topological associating domain and loop annotation across different cell types, species, and sequencing depths.

RevDate: 2023-01-21
CmpDate: 2023-01-20

Miyata M, Yoshida J, Takagishi I, et al (2023)

Comparison of CRISPR-Cas9-mediated megabase-scale genome deletion methods in mouse embryonic stem cells.

DNA research : an international journal for rapid publication of reports on genes and genomes, 30(1):.

The genome contains large functional units ranging in size from hundreds of kilobases to megabases, such as gene clusters and topologically associating domains. To analyse these large functional units, the technique of deleting the entire functional unit is effective. However, deletion of such large regions is less efficient than conventional genome editing, especially in cultured cells, and a method that can ensure success is anticipated. Here, we compared methods to delete the 2.5-Mb Krüppel-associated box zinc finger protein (KRAB-ZFP) gene cluster in mouse embryonic stem cells using CRISPR-Cas9. Three methods were used: first, deletion by non-homologous end joining (NHEJ); second, homology-directed repair (HDR) using a single-stranded oligodeoxynucleotide (ssODN); and third, HDR employing targeting vectors with a selectable marker and 1-kb homology arms. NHEJ-mediated deletion was achieved in 9% of the transfected cells. Inversion was also detected at similar efficiency. The deletion frequency of NHEJ and HDR was found to be comparable when the ssODN was transfected. Deletion frequency was highest when targeting vectors were introduced, with deletions occurring in 31-63% of the drug-resistant clones. Biallelic deletion was observed when targeting vectors were used. This study will serve as a benchmark for the introduction of large deletions into the genome.

RevDate: 2023-01-13
CmpDate: 2022-11-30

Wang Y, Mak TSH, Dattani S, et al (2022)

Whole genome sequencing reveals epistasis effects within RET for Hirschsprung disease.

Scientific reports, 12(1):20423.

Common variants in RET and NRG1 have been associated with Hirschsprung disease (HSCR), a congenital disorder characterised by incomplete innervation of distal gut, in East Asian (EA) populations. However, the allelic effects so far identified do not fully explain its heritability, suggesting the presence of epistasis, where effect of one genetic variant differs depending on other (modifier) variants. Few instances of epistasis have been documented in complex diseases due to modelling complexity and data challenges. We proposed four epistasis models to comprehensively capture epistasis for HSCR between and within RET and NRG1 loci using whole genome sequencing (WGS) data in EA samples. 65 variants within the Topologically Associating Domain (TAD) of RET demonstrated significant epistasis with the lead enhancer variant (RET+3; rs2435357). These epistatic variants formed two linkage disequilibrium (LD) clusters represented by rs2506026 and rs2506028 that differed in minor allele frequency and the best-supported epistatic model. Intriguingly, rs2506028 is in high LD with one cis-regulatory variant (rs2506030) highlighted previously, suggesting that detected epistasis might be mediated through synergistic effects on transcription regulation of RET. Our findings demonstrated the advantages of WGS data for detecting epistasis, and support the presence of interactive effects of regulatory variants in RET for HSCR.

RevDate: 2023-02-02
CmpDate: 2022-11-23

Zhao H, Yang M, Bishop J, et al (2022)

Identification and functional validation of super-enhancers in Arabidopsis thaliana.

Proceedings of the National Academy of Sciences of the United States of America, 119(48):e2215328119.

Super-enhancers (SEs) are exceptionally large enhancers and are recognized to play prominent roles in cell identity in mammalian species. We surveyed the genomic regions containing large clusters of accessible chromatin regions (ACRs) marked by deoxyribonuclease (DNase) I hypersensitivity in Arabidopsis thaliana. We identified a set of 749 putative SEs, which have a minimum length of 1.5 kilobases and represent the top 2.5% of the largest ACR clusters. We demonstrate that the genomic regions associating with these SEs were more sensitive to DNase I than other nonpromoter ACRs. The SEs were preferentially associated with topologically associating domains. Furthermore, the SEs and their predicted cognate genes were frequently associated with organ development and tissue identity in A. thaliana. Therefore, the A. thaliana SEs and their cognate genes mirror the functional characteristics of those reported in mammalian species. We developed CRISPR/Cas-mediated deletion lines of a 3,578-bp SE associated with the thalianol biosynthetic gene cluster (BGC). Small deletions (131-157 bp) within the SE resulted in distinct phenotypic changes and transcriptional repression of all five thalianol genes. In addition, T-DNA insertions in the SE region resulted in transcriptional alteration of all five thalianol genes. Thus, this SE appears to play a central role in coordinating the operon-like expression pattern of the thalianol BGC.

RevDate: 2022-11-10

Yancoskie MN, Maritz C, van Eijk P, et al (2022)

To incise or not and where: SET-domain methyltransferases know.

Trends in biochemical sciences pii:S0968-0004(22)00274-2 [Epub ahead of print].

The concept of the histone code posits that histone modifications regulate gene functions once interpreted by epigenetic readers. A well-studied case is trimethylation of lysine 4 of histone H3 (H3K4me3), which is enriched at gene promoters. However, H3K4me3 marks are not needed for the expression of most genes, suggesting extra roles, such as influencing the 3D genome architecture. Here, we highlight an intriguing analogy between the H3K4me3-dependent induction of double-strand breaks in several recombination events and the impact of this same mark on DNA incisions for the repair of bulky lesions. We propose that Su(var)3-9, Enhancer-of-zeste and Trithorax (SET)-domain methyltransferases generate H3K4me3 to guide nucleases into chromatin spaces, the favorable accessibility of which ensures that DNA break intermediates are readily processed, thereby safeguarding genome stability.

RevDate: 2022-12-22
CmpDate: 2022-11-08

Kim J, Jimenez DS, Ragipani B, et al (2022)

Condensin DC loads and spreads from recruitment sites to create loop-anchored TADs in C. elegans.

eLife, 11:.

Condensins are molecular motors that compact DNA via linear translocation. In Caenorhabditis elegans, the X-chromosome harbors a specialized condensin that participates in dosage compensation (DC). Condensin DC is recruited to and spreads from a small number of recruitment elements on the X-chromosome (rex) and is required for the formation of topologically associating domains (TADs). We take advantage of autosomes that are largely devoid of condensin DC and TADs to address how rex sites and condensin DC give rise to the formation of TADs. When an autosome and X-chromosome are physically fused, despite the spreading of condensin DC into the autosome, no TAD was created. Insertion of a strong rex on the X-chromosome results in the TAD boundary formation regardless of sequence orientation. When the same rex is inserted on an autosome, despite condensin DC recruitment, there was no spreading or features of a TAD. On the other hand, when a 'super rex' composed of six rex sites or three separate rex sites are inserted on an autosome, recruitment and spreading of condensin DC led to the formation of TADs. Therefore, recruitment to and spreading from rex sites are necessary and sufficient for recapitulating loop-anchored TADs observed on the X-chromosome. Together our data suggest a model in which rex sites are both loading sites and bidirectional barriers for condensin DC, a one-sided loop-extruder with movable inactive anchor.

RevDate: 2023-01-06
CmpDate: 2022-12-23

Attou A, Zülske T, G Wedemann (2022)

Cohesin and CTCF complexes mediate contacts in chromatin loops depending on nucleosome positions.

Biophysical journal, 121(24):4788-4799.

