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Bibliography on: Topologically Associating Domains

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ESP: PubMed Auto Bibliography 29 Jun 2022 at 01:52 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: 2022-06-27

Sawh AN, SE Mango (2022)

Chromosome organization in 4D: insights from C. elegans development.

Current opinion in genetics & development, 75:101939 pii:S0959-437X(22)00048-X [Epub ahead of print].

Eukaryotic genome organization is ordered and multilayered, from the nucleosome to chromosomal scales. These layers are not static during development, but are remodeled over time and between tissues. Thus, animal model studies with high spatiotemporal resolution are necessary to understand the various forms and functions of genome organization in vivo. In C. elegans, sequencing- and imaging-based advances have provided insight on how histone modifications, regulatory elements, and large-scale chromosome conformations are established and changed. Recent observations include unexpected physiological roles for topologically associating domains, different roles for the nuclear lamina at different chromatin scales, cell-type-specific enhancer and promoter regulatory grammars, and prevalent compartment variability in early development. Here, we summarize these and other recent findings in C. elegans, and suggest future avenues of research to enrich our in vivo knowledge of the forms and functions of nuclear organization.

RevDate: 2022-06-24

Liu T, Z Wang (2022)

scHiCEmbed: Bin-Specific Embeddings of Single-Cell Hi-C Data Using Graph Auto-Encoders.

Genes, 13(6): pii:genes13061048.

Most publicly accessible single-cell Hi-C data are sparse and cannot reach a higher resolution. Therefore, learning latent representations (bin-specific embeddings) of sparse single-cell Hi-C matrices would provide us with a novel way of mining valuable information hidden in the limited number of single-cell Hi-C contacts. We present scHiCEmbed, an unsupervised computational method for learning bin-specific embeddings of single-cell Hi-C data, and the computational system is applied to the tasks of 3D structure reconstruction of whole genomes and detection of topologically associating domains (TAD). The only input of scHiCEmbed is a raw or scHiCluster-imputed single-cell Hi-C matrix. The main process of scHiCEmbed is to embed each node/bin in a higher dimensional space using graph auto-encoders. The learned n-by-3 bin-specific embedding/latent matrix is considered the final reconstructed 3D genome structure. For TAD detection, we use constrained hierarchical clustering on the latent matrix to classify bins: S_Dbw is used to determine the optimal number of clusters, and each cluster is considered as one potential TAD. Our reconstructed 3D structures for individual chromatins at different cell stages reveal the expanding process of chromatins during the cell cycle. We observe that the TADs called from single-cell Hi-C data are not shared across individual cells and that the TAD boundaries called from raw or imputed single-cell Hi-C are significantly different from those called from bulk Hi-C, confirming the cell-to-cell variability in terms of TAD definitions. The source code for scHiCEmbed is publicly available, and the URL can be found in the conclusion section.

RevDate: 2022-06-22

Tian GG, Hou C, Li J, et al (2022)

Three-dimensional genome structure shapes the recombination landscape of chromatin features during female germline stem cell development.

Clinical and translational medicine, 12(6):e927.

BACKGROUND: During meiosis of mammalian cells, chromatin undergoes drastic reorganization. However, the dynamics of the three-dimensional (3D) chromatin structure during the development of female germline stem cells (FGSCs) are poorly understood.

METHODS: The high-throughput chromosome conformation capture technique was used to probe the 3D structure of chromatin in mouse germ cells at each stage of FGSC development.

RESULTS: The global 3D genome was dramatically reorganized during FGSC development. In topologically associating domains, the chromatin structure was weakened in germinal vesicle stage oocytes and still present in meiosis I stage oocytes but had vanished in meiosis II oocytes. This switch between topologically associating domains was related to the biological process of FGSC development. Moreover, we constructed a landscape of chromosome X organization, which showed that the X chromosome occupied a smaller proportion of the active (A) compartment than the autosome during FGSC development. By comparing the high-order chromatin structure between female and male germline development, we found that 3D genome organization was remodelled by two different potential mechanisms during gamete development, in which interchromosomal interactions, compartments, and topologically associating domain were decreased during FGSC development but reorganized and recovered during spermatogenesis. Finally, we identified conserved chromatin structures between FGSC development and early embryonic development.

CONCLUSIONS: These results provide a valuable resource to characterize chromatin organization and for further studies of FGSC development.

RevDate: 2022-06-21

Wang W, Chandra A, Goldman N, et al (2022)

TCF-1 promotes chromatin interactions across topologically associating domains in T cell progenitors.

Nature immunology [Epub ahead of print].

The high mobility group (HMG) transcription factor TCF-1 is essential for early T cell development. Although in vitro biochemical assays suggest that HMG proteins can serve as architectural elements in the assembly of higher-order nuclear organization, the contribution of TCF-1 on the control of three-dimensional (3D) genome structures during T cell development remains unknown. Here, we investigated the role of TCF-1 in 3D genome reconfiguration. Using gain- and loss-of-function experiments, we discovered that the co-occupancy of TCF-1 and the architectural protein CTCF altered the structure of topologically associating domains in T cell progenitors, leading to interactions between previously insulated regulatory elements and target genes at late stages of T cell development. The TCF-1-dependent gain in long-range interactions was linked to deposition of active enhancer mark H3K27ac and recruitment of the cohesin-loading factor NIPBL at active enhancers. These data indicate that TCF-1 has a role in controlling global genome organization during T cell development.

RevDate: 2022-06-21

Liu E, Lyu H, Peng Q, et al (2022)

TADfit is a multivariate linear regression model for profiling hierarchical chromatin domains on replicate Hi-C data.

Communications biology, 5(1):608.

Topologically associating domains (TADs) are fundamental building blocks of three dimensional genome, and organized into complex hierarchies. Identifying hierarchical TADs on Hi-C data helps to understand the relationship between genome architectures and gene regulation. Herein we propose TADfit, a multivariate linear regression model for profiling hierarchical chromatin domains, which tries to fit the interaction frequencies in Hi-C contact matrix with and without replicates using all-possible hierarchical TADs, and the significant ones can be determined by the regression coefficients obtained with the help of an online learning solver called Follow-The-Regularized-Leader (FTRL). Beyond the existing methods, TADfit has an ability to handle multiple contact matrix replicates and find partially overlapping TADs on them, which helps to find the comprehensive underlying TADs across replicates from different experiments. The comparative results tell that TADfit has better accuracy and reproducibility, and the hierarchical TADs called by it exhibit a reasonable biological relevance.