The spatial organization of the eukaryotic genome plays an important role in regulating transcriptional activity. In the nucleus, chromatin forms loops that assemble into fundamental units called topologically associating domains that facilitate or inhibit long-range contacts. These loops are formed and held together by the ring-shaped cohesin protein complex, and this can involve binding of CCCTC-binding factor (CTCF). High-resolution conformation capture experiments provide the frequency at which two DNA fragments physically associate in three-dimensional space. However, technical limitations of this approach, such as low throughput, low resolution, or noise in contact maps, make data interpretation and identification of chromatin intraloop contacts, e.g., between distal regulatory elements and their target genes, challenging. Herein, an existing coarse-grained model of chromatin at single-nucleosome resolution was extended by integrating potentials describing CTCF and cohesin. We performed replica-exchange Monte Carlo simulations with regularly spaced nucleosomes and experimentally determined nucleosome positions in the presence of cohesin-CTCF, as well as depleted systems as controls. In fully extruded loops caused by the presence of cohesin and CTCF, the number of contacts within the formed loops was increased. The number and types of these contacts were impacted by the nucleosome distribution and loop size. Microloops were observed within cohesin-mediated loops due to thermal fluctuations without additional influence of other factors, and the number, size, and shape of microloops were determined by nucleosome distribution and loop size. Nucleosome positions directly affect the spatial structure and contact probability within a loop, with presumed consequences for transcriptional activity.

RevDate: 2022-12-13
CmpDate: 2022-11-04

Zhu X, Qi C, Wang R, et al (2022)

Acute depletion of human core nucleoporin reveals direct roles in transcription control but dispensability for 3D genome organization.

Cell reports, 41(5):111576.

The nuclear pore complex (NPC) comprises more than 30 nucleoporins (NUPs) and is a hallmark of eukaryotes. NUPs have been suggested to be important in regulating gene transcription and 3D genome organization. However, evidence in support of their direct roles remains limited. Here, by Cut&Run, we find that core NUPs display broad but also cell-type-specific association with active promoters and enhancers in human cells. Auxin-mediated rapid depletion of two NUPs demonstrates that NUP93, but not NUP35, directly and specifically controls gene transcription. NUP93 directly activates genes with high levels of RNA polymerase II loading and transcriptional elongation by facilitating full BRD4 recruitment to their active enhancers. dCas9-based tethering confirms a direct and causal role of NUP93 in gene transcriptional activation. Unexpectedly, in situ Hi-C and H3K27ac or H3K4me1 HiChIP results upon acute NUP93 depletion show negligible changesS2211-1247(22)01437-1 of 3D genome organization ranging from A/B compartments and topologically associating domains (TADs) to enhancer-promoter contacts.

RevDate: 2023-01-28
CmpDate: 2023-01-18

Wang Y, Song C, Zhao J, et al (2023)

SEdb 2.0: a comprehensive super-enhancer database of human and mouse.

Nucleic acids research, 51(D1):D280-D290.

Super-enhancers (SEs) are cell-specific DNA cis-regulatory elements that can supervise the transcriptional regulation processes of downstream genes. SEdb 2.0 (http://www.licpathway.net/sedb) aims to provide a comprehensive SE resource and annotate their potential roles in gene transcriptions. Compared with SEdb 1.0, we have made the following improvements: (i) Newly added the mouse SEs and expanded the scale of human SEs. SEdb 2.0 contained 1 167 518 SEs from 1739 human H3K27ac chromatin immunoprecipitation sequencing (ChIP-seq) samples and 550 226 SEs from 931 mouse H3K27ac ChIP-seq samples, which was five times that of SEdb 1.0. (ii) Newly added transcription factor binding sites (TFBSs) in SEs identified by TF motifs and TF ChIP-seq data. (iii) Added comprehensive (epi)genetic annotations of SEs, including chromatin accessibility regions, methylation sites, chromatin interaction regions and topologically associating domains (TADs). (iv) Newly embedded and updated search and analysis tools, including 'Search SE by TF-based', 'Differential-Overlapping-SE analysis' and 'SE-based TF-Gene analysis'. (v) Newly provided quality control (QC) metrics for ChIP-seq processing. In summary, SEdb 2.0 is a comprehensive update of SEdb 1.0, which curates more SEs and annotation information than SEdb 1.0. SEdb 2.0 provides a friendly platform for researchers to more comprehensively clarify the important role of SEs in the biological process.

RevDate: 2022-12-24
CmpDate: 2022-11-02

Telonis AG, Yang Q, Huang HT, et al (2022)

MIR retrotransposons link the epigenome and the transcriptome of coding genes in acute myeloid leukemia.

Nature communications, 13(1):6524.

DNMT3A and IDH1/2 mutations combinatorically regulate the transcriptome and the epigenome in acute myeloid leukemia; yet the mechanisms of this interplay are unknown. Using a systems approach within topologically associating domains, we find that genes with significant expression-methylation correlations are enriched in signaling and metabolic pathways. The common denominator across these methylation-regulated genes is the density in MIR retrotransposons of their introns. Moreover, a discrete number of CpGs overlapping enhancers are responsible for regulating most of these genes. Established mouse models recapitulate the dependency of MIR-rich genes on the balanced expression of epigenetic modifiers, while projection of leukemic profiles onto normal hematopoiesis ones further consolidates the dependencies of methylation-regulated genes on MIRs. Collectively, MIR elements on genes and enhancers are susceptible to changes in DNA methylation activity and explain the cooperativity of proteins in this pathway in normal and malignant hematopoiesis.

RevDate: 2022-12-24
CmpDate: 2022-11-01

Schöpflin R, Melo US, Moeinzadeh H, et al (2022)

Integration of Hi-C with short and long-read genome sequencing reveals the structure of germline rearranged genomes.

Nature communications, 13(1):6470.

Structural variants are a common cause of disease and contribute to a large extent to inter-individual variability, but their detection and interpretation remain a challenge. Here, we investigate 11 individuals with complex genomic rearrangements including germline chromothripsis by combining short- and long-read genome sequencing (GS) with Hi-C. Large-scale genomic rearrangements are identified in Hi-C interaction maps, allowing for an independent assessment of breakpoint calls derived from the GS methods, resulting in >300 genomic junctions. Based on a comprehensive breakpoint detection and Hi-C, we achieve a reconstruction of whole rearranged chromosomes. Integrating information on the three-dimensional organization of chromatin, we observe that breakpoints occur more frequently than expected in lamina-associated domains (LADs) and that a majority reshuffle topologically associating domains (TADs). By applying phased RNA-seq, we observe an enrichment of genes showing allelic imbalanced expression (AIG) within 100 kb around the breakpoints. Interestingly, the AIGs hit by a breakpoint (19/22) display both up- and downregulation, thereby suggesting different mechanisms at play, such as gene disruption and rearrangements of regulatory information. However, the majority of interpretable genes located 200 kb around a breakpoint do not show significant expression changes. Thus, there is an overall robustness in the genome towards large-scale chromosome rearrangements.

RevDate: 2022-12-22
CmpDate: 2022-11-18

Xu J, Song F, Lyu H, et al (2022)

Subtype-specific 3D genome alteration in acute myeloid leukaemia.

Nature, 611(7935):387-398.

Acute myeloid leukaemia (AML) represents a set of heterogeneous myeloid malignancies, and hallmarks include mutations in epigenetic modifiers, transcription factors and kinases[1-5]. The extent to which mutations in AML drive alterations in chromatin 3D structure and contribute to myeloid transformation is unclear. Here we use Hi-C and whole-genome sequencing to analyse 25 samples from patients with AML and 7 samples from healthy donors. Recurrent and subtype-specific alterations in A/B compartments, topologically associating domains and chromatin loops were identified. RNA sequencing, ATAC with sequencing and CUT&Tag for CTCF, H3K27ac and H3K27me3 in the same AML samples also revealed extensive and recurrent AML-specific promoter-enhancer and promoter-silencer loops. We validated the role of repressive loops on their target genes by CRISPR deletion and interference. Structural variation-induced enhancer-hijacking and silencer-hijacking events were further identified in AML samples. Hijacked enhancers play a part in AML cell growth, as demonstrated by CRISPR screening, whereas hijacked silencers have a downregulating role, as evidenced by CRISPR-interference-mediated de-repression. Finally, whole-genome bisulfite sequencing of 20 AML and normal samples revealed the delicate relationship between DNA methylation, CTCF binding and 3D genome structure. Treatment of AML cells with a DNA hypomethylating agent and triple knockdown of DNMT1, DNMT3A and DNMT3B enabled the manipulation of DNA methylation to revert 3D genome organization and gene expression. Overall, this study provides a resource for leukaemia studies and highlights the role of repressive loops and hijacked cis elements in human diseases.