RevDate: 2022-06-21
CmpDate: 2022-06-21

Bolt CC, Lopez-Delisle L, Hintermann A, et al (2022)

Context-dependent enhancer function revealed by targeted inter-TAD relocation.

Nature communications, 13(1):3488.

The expression of some genes depends on large, adjacent regions of the genome that contain multiple enhancers. These regulatory landscapes frequently align with Topologically Associating Domains (TADs), where they integrate the function of multiple similar enhancers to produce a global, TAD-specific regulation. We asked if an individual enhancer could overcome the influence of one of these landscapes, to drive gene transcription. To test this, we transferred an enhancer from its native location, into a nearby TAD with a related yet different functional specificity. We used the biphasic regulation of Hoxd genes during limb development as a paradigm. These genes are first activated in proximal limb cells by enhancers located in one TAD, which is then silenced when the neighboring TAD activates its enhancers in distal limb cells. We transferred a distal limb enhancer into the proximal limb TAD and found that its new context suppresses its normal distal specificity, even though it is bound by HOX13 transcription factors, which are responsible for the distal activity. This activity can be rescued only when a large portion of the surrounding environment is removed. These results indicate that, at least in some cases, the functioning of enhancer elements is subordinated to the host chromatin context, which can exert a dominant control over its activity.

RevDate: 2022-06-21
CmpDate: 2022-06-21

Michieletto D, WA Bickmore (2022)

TADs do not stay in the loop.

Molecular cell, 82(12):2188-2189.

In a recent issue of Science, Gabriele et al. have, for the first time, quantified the dynamics of a topologically associating domain (TAD) in live cells by coupling super-resolution imaging and computational modelling, concluding that a TAD spends most of its life in a "partially extruded state" and that CTCF-CTCF loops are rare.

RevDate: 2022-06-16

Sudarshan D, Avvakumov N, Lalonde ME, et al (2022)

Recurrent chromosomal translocations in sarcomas create a megacomplex that mislocalizes NuA4/TIP60 to Polycomb target loci.

Genes & development pii:gad.348982.121 [Epub ahead of print].

Chromosomal translocations frequently promote carcinogenesis by producing gain-of-function fusion proteins. Recent studies have identified highly recurrent chromosomal translocations in patients with endometrial stromal sarcomas (ESSs) and ossifying fibromyxoid tumors (OFMTs), leading to an in-frame fusion of PHF1 (PCL1) to six different subunits of the NuA4/TIP60 complex. While NuA4/TIP60 is a coactivator that acetylates chromatin and loads the H2A.Z histone variant, PHF1 is part of the Polycomb repressive complex 2 (PRC2) linked to transcriptional repression of key developmental genes through methylation of histone H3 on lysine 27. In this study, we characterize the fusion protein produced by the EPC1-PHF1 translocation. The chimeric protein assembles a megacomplex harboring both NuA4/TIP60 and PRC2 activities and leads to mislocalization of chromatin marks in the genome, in particular over an entire topologically associating domain including part of the HOXD cluster. This is linked to aberrant gene expression-most notably increased expression of PRC2 target genes. Furthermore, we show that JAZF1-implicated with a PRC2 component in the most frequent translocation in ESSs, JAZF1-SUZ12-is a potent transcription activator that physically associates with NuA4/TIP60, its fusion creating outcomes similar to those of EPC1-PHF1 Importantly, the specific increased expression of PRC2 targets/HOX genes was also confirmed with ESS patient samples. Altogether, these results indicate that most chromosomal translocations linked to these sarcomas use the same molecular oncogenic mechanism through a physical merge of NuA4/TIP60 and PRC2 complexes, leading to mislocalization of histone marks and aberrant Polycomb target gene expression.

RevDate: 2022-06-16

Taylor T, Sikorska N, Shchuka VM, et al (2022)

Transcriptional regulation and chromatin architecture maintenance are decoupled functions at the Sox2 locus.

Genes & development pii:gad.349489.122 [Epub ahead of print].

How distal regulatory elements control gene transcription and chromatin topology is not clearly defined, yet these processes are closely linked in lineage specification during development. Through allele-specific genome editing and chromatin interaction analyses of the Sox2 locus in mouse embryonic stem cells, we found a striking disconnection between transcriptional control and chromatin architecture. We traced nearly all Sox2 transcriptional activation to a small number of key transcription factor binding sites, whose deletions have no effect on promoter-enhancer interaction frequencies or topological domain organization. Local chromatin architecture maintenance, including at the topologically associating domain (TAD) boundary downstream from the Sox2 enhancer, is widely distributed over multiple transcription factor-bound regions and maintained in a CTCF-independent manner. Furthermore, partial disruption of promoter-enhancer interactions by ectopic chromatin loop formation has no effect on Sox2 transcription. These findings indicate that many transcription factors are involved in modulating chromatin architecture independently of CTCF.

RevDate: 2022-06-15

Lei Z, Meng H, Liu L, et al (2022)

Mitochondrial base editor induces substantial nuclear off-target mutations.

Nature [Epub ahead of print].

DddA-derived cytosine base editors (DdCBEs)-which are fusions of split DddA halves and transcription activator-like effector (TALE) array proteins from bacteria-enable targeted C•G-to-T•A conversions in mitochondrial DNA1. However, their genome-wide specificity is poorly understood. Here we show that the mitochondrial base editor induces extensive off-target editing in the nuclear genome. Genome-wide, unbiased analysis of its editome reveals hundreds of off-target sites that are TALE array sequence (TAS)-dependent or TAS-independent. TAS-dependent off-target sites in the nuclear DNA are often specified by only one of the two TALE repeats, challenging the principle that DdCBEs are guided by paired TALE proteins positioned in close proximity. TAS-independent off-target sites on nuclear DNA are frequently shared among DdCBEs with distinct TALE arrays. Notably, they co-localize strongly with binding sites for the transcription factor CTCF and are enriched in topologically associating domain boundaries. We engineered DdCBE to alleviate such off-target effects. Collectively, our results have implications for the use of DdCBEs in basic research and therapeutic applications, and suggest the need to thoroughly define and evaluate the off-target effects of base-editing tools.

RevDate: 2022-06-10

Shukla V, Cetnarowska A, Hyldahl M, et al (2022)

Interplay between regulatory elements and chromatin topology in cellular lineage determination.

Trends in genetics : TIG pii:S0168-9525(22)00119-6 [Epub ahead of print].