RevDate: 2023-01-24
CmpDate: 2022-11-10

Mohajeri K, Yadav R, D'haene E, et al (2022)

Transcriptional and functional consequences of alterations to MEF2C and its topological organization in neuronal models.

American journal of human genetics, 109(11):2049-2067.

Point mutations and structural variants that directly disrupt the coding sequence of MEF2C have been associated with a spectrum of neurodevelopmental disorders (NDDs). However, the impact of MEF2C haploinsufficiency on neurodevelopmental pathways and synaptic processes is not well understood, nor are the complex mechanisms that govern its regulation. To explore the functional changes associated with structural variants that alter MEF2C expression and/or regulation, we generated an allelic series of 204 isogenic human induced pluripotent stem cell (hiPSC)-derived neural stem cells and glutamatergic induced neurons. These neuronal models harbored CRISPR-engineered mutations that involved direct deletion of MEF2C or deletion of the boundary points for topologically associating domains (TADs) and chromatin loops encompassing MEF2C. Systematic profiling of mutation-specific alterations, contrasted to unedited controls that were exposed to the same guide RNAs for each edit, revealed that deletion of MEF2C caused differential expression of genes associated with neurodevelopmental pathways and synaptic function. We also discovered significant reduction in synaptic activity measured by multielectrode arrays (MEAs) in neuronal cells. By contrast, we observed robust buffering against MEF2C regulatory disruption following deletion of a distal 5q14.3 TAD and loop boundary, whereas homozygous loss of a proximal loop boundary resulted in down-regulation of MEF2C expression and reduced electrophysiological activity on MEA that was comparable to direct gene disruption. Collectively, these studies highlight the considerable functional impact of MEF2C deletion in neuronal cells and systematically characterize the complex interactions that challenge a priori predictions of regulatory consequences from structural variants that disrupt three-dimensional genome organization.

RevDate: 2023-01-07
CmpDate: 2022-11-09

Torosin NS, Golla TR, Lawlor MA, et al (2022)

Mode and Tempo of 3D Genome Evolution in Drosophila.

Molecular biology and evolution, 39(11):.

Topologically associating domains (TADs) are thought to play an important role in preventing gene misexpression by spatially constraining enhancer-promoter contacts. The deleterious nature of gene misexpression implies that TADs should, therefore, be conserved among related species. Several early studies comparing chromosome conformation between species reported high levels of TAD conservation; however, more recent studies have questioned these results. Furthermore, recent work suggests that TAD reorganization is not associated with extensive changes in gene expression. Here, we investigate the evolutionary conservation of TADs among 11 species of Drosophila. We use Hi-C data to identify TADs in each species and employ a comparative phylogenetic approach to derive empirical estimates of the rate of TAD evolution. Surprisingly, we find that TADs evolve rapidly. However, we also find that the rate of evolution depends on the chromatin state of the TAD, with TADs enriched for developmentally regulated chromatin evolving significantly slower than TADs enriched for broadly expressed, active chromatin. We also find that, after controlling for differences in chromatin state, highly conserved TADs do not exhibit higher levels of gene expression constraint. These results suggest that, in general, most TADs evolve rapidly and their divergence is not associated with widespread changes in gene expression. However, higher levels of evolutionary conservation and gene expression constraints in TADs enriched for developmentally regulated chromatin suggest that these TAD subtypes may be more important for regulating gene expression, likely due to the larger number of long-distance enhancer-promoter contacts associated with developmental genes.

RevDate: 2022-10-24
CmpDate: 2022-10-19

Simmons JR, An R, Amankwaa B, et al (2022)

Phosphorylated histone variant γH2Av is associated with chromatin insulators in Drosophila.

PLoS genetics, 18(10):e1010396.

Chromatin insulators are responsible for orchestrating long-range interactions between enhancers and promoters throughout the genome and align with the boundaries of Topologically Associating Domains (TADs). Here, we demonstrate an association between gypsy insulator proteins and the phosphorylated histone variant H2Av (γH2Av), normally a marker of DNA double strand breaks. Gypsy insulator components colocalize with γH2Av throughout the genome, in polytene chromosomes and in diploid cells in which Chromatin IP data shows it is enriched at TAD boundaries. Mutation of insulator components su(Hw) and Cp190 results in a significant reduction in γH2Av levels in chromatin and phosphatase inhibition strengthens the association between insulator components and γH2Av and rescues γH2Av localization in insulator mutants. We also show that γH2Av, but not H2Av, is a component of insulator bodies, which are protein condensates that form during osmotic stress. Phosphatase activity is required for insulator body dissolution after stress recovery. Together, our results implicate the H2A variant with a novel mechanism of insulator function and boundary formation.

RevDate: 2022-12-21
CmpDate: 2022-12-16

Jouret G, Egloff M, Landais E, et al (2023)

Clinical and genomic delineation of the new proximal 19p13.3 microduplication syndrome.

American journal of medical genetics. Part A, 191(1):52-63.

A small but growing body of scientific literature is emerging about clinical findings in patients with 19p13.3 microdeletion or duplication. Recently, a proximal 19p13.3 microduplication syndrome was described, associated with growth delay, microcephaly, psychomotor delay and dysmorphic features. The aim of our study was to better characterize the syndrome associated with duplications in the proximal 19p13.3 region (prox 19p13.3 dup), and to propose a comprehensive analysis of the underlying genomic mechanism. We report the largest cohort of patients with prox 19p13.3 dup through a collaborative study. We collected 24 new patients with terminal or interstitial 19p13.3 duplication characterized by array-based Comparative Genomic Hybridization (aCGH). We performed mapping, phenotype-genotype correlations analysis, critical region delineation and explored three-dimensional chromatin interactions by analyzing Topologically Associating Domains (TADs). We define a new 377 kb critical region (CR 1) in chr19: 3,116,922-3,494,377, GRCh37, different from the previously described critical region (CR 2). The new 377 kb CR 1 includes a TAD boundary and two enhancers whose common target is PIAS4. We hypothesize that duplications of CR 1 are responsible for tridimensional structural abnormalities by TAD disruption and misregulation of genes essentials for the control of head circumference during development, by breaking down the interactions between enhancers and the corresponding targeted gene.

RevDate: 2023-01-25
CmpDate: 2022-10-04

Ringel AR, Szabo Q, Chiariello AM, et al (2022)

Repression and 3D-restructuring resolves regulatory conflicts in evolutionarily rearranged genomes.

Cell, 185(20):3689-3704.e21.

Regulatory landscapes drive complex developmental gene expression, but it remains unclear how their integrity is maintained when incorporating novel genes and functions during evolution. Here, we investigated how a placental mammal-specific gene, Zfp42, emerged in an ancient vertebrate topologically associated domain (TAD) without adopting or disrupting the conserved expression of its gene, Fat1. In ESCs, physical TAD partitioning separates Zfp42 and Fat1 with distinct local enhancers that drive their independent expression. This separation is driven by chromatin activity and not CTCF/cohesin. In contrast, in embryonic limbs, inactive Zfp42 shares Fat1's intact TAD without responding to active Fat1 enhancers. However, neither Fat1 enhancer-incompatibility nor nuclear envelope-attachment account for Zfp42's unresponsiveness. Rather, Zfp42's promoter is rendered inert to enhancers by context-dependent DNA methylation. Thus, diverse mechanisms enabled the integration of independent Zfp42 regulation in the Fat1 locus. Critically, such regulatory complexity appears common in evolution as, genome wide, most TADs contain multiple independently expressed genes.

RevDate: 2022-11-28
CmpDate: 2022-09-28

Martinez-Fundichely A, Dixon A, E Khurana (2022)

Modeling tissue-specific breakpoint proximity of structural variations from whole-genomes to identify cancer drivers.

Nature communications, 13(1):5640.

Structural variations (SVs) in cancer cells often impact large genomic regions with functional consequences. However, identification of SVs under positive selection is a challenging task because little is known about the genomic features related to the background breakpoint distribution in different cancers. We report a method that uses a generalized additive model to investigate the breakpoint proximity curves from 2,382 whole-genomes of 32 cancer types. We find that a multivariate model, which includes linear and nonlinear partial contributions of various tissue-specific features and their interaction terms, can explain up to 57% of the observed deviance of breakpoint proximity. In particular, three-dimensional genomic features such as topologically associating domains (TADs), TAD-boundaries and their interaction with other features show significant contributions. The model is validated by identification of known cancer genes and revealed putative drivers in cancers different than those with previous evidence of positive selection.

RevDate: 2022-10-25
CmpDate: 2022-09-28

Damas J, Corbo M, Kim J, et al (2022)

Evolution of the ancestral mammalian karyotype and syntenic regions.

Proceedings of the National Academy of Sciences of the United States of America, 119(40):e2209139119.

Decrypting the rearrangements that drive mammalian chromosome evolution is critical to understanding the molecular bases of speciation, adaptation, and disease susceptibility. Using 8 scaffolded and 26 chromosome-scale genome assemblies representing 23/26 mammal orders, we computationally reconstructed ancestral karyotypes and syntenic relationships at 16 nodes along the mammalian phylogeny. Three different reference genomes (human, sloth, and cattle) representing phylogenetically distinct mammalian superorders were used to assess reference bias in the reconstructed ancestral karyotypes and to expand the number of clades with reconstructed genomes. The mammalian ancestor likely had 19 pairs of autosomes, with nine of the smallest chromosomes shared with the common ancestor of all amniotes (three still conserved in extant mammals), demonstrating a striking conservation of synteny for ∼320 My of vertebrate evolution. The numbers and types of chromosome rearrangements were classified for transitions between the ancestral mammalian karyotype, descendent ancestors, and extant species. For example, 94 inversions, 16 fissions, and 14 fusions that occurred over 53 My differentiated the therian from the descendent eutherian ancestor. The highest breakpoint rate was observed between the mammalian and therian ancestors (3.9 breakpoints/My). Reconstructed mammalian ancestor chromosomes were found to have distinct evolutionary histories reflected in their rates and types of rearrangements. The distributions of genes, repetitive elements, topologically associating domains, and actively transcribed regions in multispecies homologous synteny blocks and evolutionary breakpoint regions indicate that purifying selection acted over millions of years of vertebrate evolution to maintain syntenic relationships of developmentally important genes and regulatory landscapes of gene-dense chromosomes.

RevDate: 2022-09-24

Chow CN, Tseng KC, Hou PF, et al (2022)

Mysteries of gene regulation: Promoters are not the sole triggers of gene expression.

Computational and structural biotechnology journal, 20:4910-4920.

Cis-regulatory elements of promoters are essential for gene regulation by transcription factors (TFs). However, the regulatory roles of nonpromoter regions, TFs, and epigenetic marks remain poorly understood in plants. In this study, we characterized the cis-regulatory regions of 53 TFs and 19 histone marks in 328 chromatin immunoprecipitation (ChIP-seq) datasets from Arabidopsis. The genome-wide maps indicated that both promoters and regions around the transcription termination sites of protein-coding genes recruit the most TFs. The maps also revealed a diverse of histone combinations. The analysis suggested that exons play critical roles in the regulation of non-coding genes. Additionally, comparative analysis between heat-stress-responsive and nonresponsive genes indicated that the genes with distinct functions also exhibited substantial differences in cis-regulatory regions, histone regulation, and topologically associating domain (TAD) boundary organization. By integrating multiple high-throughput sequencing datasets, this study generated regulatory models for protein-coding genes, non-coding genes, and TAD boundaries to explain the complexity of transcriptional regulation.

RevDate: 2022-10-25

Doyle EJ, Morey L, E Conway (2022)

Know when to fold 'em: Polycomb complexes in oncogenic 3D genome regulation.

Frontiers in cell and developmental biology, 10:986319.

Chromatin is spatially and temporally regulated through a series of orchestrated processes resulting in the formation of 3D chromatin structures such as topologically associating domains (TADs), loops and Polycomb Bodies. These structures are closely linked to transcriptional regulation, with loss of control of these processes a frequent feature of cancer and developmental syndromes. One such oncogenic disruption of the 3D genome is through recurrent dysregulation of Polycomb Group Complex (PcG) functions either through genetic mutations, amplification or deletion of genes that encode for PcG proteins. PcG complexes are evolutionarily conserved epigenetic complexes. They are key for early development and are essential transcriptional repressors. PcG complexes include PRC1, PRC2 and PR-DUB which are responsible for the control of the histone modifications H2AK119ub1 and H3K27me3. The spatial distribution of the complexes within the nuclear environment, and their associated modifications have profound effects on the regulation of gene transcription and the 3D genome. Nevertheless, how PcG complexes regulate 3D chromatin organization is still poorly understood. Here we glean insights into the role of PcG complexes in 3D genome regulation and compaction, how these processes go awry during tumorigenesis and the therapeutic implications that result from our insights into these mechanisms.

RevDate: 2022-11-19
CmpDate: 2022-09-16

Götz M, Messina O, Espinola S, et al (2022)

Multiple parameters shape the 3D chromatin structure of single nuclei at the doc locus in Drosophila.

Nature communications, 13(1):5375.

The spatial organization of chromatin at the scale of topologically associating domains (TADs) and below displays large cell-to-cell variations. Up until now, how this heterogeneity in chromatin conformation is shaped by chromatin condensation, TAD insulation, and transcription has remained mostly elusive. Here, we used Hi-M, a multiplexed DNA-FISH imaging technique providing developmental timing and transcriptional status, to show that the emergence of TADs at the ensemble level partially segregates the conformational space explored by single nuclei during the early development of Drosophila embryos. Surprisingly, a substantial fraction of nuclei display strong insulation even before TADs emerge. Moreover, active transcription within a TAD leads to minor changes to the local inter- and intra-TAD chromatin conformation in single nuclei and only weakly affects insulation to the neighboring TAD. Overall, our results indicate that multiple parameters contribute to shaping the chromatin architecture of single nuclei at the TAD scale.

RevDate: 2022-11-19
CmpDate: 2022-09-28

Kane L, Williamson I, Flyamer IM, et al (2022)

Cohesin is required for long-range enhancer action at the Shh locus.

Nature structural & molecular biology, 29(9):891-897.

The regulatory landscapes of developmental genes in mammals can be complex, with enhancers spread over many hundreds of kilobases. It has been suggested that three-dimensional genome organization, particularly topologically associating domains formed by cohesin-mediated loop extrusion, is important for enhancers to act over such large genomic distances. By coupling acute protein degradation with synthetic activation by targeted transcription factor recruitment, here we show that cohesin, but not CTCF, is required for activation of the target gene Shh by distant enhancers in mouse embryonic stem cells. Cohesin is not required for activation directly at the promoter or by an enhancer located closer to the Shh gene. Our findings support the hypothesis that chromatin compaction via cohesin-mediated loop extrusion allows for genes to be activated by enhancers that are located many hundreds of kilobases away in the linear genome and suggests that cohesin is dispensable for enhancers located more proximally.