Cellular lineage determination is controlled by combinations of lineage-selective transcription factors (TFs) and associated coregulators that bind to cis-regulatory elements in DNA and regulate gene expression. The ability of these factors to regulate transcription is determined not only by their cooperativity, but also by biochemical and structural properties of the chromatin, sculpting higher-order genome organization. Here, we review recent advances in the understanding of the interplay between chromatin topology and transcription. Studies from many different fields, including adipocyte lineage determination, indicate that lineage determination and differentiation are dependent on elaborate crosstalk between cis-regulatory elements, leading to the formation of transcriptional hubs. Chromatin topology appears to provide a dynamic and supportive, rather than a deterministic, scaffold for this crosstalk.

RevDate: 2022-06-10

Tsujikawa LM, Kharenko OA, Stotz SC, et al (2022)

Breaking boundaries: Pan BETi disrupt 3D chromatin structure, BD2-selective BETi are strictly epigenetic transcriptional regulators.

Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie, 152:113230 pii:S0753-3322(22)00619-9 [Epub ahead of print].

BACKGROUND: Bromodomain and extraterminal proteins (BETs) are more than just epigenetic regulators of transcription. Here we highlight a new role for the BET protein BRD4 in the maintenance of higher order chromatin structure at Topologically Associating Domain Boundaries (TADBs). BD2-selective and pan (non-selective) BET inhibitors (BETi) differentially support chromatin structure, selectively affecting transcription and cell viability.

METHODS: Using RNA-seq and BRD4 ChIP-seq, the differential effect of BETi treatment on the transcriptome and BRD4 chromatin occupancy of human aortic endothelial cells from diabetic patients (dHAECs) stimulated with TNFα was evaluated. Chromatin decondensation and DNA fragmentation was assessed by immunofluorescence imaging and quantification. Key dHAEC findings were verified in proliferating monocyte-like THP-1 cells using real time-PCR, BRD4 co-immunoprecipitation studies, western blots, proliferation and apoptosis assays.

FINDINGS: We discovered that 1) BRD4 co-localizes with Ying-Yang 1 (YY1) at TADBs, critical chromatin structure complexes proximal to many DNA repair genes. 2) BD2-selective BETi enrich BRD4/YY1 associations, while pan-BETi do not. 3) Failure to support chromatin structures through BRD4/YY1 enrichment inhibits DNA repair gene transcription, which induces DNA damage responses, and causes widespread chromatin decondensation, DNA fragmentation, and apoptosis. 4) BD2-selective BETi maintain high order chromatin structure and cell viability, while reducing deleterious pro-inflammatory transcription.

INTERPRETATION: BRD4 plays a previously unrecognized role at TADBs. BETi differentially impact TADB stability. Our results provide translational insight for the development of BETi as therapeutics for a range of diseases including CVD, chronic kidney disease, cancer, and COVID-19.

RevDate: 2022-06-10

Segueni J, D Noordermeer (2022)

CTCF: A misguided jack-of-all-trades in cancer cells.

Computational and structural biotechnology journal, 20:2685-2698 pii:S2001-0370(22)00200-8.

The emergence and progression of cancers is accompanied by a dysregulation of transcriptional programs. The three-dimensional (3D) organization of the human genome has emerged as an important multi-level mediator of gene transcription and regulation. In cancer cells, this organization can be restructured, providing a framework for the deregulation of gene activity. The CTCF protein, initially identified as the product from a tumor suppressor gene, is a jack-of-all-trades for the formation of 3D genome organization in normal cells. Here, we summarize how CTCF is involved in the multi-level organization of the human genome and we discuss emerging insights into how perturbed CTCF function and DNA binding causes the activation of oncogenes in cancer cells, mostly through a process of enhancer hijacking. Moreover, we highlight non-canonical functions of CTCF that can be relevant for the emergence of cancers as well. Finally, we provide guidelines for the computational identification of perturbed CTCF binding and reorganized 3D genome structure in cancer cells.

RevDate: 2022-06-08

Emerson DJ, Zhao PA, Cook AL, et al (2022)

Cohesin-mediated loop anchors confine the locations of human replication origins.

Nature [Epub ahead of print].

DNA replication occurs through an intricately regulated series of molecular events and is fundamental for genome stability1,2. At present, it is unknown how the locations of replication origins are determined in the human genome. Here we dissect the role of topologically associating domains (TADs)3-6, subTADs7 and loops8 in the positioning of replication initiation zones (IZs). We stratify TADs and subTADs by the presence of corner-dots indicative of loops and the orientation of CTCF motifs. We find that high-efficiency, early replicating IZs localize to boundaries between adjacent corner-dot TADs anchored by high-density arrays of divergently and convergently oriented CTCF motifs. By contrast, low-efficiency IZs localize to weaker dotless boundaries. Following ablation of cohesin-mediated loop extrusion during G1, high-efficiency IZs become diffuse and delocalized at boundaries with complex CTCF motif orientations. Moreover, G1 knockdown of the cohesin unloading factor WAPL results in gained long-range loops and narrowed localization of IZs at the same boundaries. Finally, targeted deletion or insertion of specific boundaries causes local replication timing shifts consistent with IZ loss or gain, respectively. Our data support a model in which cohesin-mediated loop extrusion and stalling at a subset of genetically encoded TAD and subTAD boundaries is an essential determinant of the locations of replication origins in human S phase.

RevDate: 2022-06-06

Zhang S, Tao W, JJ Han (2022)

3D chromatin structure changes during spermatogenesis and oogenesis.

Computational and structural biotechnology journal, 20:2434-2441 pii:S2001-0370(22)00188-X.

Gametogenesis, including spermatogenesis and oogenesis, are unique differentiation processes involving extraordinarily complex and precise regulatory mechanisms that require the interactions of multiple cell types, hormones, paracrine factors, genes and epigenetic regulators, and extensive chromatin 3D structure re-organization. In recent years, the development of 3D genome technology represented by Hi-C, enabled mapping of the 3D re-organization of chromosomes during zygogenesis at an unprecedented resolution. The 3D remodeling is achieved by folding chromatin into loops, topologically associating domains (TADs), and compartments (A and B), which ultimately affect transcriptional activity. In this review, we summarize the research progresses and findings on chromatin 3D structure changes during spermatogenesis and oogenesis.

RevDate: 2022-05-28

Zhang Y, Tian GG, Wang X, et al (2022)

Retinoic acid induced meiosis initiation in female germline stem cells by remodelling three-dimensional chromatin structure.

Cell proliferation [Epub ahead of print].

OBJECTIVES: This study aimed to clarify the regulation and mechanism of meiotic initiation in FGSC development.