RevDate: 2022-11-01
CmpDate: 2022-09-13

Zheng L, W Wang (2022)

Regulation associated modules reflect 3D genome modularity associated with chromatin activity.

Nature communications, 13(1):5281.

The 3D genome has been shown to be organized into modules including topologically associating domains (TADs) and compartments that are primarily defined by spatial contacts from Hi-C. There exists a gap to investigate whether and how the spatial modularity of the chromatin is related to the functional modularity resulting from chromatin activity. Despite histone modifications reflecting chromatin activity, inferring spatial modularity of the genome directly from the histone modification patterns has not been well explored. Here, we report that histone modifications show a modular pattern (referred to as regulation associated modules, RAMs) that reflects spatial chromatin modularity. Enhancer-promoter interactions, loop anchors, super-enhancer clusters and extrachromosomal DNAs (ecDNAs) are found to occur more often within the same RAMs than within the same TADs. Consistently, compared to the TAD boundaries, deletions of RAM boundaries perturb the chromatin structure more severely (may even cause cell death) and somatic variants in cancer samples are more enriched in RAM boundaries. These observations suggest that RAMs reflect a modular organization of the 3D genome at a scale better aligned with chromatin activity, providing a bridge connecting the structural and functional modularity of the genome.

RevDate: 2022-09-06

Sun Y, Dotson GA, Muir LA, et al (2022)

Rearrangement of T Cell genome architecture regulates GVHD.

iScience, 25(9):104846.

WAPL, cohesin's DNA release factor, regulates three-dimensional (3D) chromatin architecture. The 3D chromatin structure and its relevance to mature T cell functions is not well understood. We show that in vivo lymphopenic expansion, and alloantigen-driven proliferation, alters the 3D structure and function of the genome in mature T cells. Conditional deletion of WAPL, cohesin's DNA release factor, in T cells reduced long-range genomic interactions and altered chromatin A/B compartments and interactions within topologically associating domains (TADs) of the chromatin in T cells at baseline. WAPL deficiency in T cells reduced loop extensions, changed expression of cell cycling genes and reduced proliferation following in vitro and in vivo stimulation, and reduced severity of graft-versus-host disease (GVHD) following experimental allogeneic hematopoietic stem cell transplantation. These data collectively characterize 3D genomic architecture of T cells in vivo and demonstrate biological and clinical implications for its disruption by cohesin release factor WAPL.

RevDate: 2022-12-22
CmpDate: 2022-10-27

Che Y, Yang X, Jia P, et al (2022)

D[2] Plot, a Matrix of DNA Density and Distance to Periphery, Reveals Functional Genome Regions.

Advanced science (Weinheim, Baden-Wurttemberg, Germany), 9(30):e2202149.

The execution of biological activities inside space-limited cell nuclei requires sophisticated organization. Current studies on the 3D genome focus on chromatin interactions and local structures, e.g., topologically associating domains (TADs). In this study, two global physical properties: DNA density and distance to nuclear periphery (DisTP), are introduced and a 2D matrix, D[2] plot, is constructed for mapping genetic and epigenetic markers. Distinct patterns of functional markers on the D[2] plot, indicating its ability to compartmentalize functional genome regions, are observed. Furthermore, enrichments of transcription-related markers are concatenated into a cross-species transcriptional activation model, where the nucleus is divided into four areas: active, intermediate, repress and histone, and repress and repeat. Based on the trajectories of the genomic regions on D[2] plot, the constantly active and newly activated genes are successfully identified during olfactory sensory neuron maturation. The analysis reveals that the D[2] plot effectively categorizes functional regions and provides a universal and transcription-related measurement for the 3D genome.

RevDate: 2022-11-03
CmpDate: 2022-10-06

Schwartz M (2022)

Can abnormal chromatin folding cause high-penetrance cancer predisposition?.

Physiological genomics, 54(10):380-388.

Sequencing cancer predisposing genes (CPGs) in evocative patients (i.e., patients with personal and family history of multiple/early-onset/unusual cancers) allows follow-up in their relatives to be adapted when a causative pathogenic variant is identified. Unfortunately, many evocative families remain unexplained. Part of this "missing heritability" could be due to CPG dysregulations caused by remote noncoding genomic alterations. Transcription levels are regulated through the ability of promoters to physically interact with their distant cis-regulatory elements. Three-dimensional chromatin contacts, mediated by a dynamic loop extrusion process, are uncovered by chromosome conformation capture (3C) and 3C-derived techniques, which have enabled the discovery of new pathological mechanisms in developmental diseases and cancers. High-penetrance cancer predisposition is caused by germline hereditary alterations otherwise found at the somatic level in sporadic cancers. Thus, data from both developmental diseases and cancers provide information about possible unknown cancer predisposition mechanisms. This mini-review aims to deduce from these data whether abnormal chromatin folding can cause high-penetrance cancer predisposition.

RevDate: 2022-11-22
CmpDate: 2022-11-22

Kulikova T, Maslova A, Starshova P, et al (2022)

Comparison of the somatic TADs and lampbrush chromomere-loop complexes in transcriptionally active prophase I oocytes.

Chromosoma, 131(4):207-223.

In diplotene oocyte nuclei of all vertebrate species, except mammals, chromosomes lack interchromosomal contacts and chromatin is linearly compartmentalized into distinct chromomere-loop complexes forming lampbrush chromosomes. However, the mechanisms underlying the formation of chromomere-loop complexes remain unexplored. Here we aimed to compare somatic topologically associating domains (TADs), recently identified in chicken embryonic fibroblasts, with chromomere-loop complexes in lampbrush meiotic chromosomes. By measuring 3D-distances and colocalization between linear equidistantly located genomic loci, positioned within one TAD or separated by a TAD border, we confirmed the presence of predicted TADs in chicken embryonic fibroblast nuclei. Using three-colored FISH with BAC probes, we mapped equidistant genomic regions included in several sequential somatic TADs on isolated chicken lampbrush chromosomes. Eight genomic regions, each comprising two or three somatic TADs, were mapped to non-overlapping neighboring lampbrush chromatin domains - lateral loops, chromomeres, or chromomere-loop complexes. Genomic loci from the neighboring somatic TADs could localize in one lampbrush chromomere-loop complex, while genomic loci belonging to the same somatic TAD could be localized in neighboring lampbrush chromomere-loop domains. In addition, FISH-mapping of BAC probes to the nascent transcripts on the lateral loops indicates transcription of at least 17 protein-coding genes and 2 non-coding RNA genes during the lampbrush stage of chicken oogenesis, including genes involved in oocyte maturation and early embryo development.

RevDate: 2022-08-26

Wang X, Yan J, Ye Z, et al (2022)

Reorganization of 3D chromatin architecture in doxorubicin-resistant breast cancer cells.

Frontiers in cell and developmental biology, 10:974750.

Background: Doxorubicin resistance remains a major therapeutic challenge leading to poor survival prognosis and treatment failure in breast cancer. Although doxorubicin induces massive changes in the transcriptional landscape are well known, potential diagnostic or therapeutic targets associated with the reorganization of three-dimensional (3D) chromatin architecture have not yet been systematically investigated. Methods: Here we performed in situ high-throughput chromosome conformation capture (Hi-C) on parental and doxorubicin-resistant MCF7 (MCF7-DR) human breast cancer cells, followed by integrative analysis of HiC, ATAC-seq, RNA-seq and TCGA data. Results: It revealed that A/B compartment switching was positively correlated to genome-wide differential gene expression. The genome of MCF7-DR cells was spatially reorganized into smaller topologically associating domains (TADs) and chromatin loops. We also revealed the contribution of increased chromatin accessibility and potential transcription factor families, including CTCF, AP-1 and bHLH, to gained TADs or loops. Intriguingly, we observed two condensed genomic regions (∼20 kb) with decreased chromatin accessibility flanking TAD boundaries, which might play a critical role in the formation or maintenance of TADs. Finally, combining data from TCGA, we identified a number of gained and lost enhancer-promoter interactions and their corresponding differentially expressed genes involved in chromatin organization and breast cancer signaling pathways, including FA2H, FOXA1 and JRKL, which might serve as potential treatment targets for breast cancer. Conclusion: These data uncovered a close connection between 3D genome reorganization, chromatin accessibility as well as gene transcription and provide novel insights into the epigenomic mechanisms involving doxorubicin resistance in breast cancer.