MATERIALS AND METHODS: FGSCs were induced to differentiate into meiosis in differentiation medium. RNA sequencing was performed to analysis the difference of transcription level. High-through chromosome conformation capture sequencing (Hi-C) was performed to analysis changes of three-dimensional chromatin structure. Chromosome conformation capture further confirmed a spatial chromatin loop. ChIP-qPCR and dual luciferase reporter were used to test the interaction between Stimulated by retinoic acid gene 8 (STRA8) protein and Trip13 promoter.

RESULTS: Compared with FGSCs, the average diameter of STRA8-positive germ cells increased from 13 μm to 16.8 μm. Furthermore, there were 4788 differentially expressed genes between the two cell stages; Meiosis and chromatin structure-associated terms were significantly enriched. Additionally, Hi-C results showed that FGSCs underwent A/B compartment switching (switch rate was 29.81%), the number of topologically associating domains (TADs) increasing, the average size of TADs decreasing, and chromatin loop changes at genome region of Trip13 from undifferentiated stage to meiosis-initiation stage. Furthermore, we validated that Trip13 promoter contacted distal enhancer to form spatial chromatin loop and STRA8 could bind Trip13 promoter to promote gene expression.

CONCLUSION: FGSCs underwent chromatin structure remodelling from undifferentiated stage to meiosis-initiation stage, which facilitated STRA8 binding to Trip13 promoter and promoting its expression.

RevDate: 2022-05-19

Bin Akhtar G, Buist M, M Rastegar (2022)

MeCP2 and transcriptional control of eukaryotic gene expression.

European journal of cell biology, 101(3):151237 pii:S0171-9335(22)00040-1 [Epub ahead of print].

Eukaryotic gene expression is controlled at multiple steps that work in harmony to ensure proper maintenance of cellular morphology and function. Such regulatory mechanisms would include transcriptional gene regulation, which is in turn controlled by chromatin remodeling, distinct topologically associating domains of the chromatin structure, cis-regulatory elements such as enhancers and promoters, action of trans-acting factors, DNA methylation, RNA modifications, and post-translational modification of histones. These guiding mechanisms of gene expression play critical roles in the epigenetic setting of individual cells within the eukaryotic systems. Some epigenetic factors may play multiple functional roles in guarding the accurate gene expression program of the eukaryotic cells, especially within the central nervous system. A well-studied example of such multi-functional factors is the methyl-CpG-binding protein 2 (MeCP2), a nuclear protein that is encoded by the X-linked MECP2 gene. Here, we aim to provide an overview of eukaryotic gene regulation, the three-dimensional chromatin organization, standard techniques to study newly synthesized RNA transcripts, and the role of MeCP2 as an important transcriptional regulator in eukaryotes.

RevDate: 2022-05-18

Dequeker BJH, Scherr MJ, Brandão HB, et al (2022)

MCM complexes are barriers that restrict cohesin-mediated loop extrusion.

Nature [Epub ahead of print].

Eukaryotic genomes are compacted into loops and topologically associating domains (TADs)1-3, which contribute to transcription, recombination and genomic stability4,5. Cohesin extrudes DNA into loops that are thought to lengthen until CTCF boundaries are encountered6-12. Little is known about whether loop extrusion is impeded by DNA-bound machines. Here we show that the minichromosome maintenance (MCM) complex is a barrier that restricts loop extrusion in G1 phase. Single-nucleus Hi-C (high-resolution chromosome conformation capture) of mouse zygotes reveals that MCM loading reduces CTCF-anchored loops and decreases TAD boundary insulation, which suggests that loop extrusion is impeded before reaching CTCF. This effect extends to HCT116 cells, in which MCMs affect the number of CTCF-anchored loops and gene expression. Simulations suggest that MCMs are abundant, randomly positioned and partially permeable barriers. Single-molecule imaging shows that MCMs are physical barriers that frequently constrain cohesin translocation in vitro. Notably, chimeric yeast MCMs that contain a cohesin-interaction motif from human MCM3 induce cohesin pausing, indicating that MCMs are 'active' barriers with binding sites. These findings raise the possibility that cohesin can arrive by loop extrusion at MCMs, which determine the genomic sites at which sister chromatid cohesion is established. On the basis of in vivo, in silico and in vitro data, we conclude that distinct loop extrusion barriers shape the three-dimensional genome.

RevDate: 2022-05-18

Zhou J (2022)

Sequence-based modeling of three-dimensional genome architecture from kilobase to chromosome scale.

Nature genetics, 54(5):725-734.

To learn how genomic sequence influences multiscale three-dimensional (3D) genome architecture, this manuscript presents a sequence-based deep-learning approach, Orca, that predicts directly from sequence the 3D genome architecture from kilobase to whole-chromosome scale. Orca captures the sequence dependencies of structures including chromatin compartments and topologically associating domains, as well as diverse types of interactions from CTCF-mediated to enhancer-promoter interactions and Polycomb-mediated interactions with cell-type specificity. Orca enables various applications including predicting structural variant effects on multiscale genome organization and it recapitulated effects of experimentally studied variants at varying sizes (300 bp to 90 Mb). Moreover, Orca enables in silico virtual screens to probe the sequence basis of 3D genome organization at different scales. At the submegabase scale, it predicted specific transcription factor motifs underlying cell-type-specific genome interactions. At the compartment scale, virtual screens of sequence activities suggest a model for the sequence basis of chromatin compartments with a prominent role of transcription start sites.

RevDate: 2022-05-10

Fan Z, Wu C, Chen M, et al (2022)

The generation of PD-L1 and PD-L2 in cancer cells: From nuclear chromatin reorganization to extracellular presentation.

Acta pharmaceutica Sinica. B, 12(3):1041-1053.

The immune checkpoint blockade (ICB) targeting on PD-1/PD-L1 has shown remarkable promise in treating cancers. However, the low response rate and frequently observed severe side effects limit its broad benefits. It is partially due to less understanding of the biological regulation of PD-L1. Here, we systematically and comprehensively summarized the regulation of PD-L1 from nuclear chromatin reorganization to extracellular presentation. In PD-L1 and PD-L2 highly expressed cancer cells, a new TAD (topologically associating domain) (chr9: 5,400,000-5,600,000) around CD274 and CD273 was discovered, which includes a reported super-enhancer to drive synchronous transcription of PD-L1 and PD-L2. The re-shaped TAD allows transcription factors such as STAT3 and IRF1 recruit to PD-L1 locus in order to guide the expression of PD-L1. After transcription, the PD-L1 is tightly regulated by miRNAs and RNA-binding proteins via the long 3'UTR. At translational level, PD-L1 protein and its membrane presentation are tightly regulated by post-translational modification such as glycosylation and ubiquitination. In addition, PD-L1 can be secreted via exosome to systematically inhibit immune response. Therefore, fully dissecting the regulation of PD-L1/PD-L2 and thoroughly detecting PD-L1/PD-L2 as well as their regulatory networks will bring more insights in ICB and ICB-based combinational therapy.