RevDate: 2022-11-08
CmpDate: 2022-10-05

Chen M, Jia L, Zheng X, et al (2022)

Ancient Human Endogenous Retroviruses Contribute to Genetic Evolution and Regulate Cancer Cell Type-Specific Gene Expression.

Cancer research, 82(19):3457-3473.

UNLABELLED: Human endogenous retroviruses (HERV), a type of transposable elements (TE), play crucial roles in human placental morphogenesis, immune response, and cancer progression. Emerging evidence suggests that TEs have been a rich source of regulatory elements in the human genome, but little is known about the global impact of HERVs on transcriptional networks in cancer. Using genome-wide approaches, we show that HERVs are composed primarily of three ancient superfamilies: ERVL-MaLR, ERVL, and ERV1. This analysis suggests that the integration of exonic, intronic, and intergenic HERVs, as well as human or Hominidae gene-specific HERVs, contributes to human genomic innovation. HERVs exonized in genes are located mainly in the 3' untranslated region (UTR) or 3' end and participate in basic biological processes. Active HERVs are located mainly in intronic and intergenic regions and tend to function as enhancers and contribute to cancer cell type-specific gene expression. More importantly, HERVs may also define chromatin topologically associating domain (TAD) and loop boundaries in a cell type-specific manner. Taken together, these findings reveal that ancient HERV elements are a source of diverse regulatory sequences, including 3' UTRs, 5' UTRs, promoters, and enhancers, and they contribute to genetic innovation and cancer cell type-specific gene expression, highlighting the previously underestimated importance of these elements.

SIGNIFICANCE: Genome-wide analyses show that human endogenous retroviruses mediate cancer cell type-specific gene expression, epigenetic modification, and 3D chromatin architecture, elucidating the relationship between HERVs and diverse cancers.

RevDate: 2022-10-05
CmpDate: 2022-08-17

Kurotaki D, Kikuchi K, Cui K, et al (2022)

Chromatin structure undergoes global and local reorganization during murine dendritic cell development and activation.

Proceedings of the National Academy of Sciences of the United States of America, 119(34):e2207009119.

Classical dendritic cells (cDCs) are essential for immune responses and differentiate from hematopoietic stem cells via intermediate progenitors, such as monocyte-DC progenitors (MDPs) and common DC progenitors (CDPs). Upon infection, cDCs are activated and rapidly express host defense-related genes, such as those encoding cytokines and chemokines. Chromatin structures, including nuclear compartments and topologically associating domains (TADs), have been implicated in gene regulation. However, the extent and dynamics of their reorganization during cDC development and activation remain unknown. In this study, we comprehensively determined higher-order chromatin structures by Hi-C in DC progenitors and cDC subpopulations. During cDC differentiation, chromatin activation was initially induced at the MDP stage. Subsequently, a shift from inactive to active nuclear compartments occurred at the cDC gene loci in CDPs, which was followed by increased intra-TAD interactions and loop formation. Mechanistically, the transcription factor IRF8, indispensable for cDC differentiation, mediated chromatin activation and changes into the active compartments in DC progenitors, thereby possibly leading to cDC-specific gene induction. Using an infection model, we found that the chromatin structures of host defense-related gene loci were preestablished in unstimulated cDCs, indicating that the formation of higher-order chromatin structures prior to infection may contribute to the rapid responses to pathogens. Overall, these results suggest that chromatin structure reorganization is closely related to the establishment of cDC-specific gene expression and immune functions. This study advances the fundamental understanding of chromatin reorganization in cDC differentiation and activation.

RevDate: 2022-08-16

Zhou T, Q Feng (2022)

Androgen receptor signaling and spatial chromatin organization in castration-resistant prostate cancer.

Frontiers in medicine, 9:924087.

Prostate cancer is one of the leading causes of cancer death and affects millions of men in the world. The American Cancer Society estimated about 34,500 deaths from prostate cancer in the United States in year 2022. The Androgen receptor (AR) signaling is a major pathway that sustains local and metastatic prostate tumor growth. Androgen-deprivation therapy (ADT) is the standard of care for metastatic prostate cancer patient and can suppress the tumor growth for a median of 2-3 years. Unfortunately, the malignancy inevitably progresses to castration-resistant prostate cancer (CRPC) which is more aggressive and no longer responsive to ADT. Surprisingly, for most of the CPRC patients, cancer growth still depends on androgen receptor signaling. Accumulating evidence suggests that CRPC cells have rewired their transcriptional program to retain AR signaling in the absence of androgens. Besides AR, other transcription factors also contribute to the resistance mechanism through multiple pathways including enhancing AR signaling pathway and activating other complementary signaling pathways for the favor of AR downstream genes expression. More recent studies have shown the role of transcription factors in reconfiguring chromatin 3D structure and regulating topologically associating domains (TADs). Pioneer factors, transcription factors and coactivators form liquid-liquid phase separation compartment that can modulate transcriptional events along with configuring TADs. The role of AR and other transcription factors on chromatin structure change and formation of condensate compartment in prostate cancer cells has only been recently investigated and appreciated. This review intends to provide an overview of transcription factors that contribute to AR signaling through activation of gene expression, governing 3D chromatin structure and establishing phase to phase separation. A more detailed understanding of the spatial role of transcription factors in CRPC might provide novel therapeutic targets for the treatment of CRPC.

RevDate: 2022-08-31
CmpDate: 2022-08-16

Giaimo BD, T Borggrefe (2022)

Enhancer-promoter communication: unraveling enhancer strength and positioning within a given topologically associating domain (TAD).

Signal transduction and targeted therapy, 7(1):281.

RevDate: 2022-10-15

Fang K, Wang J, Liu L, et al (2022)

Mapping nucleosome and chromatin architectures: A survey of computational methods.

Computational and structural biotechnology journal, 20:3955-3962.

With ever-growing genomic sequencing data, the data variabilities and the underlying biases of the sequencing technologies pose significant computational challenges ranging from the need for accurately detecting the nucleosome positioning or chromatin interaction to the need for developing normalization methods to eliminate systematic biases. This review mainly surveys the computational methods for mapping the higher-resolution nucleosome and higher-order chromatin architectures. While a detailed discussion of the underlying algorithms is beyond the scope of our survey, we have discussed the methods and tools that can detect the nucleosomes in the genome, then demonstrated the computational methods for identifying 3D chromatin domains and interactions. We further illustrated computational approaches for integrating multi-omics data with Hi-C data and the advance of single-cell (sc)Hi-C data analysis. Our survey provides a comprehensive and valuable resource for biomedical scientists interested in studying nucleosome organization and chromatin structures as well as for computational scientists who are interested in improving upon them.

RevDate: 2022-08-09

Ahn J, Lee J, Kim DH, et al (2022)

Loss of Monoallelic Expression of IGF2 in the Adult Liver Via Alternative Promoter Usage and Chromatin Reorganization.

Frontiers in genetics, 13:920641.