RevDate: 2022-05-07

Poszewiecka B, Pienkowski VM, Nowosad K, et al (2022)

TADeus2: a web server facilitating the clinical diagnosis by pathogenicity assessment of structural variations disarranging 3D chromatin structure.

Nucleic acids research pii:6582161 [Epub ahead of print].

In recent years great progress has been made in identification of structural variants (SV) in the human genome. However, the interpretation of SVs, especially located in non-coding DNA, remains challenging. One of the reasons stems in the lack of tools exclusively designed for clinical SVs evaluation acknowledging the 3D chromatin architecture. Therefore, we present TADeus2 a web server dedicated for a quick investigation of chromatin conformation changes, providing a visual framework for the interpretation of SVs affecting topologically associating domains (TADs). This tool provides a convenient visual inspection of SVs, both in a continuous genome view as well as from a rearrangement's breakpoint perspective. Additionally, TADeus2 allows the user to assess the influence of analyzed SVs within flaking coding/non-coding regions based on the Hi-C matrix. Importantly, the SVs pathogenicity is quantified and ranked using TADA, ClassifyCNV tools and sampling-based P-value. TADeus2 is publicly available at

RevDate: 2022-05-10
CmpDate: 2022-05-10

Li D, He M, Tang Q, et al (2022)

Comparative 3D genome architecture in vertebrates.

BMC biology, 20(1):99.

BACKGROUND: The three-dimensional (3D) architecture of the genome has a highly ordered and hierarchical nature, which influences the regulation of essential nuclear processes at the basis of gene expression, such as gene transcription. While the hierarchical organization of heterochromatin and euchromatin can underlie differences in gene expression that determine evolutionary differences among species, the way 3D genome architecture is affected by evolutionary forces within major lineages remains unclear. Here, we report a comprehensive comparison of 3D genomes, using high resolution Hi-C data in fibroblast cells of fish, chickens, and 10 mammalian species.

RESULTS: This analysis shows a correlation between genome size and chromosome length that affects chromosome territory (CT) organization in the upper hierarchy of genome architecture, whereas lower hierarchical features, including local transcriptional availability of DNA, are selected through the evolution of vertebrates. Furthermore, conservation of topologically associating domains (TADs) appears strongly associated with the modularity of expression profiles across species. Additionally, LINE and SINE transposable elements likely contribute to heterochromatin and euchromatin organization, respectively, during the evolution of genome architecture.

CONCLUSIONS: Our analysis uncovers organizational features that appear to determine the conservation and transcriptional regulation of functional genes across species. These findings can guide ongoing investigations of genome evolution by extending our understanding of the mechanisms shaping genome architecture.

RevDate: 2022-05-17

Deng S, Feng Y, S Pauklin (2022)

3D chromatin architecture and transcription regulation in cancer.

Journal of hematology & oncology, 15(1):49.

Chromatin has distinct three-dimensional (3D) architectures important in key biological processes, such as cell cycle, replication, differentiation, and transcription regulation. In turn, aberrant 3D structures play a vital role in developing abnormalities and diseases such as cancer. This review discusses key 3D chromatin structures (topologically associating domain, lamina-associated domain, and enhancer-promoter interactions) and corresponding structural protein elements mediating 3D chromatin interactions [CCCTC-binding factor, polycomb group protein, cohesin, and Brother of the Regulator of Imprinted Sites (BORIS) protein] with a highlight of their associations with cancer. We also summarise the recent development of technologies and bioinformatics approaches to study the 3D chromatin interactions in gene expression regulation, including crosslinking and proximity ligation methods in the bulk cell population (ChIA-PET and HiChIP) or single-molecule resolution (ChIA-drop), and methods other than proximity ligation, such as GAM, SPRITE, and super-resolution microscopy techniques.

RevDate: 2022-05-06
CmpDate: 2022-05-06

Gilbertson SE, Walter HC, Gardner K, et al (2022)

Topologically associating domains are disrupted by evolutionary genome rearrangements forming species-specific enhancer connections in mice and humans.

Cell reports, 39(5):110769.

Distinguishing between conserved and divergent regulatory mechanisms is essential for translating preclinical research from mice to humans, yet there is a lack of information about how evolutionary genome rearrangements affect the regulation of the immune response, a rapidly evolving system. The current model is topologically associating domains (TADs) are conserved between species, buffering evolutionary rearrangements and conserving long-range interactions within a TAD. However, we find that TADs frequently span evolutionary translocation and inversion breakpoints near genes with species-specific expression in immune cells, creating unique enhancer-promoter interactions exclusive to the mouse or human genomes. This includes TADs encompassing immune-related transcription factors, cytokines, and receptors. For example, we uncover an evolutionary rearrangement that created a shared LPS-inducible regulatory module between OASL and P2RX7 in human macrophages that is absent in mice. Therefore, evolutionary genome rearrangements disrupt TAD boundaries, enabling sequence-conserved enhancer elements from divergent genomic locations between species to create unique regulatory modules.

RevDate: 2022-05-09
CmpDate: 2022-05-09

Galupa R, Picard C, Servant N, et al (2022)

Inversion of a topological domain leads to restricted changes in its gene expression and affects interdomain communication.

Development (Cambridge, England), 149(9):.

The interplay between the topological organization of the genome and the regulation of gene expression remains unclear. Depletion of molecular factors (e.g. CTCF) underlying topologically associating domains (TADs) leads to modest alterations in gene expression, whereas genomic rearrangements involving TAD boundaries disrupt normal gene expression and can lead to pathological phenotypes. Here, we targeted the TAD neighboring that of the noncoding transcript Xist, which controls X-chromosome inactivation. Inverting 245 kb within the TAD led to expected rearrangement of CTCF-based contacts but revealed heterogeneity in the 'contact' potential of different CTCF sites. Expression of most genes therein remained unaffected in mouse embryonic stem cells and during differentiation. Interestingly, expression of Xist was ectopically upregulated. The same inversion in mouse embryos led to biased Xist expression. Smaller inversions and deletions of CTCF clusters led to similar results: rearrangement of contacts and limited changes in local gene expression, but significant changes in Xist expression in embryos. Our study suggests that the wiring of regulatory interactions within a TAD can influence the expression of genes in neighboring TADs, highlighting the existence of mechanisms of inter-TAD communication.