In mammals, genomic imprinting operates via gene silencing mechanisms. Although conservation of the imprinting mechanism at the H19/IGF2 locus has been generally described in pigs, tissue-specific imprinting at the transcript level, monoallelic-to-biallelic conversion, and spatio-temporal chromatin reorganization remain largely uninvestigated. Here, we delineate spatially regulated imprinting of IGF2 transcripts, age-dependent hepatic mono- to biallelic conversion, and reorganization of topologically associating domains at the porcine H19/IGF2 locus for better translation to human and animal research. Whole-genome bisulfite sequencing (WGBS) and RNA sequencing (RNA-seq) of normal and parthenogenetic porcine embryos revealed the paternally hypermethylated H19 differentially methylated region and paternal expression of IGF2. Using a polymorphism-based approach and omics datasets from chromatin immunoprecipitation sequencing (ChIP-seq), whole-genome sequencing (WGS), RNA-seq, and Hi-C, regulation of IGF2 during development was analyzed. Regulatory elements in the liver were distinguished from those in the muscle where the porcine IGF2 transcript was monoallelically expressed. The IGF2 transcript from the liver was biallelically expressed at later developmental stages in both pigs and humans. Chromatin interaction was less frequent in the adult liver compared to the fetal liver and skeletal muscle. The duration of genomic imprinting effects within the H19/IGF2 locus might be reduced in the liver with biallelic conversion through alternative promoter usage and chromatin remodeling. Our integrative omics analyses of genome, epigenome, and transcriptome provided a comprehensive view of imprinting status at the H19/IGF2 cluster.

RevDate: 2022-12-27
CmpDate: 2022-12-15

Sun Z, Wang Y, Song Z, et al (2022)

DNA methylation in transposable elements buffers the connection between three-dimensional chromatin organization and gene transcription upon rice genome duplication.

Journal of advanced research, 42:41-53.

INTRODUCTION: Polyploidy is a major force in plant evolution and the domestication of cultivated crops.

OBJECTIVES: The study aimed to explore the relationship and underlying mechanism between three-dimensional (3D) chromatin organization and gene transcription upon rice genome duplication.

METHODS: The 3D chromatin structures between diploid (2C) and autotetraploid (4C) rice were compared using high-throughput chromosome conformation capture (Hi-C) analysis. The study combined genetics, transcriptomics, whole-genome bisulfite sequencing (WGBS-seq) and 3D genomics approaches to uncover the mechanism for DNA methylation in modulating gene transcription through 3D chromatin architectures upon rice genome duplication.

RESULTS: We found that 4C rice presents weakened intra-chromosomal interactions compared to its 2C progenitor in some chromosomes. In addition, we found that changes of 3D chromatin organizations including chromatin compartments, topologically associating domains (TADs), and loops, are uncorrelated with gene transcription. Moreover, DNA methylations in the regulatory sequences of genes in compartment A/B switched regions and TAD boundaries are unrelated to their expression. Importantly, although there was no significant difference in the methylation levels in transposable elements (TEs) in differentially expressed gene (DEG) and non-DEG promoters between 2C and 4C rice, we found that the hypermethylated TEs across genes in compartment A/B switched regions and TAD boundaries may suppress the expression of these genes.

CONCLUSION: The study proposed that the rice genome doubling might modulate TE methylation to buffer the effects of chromatin architecture on gene transcription in compartment A/B switched regions and TAD boundaries, resulting in the disconnection between 3D chromatin structure alteration and gene transcription upon rice genome duplication.

RevDate: 2022-08-09

Li D, Strong A, Hou C, et al (2022)

Interstitial deletion 4p15.32p16.1 and complex chromoplexy in a female proband with severe neurodevelopmental delay, growth failure and dysmorphism.

Molecular cytogenetics, 15(1):33.

Complex chromosomal rearrangements involve the restructuring of genetic material within a single chromosome or across multiple chromosomes. These events can cause serious human disease by disrupting coding DNA and gene regulatory elements via deletions, duplications, and structural rearrangements. Here we describe a 5-year-old female with severe developmental delay, dysmorphic features, multi-suture craniosynostosis, and growth failure found to have a complex series of balanced intra- and inter-chromosomal rearrangements involving chromosomes 4, 11, 13, and X. Initial clinical studies were performed by karyotype, chromosomal microarray, and FISH with research-based short-read genome sequencing coupled with sanger sequencing to precisely map her breakpoints to the base pair resolution to understand the molecular basis of her phenotype. Genome analysis revealed two pathogenic deletions at 4p16.1-p15.32 and 4q31.1, accounting for her developmental delay and dysmorphism. We identified over 60 breakpoints, many with blunt ends and limited homology, supporting a role for non-homologous end joining in restructuring and resolution of the seminal chromoplexy event. We propose that the complexity of our patient's genomic rearrangements with a high number of breakpoints causes dysregulation of gene expression by three-dimensional chromatin interactions or topologically associating domains leading to growth failure and craniosynostosis. Our work supports an important role for genome sequencing in understanding the molecular basis of complex chromosomal rearrangements in human disease.

RevDate: 2023-01-25
CmpDate: 2022-07-29

Campbell M, Chantarasrivong C, Yanagihashi Y, et al (2022)

KSHV Topologically Associating Domains in Latent and Reactivated Viral Chromatin.

Journal of virology, 96(14):e0056522.

Eukaryotic genomes are structurally organized via the formation of multiple loops that create gene expression regulatory units called topologically associating domains (TADs). Here we revealed the KSHV TAD structure at 500 bp resolution and constructed a 3D KSHV genomic structural model with 2 kb binning. The latent KSHV genome formed very similar genomic architectures in three different naturally infected PEL cell lines and in an experimentally infected epithelial cell line. The majority of the TAD boundaries were occupied by structural maintenance of chromosomes (SMC1) cohesin complex and CCCTC-binding factor (CTCF), and the KSHV transactivator was recruited to those sites during reactivation. Triggering KSHV gene expression decreased prewired genomic loops within the regulatory unit, while contacts extending outside of regulatory borders increased, leading to formation of a larger regulatory unit with a shift from repressive to active compartments (B to A). The 3D genomic structural model proposes that the immediate early promoter region is localized on the periphery of the 3D viral genome during latency, while highly inducible noncoding RNA regions moved toward the inner space of the structure, resembling the configuration of a "bird cage" during reactivation. The compartment-like properties of viral episomal chromatin structure and its reorganization during the transition from latency may help facilitate viral gene transcription. IMPORTANCE The 3D architecture of chromatin allows for efficient arrangement, expression, and replication of genetic material. The genomes of all organisms studied to date have been found to be organized through some form of tiered domain structures. However, the architectural framework of the genomes of large double-stranded DNA viruses such as the herpesvirus family has not been reported. Prior studies with Kaposi's sarcoma-associated herpesvirus (KSHV) have indicated that the viral chromatin shares many biological properties exhibited by the host cell genome, essentially behaving as a mini human chromosome. Thus, we hypothesized that the KSHV genome may be organized in a similar manner. In this report, we describe the domain structure of the latent and lytic KSHV genome at 500 bp resolution and present a 3D genomic structural model for KSHV under each condition. These results add new insights into the complex regulation of the viral life cycle.

RevDate: 2022-09-13
CmpDate: 2022-07-26

Sabaté T, Zimmer C, E Bertrand (2022)

Versatile CRISPR-Based Method for Site-Specific Insertion of Repeat Arrays to Visualize Chromatin Loci in Living Cells.

Methods in molecular biology (Clifton, N.J.), 2532:275-290.