RevDate: 2022-04-29
CmpDate: 2022-04-29

Panarotto M, Davidson IF, Litos G, et al (2022)

Cornelia de Lange syndrome mutations in NIPBL can impair cohesin-mediated DNA loop extrusion.

Proceedings of the National Academy of Sciences of the United States of America, 119(18):e2201029119.

Cornelia de Lange syndrome (CdLS) is a developmental multisystem disorder frequently associated with mutations in NIPBL. CdLS is thought to arise from developmental gene regulation defects, but how NIPBL mutations cause these is unknown. Here we show that several NIPBL mutations impair the DNA loop extrusion activity of cohesin. Because this activity is required for the formation of chromatin loops and topologically associating domains, which have important roles in gene regulation, our results suggest that defects in cohesin-mediated loop extrusion contribute to the etiology of CdLS by altering interactions between developmental genes and their enhancers.

RevDate: 2022-04-26

Jouret G, Heide S, Sorlin A, et al (2022)

Understanding the new BRD4-related syndrome: Clinical and genomic delineation with an international cohort study.

Clinical genetics [Epub ahead of print].

BRD4 is part of a multiprotein complex involved in loading the cohesin complex onto DNA, a fundamental process required for cohesin-mediated loop extrusion and formation of Topologically Associating Domains. Pathogenic variations in this complex have been associated with a growing number of syndromes, collectively known as cohesinopathies, the most classic being Cornelia de Lange syndrome. However, no cohort study has been conducted to delineate the clinical and molecular spectrum of BRD4-related disorder. We formed an international collaborative study, and collected 14 new patients, including two fetuses. We performed phenotype and genotype analysis, integrated prenatal findings from fetopathological examinations, phenotypes of pediatric patients and adults. We report the first cohort of patients with BRD4-related disorder and delineate the dysmorphic features at different ages. This work extends the phenotypic spectrum of cohesinopathies and characterize a new clinically relevant and recognizable pattern, distinguishable from the other cohesinopathies.

RevDate: 2022-04-29

Habash NW, Sehrawat TS, Shah VH, et al (2022)

Epigenetics of alcohol-related liver diseases.

JHEP reports : innovation in hepatology, 4(5):100466.

Alcohol-related liver disease (ARLD) is a primary cause of chronic liver disease in the United States. Despite advances in the diagnosis and management of ARLD, it remains a major public health problem associated with significant morbidity and mortality, emphasising the need to adopt novel approaches to the study of ARLD and its complications. Epigenetic changes are increasingly being recognised as contributing to the pathogenesis of multiple disease states. Harnessing the power of innovative technologies for the study of epigenetics (e.g., next-generation sequencing, DNA methylation assays, histone modification profiling and computational techniques like machine learning) has resulted in a seismic shift in our understanding of the pathophysiology of ARLD. Knowledge of these techniques and advances is of paramount importance for the practicing hepatologist and researchers alike. Accordingly, in this review article we will summarise the current knowledge about alcohol-induced epigenetic alterations in the context of ARLD, including but not limited to, DNA hyper/hypo methylation, histone modifications, changes in non-coding RNA, 3D chromatin architecture and enhancer-promoter interactions. Additionally, we will discuss the state-of-the-art techniques used in the study of ARLD (e.g. single-cell sequencing). We will also highlight the epigenetic regulation of chemokines and their proinflammatory role in the context of ARLD. Lastly, we will examine the clinical applications of epigenetics in the diagnosis and management of ARLD.

RevDate: 2022-05-07

Liu W, Zhong W, Chen J, et al (2022)

Understanding Regulatory Mechanisms of Brain Function and Disease through 3D Genome Organization.

Genes, 13(4):.

The human genome has a complex and dynamic three-dimensional (3D) organization, which plays a critical role for gene regulation and genome function. The importance of 3D genome organization in brain development and function has been well characterized in a region- and cell-type-specific fashion. Recent technological advances in chromosome conformation capture (3C)-based techniques, imaging approaches, and ligation-free methods, along with computational methods to analyze the data generated, have revealed 3D genome features at different scales in the brain that contribute to our understanding of genetic mechanisms underlying neuropsychiatric diseases and other brain-related traits. In this review, we discuss how these advances aid in the genetic dissection of brain-related traits.

RevDate: 2022-04-29
CmpDate: 2022-04-21

Aljahani A, Hua P, Karpinska MA, et al (2022)

Analysis of sub-kilobase chromatin topology reveals nano-scale regulatory interactions with variable dependence on cohesin and CTCF.

Nature communications, 13(1):2139.

Enhancers and promoters predominantly interact within large-scale topologically associating domains (TADs), which are formed by loop extrusion mediated by cohesin and CTCF. However, it is unclear whether complex chromatin structures exist at sub-kilobase-scale and to what extent fine-scale regulatory interactions depend on loop extrusion. To address these questions, we present an MNase-based chromosome conformation capture (3C) approach, which has enabled us to generate the most detailed local interaction data to date (20 bp resolution) and precisely investigate the effects of cohesin and CTCF depletion on chromatin architecture. Our data reveal that cis-regulatory elements have distinct internal nano-scale structures, within which local insulation is dependent on CTCF, but which are independent of cohesin. In contrast, we find that depletion of cohesin causes a subtle reduction in longer-range enhancer-promoter interactions and that CTCF depletion can cause rewiring of regulatory contacts. Together, our data show that loop extrusion is not essential for enhancer-promoter interactions, but contributes to their robustness and specificity and to precise regulation of gene expression.

RevDate: 2022-05-07

Gabriele M, Brandão HB, Grosse-Holz S, et al (2022)

Dynamics of CTCF- and cohesin-mediated chromatin looping revealed by live-cell imaging.

Science (New York, N.Y.), 376(6592):496-501.

Animal genomes are folded into loops and topologically associating domains (TADs) by CTCF and loop-extruding cohesins, but the live dynamics of loop formation and stability remain unknown. Here, we directly visualized chromatin looping at the Fbn2 TAD in mouse embryonic stem cells using super-resolution live-cell imaging and quantified looping dynamics by Bayesian inference. Unexpectedly, the Fbn2 loop was both rare and dynamic, with a looped fraction of approximately 3 to 6.5% and a median loop lifetime of approximately 10 to 30 minutes. Our results establish that the Fbn2 TAD is highly dynamic, and about 92% of the time, cohesin-extruded loops exist within the TAD without bridging both CTCF boundaries. This suggests that single CTCF boundaries, rather than the fully CTCF-CTCF looped state, may be the primary regulators of functional interactions.

RevDate: 2022-05-09

Zuin J, Roth G, Zhan Y, et al (2022)

Nonlinear control of transcription through enhancer-promoter interactions.

Nature, 604(7906):571-577.

Chromosome structure in mammals is thought to regulate transcription by modulating three-dimensional interactions between enhancers and promoters, notably through CTCF-mediated loops and topologically associating domains (TADs)1-4. However, how chromosome interactions are actually translated into transcriptional outputs remains unclear. Here, to address this question, we use an assay to position an enhancer at large numbers of densely spaced chromosomal locations relative to a fixed promoter, and measure promoter output and interactions within a genomic region with minimal regulatory and structural complexity. A quantitative analysis of hundreds of cell lines reveals that the transcriptional effect of an enhancer depends on its contact probabilities with the promoter through a nonlinear relationship. Mathematical modelling suggests that nonlinearity might arise from transient enhancer-promoter interactions being translated into slower promoter bursting dynamics in individual cells, therefore uncoupling the temporal dynamics of interactions from those of transcription. This uncovers a potential mechanism of how distal enhancers act from large genomic distances, and of how topologically associating domain boundaries block distal enhancers. Finally, we show that enhancer strength also determines absolute transcription levels as well as the sensitivity of a promoter to CTCF-mediated transcriptional insulation. Our measurements establish general principles for the context-dependent role of chromosome structure in long-range transcriptional regulation.

RevDate: 2022-05-06
CmpDate: 2022-04-14

Sefer E (2022)

A comparison of topologically associating domain callers over mammals at high resolution.

BMC bioinformatics, 23(1):127.

BACKGROUND: Topologically associating domains (TADs) are locally highly-interacting genome regions, which also play a critical role in regulating gene expression in the cell. TADs have been first identified while investigating the 3D genome structure over High-throughput Chromosome Conformation Capture (Hi-C) interaction dataset. Substantial degree of efforts have been devoted to develop techniques for inferring TADs from Hi-C interaction dataset. Many TAD-calling methods have been developed which differ in their criteria and assumptions in TAD inference. Correspondingly, TADs inferred via these callers vary in terms of both similarities and biological features they are enriched in.

RESULT: We have carried out a systematic comparison of 27 TAD-calling methods over mammals. We use Micro-C, a recent high-resolution variant of Hi-C, to compare TADs at a very high resolution, and classify the methods into 3 categories: feature-based methods, Clustering methods, Graph-partitioning methods. We have evaluated TAD boundaries, gaps between adjacent TADs, and quality of TADs across various criteria. We also found particularly CTCF and Cohesin proteins to be effective in formation of TADs with corner dots. We have also assessed the callers performance on simulated datasets since a gold standard for TADs is missing. TAD sizes and numbers change remarkably between TAD callers and dataset resolutions, indicating that TADs are hierarchically-organized domains, instead of disjoint regions. A core subset of feature-based TAD callers regularly perform the best while inferring reproducible domains, which are also enriched for TAD related biological properties.

CONCLUSION: We have analyzed the fundamental principles of TAD-calling methods, and identified the existing situation in TAD inference across high resolution Micro-C interaction datasets over mammals. We come up with a systematic, comprehensive, and concise framework to evaluate the TAD-calling methods performance across Micro-C datasets. Our research will be useful in selecting appropriate methods for TAD inference and evaluation based on available data, experimental design, and biological question of interest. We also introduce our analysis as a benchmarking tool with publicly available source code.

RevDate: 2022-05-15
CmpDate: 2022-04-14

Xie L, Dong P, Qi Y, et al (2022)

BRD2 compartmentalizes the accessible genome.

Nature genetics, 54(4):481-491.

Mammalian chromosomes are organized into megabase-sized compartments that are further subdivided into topologically associating domains (TADs). While the formation of TADs is dependent on cohesin, the mechanism behind compartmentalization remains enigmatic. Here, we show that the bromodomain and extraterminal (BET) family scaffold protein BRD2 promotes spatial mixing and compartmentalization of active chromatin after cohesin loss. This activity is independent of transcription but requires BRD2 to recognize acetylated targets through its double bromodomain and interact with binding partners with its low-complexity domain. Notably, genome compartmentalization mediated by BRD2 is antagonized on the one hand by cohesin and on the other hand by the BET homolog protein BRD4, both of which inhibit BRD2 binding to chromatin. Polymer simulation of our data supports a BRD2-cohesin interplay model of nuclear topology, in which genome compartmentalization results from a competition between loop extrusion and chromatin-state-specific affinity interactions.

RevDate: 2022-05-06

Feng Y, Cai L, Hong W, et al (2022)

Rewiring of 3D Chromatin Topology Orchestrates Transcriptional Reprogramming and the Development of Human Dilated Cardiomyopathy.

Circulation [Epub ahead of print].

Background: Transcriptional reconfiguration is central to heart failure, the common cause of which is dilated cardiomyopathy (DCM). However, the impact of three-dimensional (3D) chromatin topology on transcriptional dysregulation and pathogenesis in human DCM remains elusive. Methods: We generated a compendium of 3D-epigenome and transcriptome maps from 101 biobanked human DCM and non-failing heart tissues through HiChIP (H3K27ac), in situ Hi-C, ChIP-seq, ATAC-seq and RNA-seq profiling. We employed human iPSC-derived cardiomyocytes (hiPSC-CMs) and mouse models to further interrogate the key transcription factor implicated in 3D chromatin organization and transcriptional regulation in DCM pathogenesis. Results: We discovered that the active regulatory elements (H3K27ac peaks) and their connectome (H3K27ac loops) were extensively reprogrammed in DCM hearts and contributed to transcriptional dysregulation implicated for DCM development. For example, we identified that non-transcribing NPPA-AS1 promoter functions as an enhancer and physically interacts with the NPPA and NPPB promoters, leading to the co-transcription of NPPA and NPPB in DCM hearts. We uncovered that DCM-enriched H3K27ac loops largely resided in conserved high-order chromatin architectures (Compartments, Topologically Associating Domains) and unexpectedly their anchors had equivalent chromatin accessibility. Intriguingly, we discovered that the DCM-enriched H3K27ac loop anchors exhibited a strong enrichment for Heart and Neural Crest Derivatives Expressed 1 (HAND1), a key transcription factor involved in early cardiogenesis. In line with this, its protein expression was upregulated in human DCM and mouse failing hearts. To further validate whether HAND1 is a causal driver for the reprogramming of enhancer/promoter connectome in DCM hearts, we performed comprehensive 3D epigenome mappings in hiPSC-CMs. We found that forced overexpression of HAND1 in hiPSC-CM induced a distinct gain of enhancer/promoter connectivity and, correspondingly, increased the expression of their connected genes implicated in DCM etiology, thus recapitulating the transcriptional signature in human DCM hearts. Moreover, electrophysiology analysis demonstrated that forced overexpression of HAND1 in hiPSC-CM induced abnormal calcium handling. Furthermore, cardiomyocyte-specific overexpression of Hand1 in the mouse hearts resulted in a dilated cardiac remodeling with impaired contractility/Ca2+ handling in cardiomyocytes, increased ratio of heart weight/body weight and compromised cardiac function, which were ascribed to recapitulation of transcriptional reprogramming in DCM. Conclusions: This study provided novel chromatin topology insights into DCM pathogenesis and illustrated a model whereby a single transcription factor (HAND1) reprograms the genome-wide enhancer/promoter connectome to drive DCM pathogenesis.