Hi-C and related sequencing-based techniques have brought a detailed understanding of the 3D genome architecture and the discovery of novel structures such as topologically associating domains (TADs) and chromatin loops, which emerge from cohesin-mediated DNA extrusion. However, these techniques require cell fixation, which precludes assessment of chromatin structure dynamics, and are generally restricted to population averages, thus masking cell-to-cell heterogeneity. By contrast, live-cell imaging allows to characterize and quantify the temporal dynamics of chromatin, potentially including TADs and loops in single cells. Specific chromatin loci can be visualized at high temporal and spatial resolution by inserting a repeat array from bacterial operator sequences bound by fluorescent tags. Using two different types of repeats allows to tag both anchors of a loop in different colors, thus enabling to track them separately even when they are in close vicinity. Here, we describe a versatile cloning method for generating many repeat array repair cassettes in parallel and inserting them by CRISPR-Cas9 into the human genome. This method should be instrumental to studying chromatin loop dynamics in single human cells.

RevDate: 2022-09-07
CmpDate: 2022-07-26

Miranda M, Noordermeer D, B Moindrot (2022)

Detection of Allele-Specific 3D Chromatin Interactions Using High-Resolution In-Nucleus 4C-seq.

Methods in molecular biology (Clifton, N.J.), 2532:15-33.

Chromosome conformation capture techniques are a set of methods used to determine 3D genome organization through the capture and identification of physical contacts between pairs of genomic loci. Among them, 4C-seq (circular chromosome conformation capture coupled to high-throughput sequencing) allows for the identification and quantification of the sequences interacting with a preselected locus of interest. 4C-seq has been widely used in the literature, mainly to study chromatin loops between enhancers and promoters or between CTCF binding sites and to identify chromatin domain boundaries. As 3D-contacts may be established in an allele-specific manner, we describe an up-to-date allele-specific 4C-seq protocol, starting from the selection of allele-specific viewpoints to Illumina sequencing. This protocol has mainly been optimized for cultured mammalian cells, but can be adapted for other cell types with relatively minor changes in initial steps.

RevDate: 2022-07-22

Sengupta K, Denkiewicz M, Chiliński M, et al (2022)

Multi-scale phase separation by explosive percolation with single-chromatin loop resolution.

Computational and structural biotechnology journal, 20:3591-3603.

The 2 m-long human DNA is tightly intertwined into the cell nucleus of the size of 10 μm. The DNA packing is explained by folding of chromatin fiber. This folding leads to the formation of such hierarchical structures as: chromosomal territories, compartments; densely-packed genomic regions known as Topologically Associating Domains (TADs), or Chromatin Contact Domains (CCDs), and loops. We propose models of dynamical human genome folding into hierarchical components in human lymphoblastoid, stem cell, and fibroblast cell lines. Our models are based on explosive percolation theory. The chromosomes are modeled as graphs where CTCF chromatin loops are represented as edges. The folding trajectory is simulated by gradually introducing loops to the graph following various edge addition strategies that are based on topological network properties, chromatin loop frequencies, compartmentalization, or epigenomic features. Finally, we propose the genome folding model - a biophysical pseudo-time process guided by a single scalar order parameter. The parameter is calculated by Linear Discriminant Analysis of chromatin features. We also include dynamics of loop formation by using Loop Extrusion Model (LEM) while adding them to the system. The chromatin phase separation, where fiber folds in 3D space into topological domains and compartments, is observed when the critical number of contacts is reached. We also observe that at least 80% of the loops are needed for chromatin fiber to condense in 3D space, and this is constant through various cell lines. Overall, our in-silico model integrates the high-throughput 3D genome interaction experimental data with the novel theoretical concept of phase separation, which allows us to model event-based time dynamics of chromatin loop formation and folding trajectories.

RevDate: 2022-07-16

Lamberti WF, C Zang (2022)

Extracting physical characteristics of higher-order chromatin structures from 3D image data.

Computational and structural biotechnology journal, 20:3387-3398.

Higher-order chromatin structures have functional impacts on gene regulation and cell identity determination. Using high-throughput sequencing (HTS)-based methods like Hi-C, active or inactive compartments and open or closed topologically associating domain (TAD) structures can be identified on a cell population level. Recently developed high-resolution three-dimensional (3D) molecular imaging techniques such as 3D electron microscopy with in situ hybridization (3D-EMSIH) and 3D structured illumination microscopy (3D-SIM) enable direct detection of physical representations of chromatin structures in a single cell. However, computational analysis of 3D image data with explainability and interpretability on functional characteristics of chromatin structures is still challenging. We developed Extracting Physical-Characteristics from Images of Chromatin Structures (EPICS), a machine-learning based computational method for processing high-resolution chromatin 3D image data. Using EPICS on images produced by 3D-EMISH or 3D-SIM techniques, we generated more direct 3D representations of higher-order chromatin structures, identified major chromatin domains, and determined the open or closed status of each domain. We identified several high-contributing features from the model as the major physical characteristics that define the open or closed chromatin domains, demonstrating the explainability and interpretability of EPICS. EPICS can be applied to the analysis of other high-resolution 3D molecular imaging data for spatial genomics studies. The R and Python codes of EPICS are available at https://github.com/zang-lab/epics.


RJR Experience and Expertise


Robbins holds BS, MS, and PhD degrees in the life sciences. He served as a tenured faculty member in the Zoology and Biological Science departments at Michigan State University. He is currently exploring the intersection between genomics, microbial ecology, and biodiversity — an area that promises to transform our understanding of the biosphere.


Robbins has extensive experience in college-level education: At MSU he taught introductory biology, genetics, and population genetics. At JHU, he was an instructor for a special course on biological database design. At FHCRC, he team-taught a graduate-level course on the history of genetics. At Bellevue College he taught medical informatics.


Robbins has been involved in science administration at both the federal and the institutional levels. At NSF he was a program officer for database activities in the life sciences, at DOE he was a program officer for information infrastructure in the human genome project. At the Fred Hutchinson Cancer Research Center, he served as a vice president for fifteen years.


Robbins has been involved with information technology since writing his first Fortran program as a college student. At NSF he was the first program officer for database activities in the life sciences. At JHU he held an appointment in the CS department and served as director of the informatics core for the Genome Data Base. At the FHCRC he was VP for Information Technology.


While still at Michigan State, Robbins started his first publishing venture, founding a small company that addressed the short-run publishing needs of instructors in very large undergraduate classes. For more than 20 years, Robbins has been operating The Electronic Scholarly Publishing Project, a web site dedicated to the digital publishing of critical works in science, especially classical genetics.


Robbins is well-known for his speaking abilities and is often called upon to provide keynote or plenary addresses at international meetings. For example, in July, 2012, he gave a well-received keynote address at the Global Biodiversity Informatics Congress, sponsored by GBIF and held in Copenhagen. The slides from that talk can be seen HERE.


Robbins is a skilled meeting facilitator. He prefers a participatory approach, with part of the meeting involving dynamic breakout groups, created by the participants in real time: (1) individuals propose breakout groups; (2) everyone signs up for one (or more) groups; (3) the groups with the most interested parties then meet, with reports from each group presented and discussed in a subsequent plenary session.


Robbins has been engaged with photography and design since the 1960s, when he worked for a professional photography laboratory. He now prefers digital photography and tools for their precision and reproducibility. He designed his first web site more than 20 years ago and he personally designed and implemented this web site. He engages in graphic design as a hobby.

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E-mail: RJR8222@gmail.com

Collection of publications by R J Robbins

Reprints and preprints of publications, slide presentations, instructional materials, and data compilations written or prepared by Robert Robbins. Most papers deal with computational biology, genome informatics, using information technology to support biomedical research, and related matters.

Research Gate page for R J Robbins

ResearchGate is a social networking site for scientists and researchers to share papers, ask and answer questions, and find collaborators. According to a study by Nature and an article in Times Higher Education , it is the largest academic social network in terms of active users.

Curriculum Vitae for R J Robbins

short personal version

Curriculum Vitae for R J Robbins

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RJR Picks from Around the Web (updated 11 MAY 2018 )