RevDate: 2022-05-17

Chu Z, Gu L, Hu Y, et al (2022)

STAG2 regulates interferon signaling in melanoma via enhancer loop reprogramming.

Nature communications, 13(1):1859.

The cohesin complex participates in the organization of 3D genome through generating and maintaining DNA loops. Stromal antigen 2 (STAG2), a core subunit of the cohesin complex, is frequently mutated in various cancers. However, the impact of STAG2 inactivation on 3D genome organization, especially the long-range enhancer-promoter contacts and subsequent gene expression control in cancer, remains poorly understood. Here we show that depletion of STAG2 in melanoma cells leads to expansion of topologically associating domains (TADs) and enhances the formation of acetylated histone H3 lysine 27 (H3K27ac)-associated DNA loops at sites where binding of STAG2 is switched to its paralog STAG1. We further identify Interferon Regulatory Factor 9 (IRF9) as a major direct target of STAG2 in melanoma cells via integrated RNA-seq, STAG2 ChIP-seq and H3K27ac HiChIP analyses. We demonstrate that loss of STAG2 activates IRF9 through modulating the 3D genome organization, which in turn enhances type I interferon signaling and increases the expression of PD-L1. Our findings not only establish a previously unknown role of the STAG2 to STAG1 switch in 3D genome organization, but also reveal a functional link between STAG2 and interferon signaling in cancer cells, which may enhance the immune evasion potential in STAG2-mutant cancer.

RevDate: 2022-05-06

Serna-Pujol N, Salinas-Pena M, Mugianesi F, et al (2022)

Coordinated changes in gene expression, H1 variant distribution and genome 3D conformation in response to H1 depletion.

Nucleic acids research, 50(7):3892-3910.

Up to seven members of the histone H1 family may contribute to chromatin compaction and its regulation in human somatic cells. In breast cancer cells, knock-down of multiple H1 variants deregulates many genes, promotes the appearance of genome-wide accessibility sites and triggers an interferon response via activation of heterochromatic repeats. However, how these changes in the expression profile relate to the re-distribution of H1 variants as well as to genome conformational changes have not been yet studied. Here, we combined ChIP-seq of five endogenous H1 variants with Chromosome Conformation Capture analysis in wild-type and H1.2/H1.4 knock-down T47D cells. The results indicate that H1 variants coexist in the genome in two large groups depending on the local GC content and that their distribution is robust with respect to H1 depletion. Despite the small changes in H1 variants distribution, knock-down of H1 translated into more isolated but de-compacted chromatin structures at the scale of topologically associating domains (TADs). Such changes in TAD structure correlated with a coordinated gene expression response of their resident genes. This is the first report describing simultaneous profiling of five endogenous H1 variants and giving functional evidence of genome topology alterations upon H1 depletion in human cancer cells.

RevDate: 2022-05-18

Brown JM, De Ornellas S, Parisi E, et al (2022)

RASER-FISH: non-denaturing fluorescence in situ hybridization for preservation of three-dimensional interphase chromatin structure.

Nature protocols, 17(5):1306-1331.

DNA fluorescence in situ hybridization (FISH) has been a central technique in advancing our understanding of how chromatin is organized within the nucleus. With the increasing resolution offered by super-resolution microscopy, the optimal maintenance of chromatin structure within the nucleus is essential for accuracy in measurements and interpretation of data. However, standard 3D-FISH requires potentially destructive heat denaturation in the presence of chaotropic agents such as formamide to allow access to the DNA strands for labeled FISH probes. To avoid the need to heat-denature, we developed Resolution After Single-strand Exonuclease Resection (RASER)-FISH, which uses exonuclease digestion to generate single-stranded target DNA for efficient probe binding over a 2 d process. Furthermore, RASER-FISH is easily combined with immunostaining of nuclear proteins or the detection of RNAs. Here, we provide detailed procedures for RASER-FISH in mammalian cultured cells to detect single loci, chromatin tracks and topologically associating domains with conventional and super-resolution 3D structured illumination microscopy. Moreover, we provide a validation and characterization of our method, demonstrating excellent preservation of chromatin structure and nuclear integrity, together with improved hybridization efficiency, compared with classic 3D-FISH protocols.

RevDate: 2022-05-12
CmpDate: 2022-04-12

Chathoth KT, Mikheeva LA, Crevel G, et al (2022)

The role of insulators and transcription in 3D chromatin organization of flies.

Genome research, 32(4):682-698.

The DNA in many organisms, including humans, is shown to be organized in topologically associating domains (TADs). In Drosophila, several architectural proteins are enriched at TAD borders, but it is still unclear whether these proteins play a functional role in the formation and maintenance of TADs. Here, we show that depletion of BEAF-32, Cp190, Chro, and Dref leads to changes in TAD organization and chromatin loops. Their depletion predominantly affects TAD borders located in regions moderately enriched in repressive modifications and depleted in active ones, whereas TAD borders located in euchromatin are resilient to these knockdowns. Furthermore, transcriptomic data has revealed hundreds of genes displaying differential expression in these knockdowns and showed that the majority of differentially expressed genes are located within reorganized TADs. Our work identifies a novel and functional role for architectural proteins at TAD borders in Drosophila and a link between TAD reorganization and subsequent changes in gene expression.

RevDate: 2022-05-16

Yin S, NandyMazumdar M, Paranjapye A, et al (2022)

Cross-talk between enhancers, structural elements and activating transcription factors maintains the 3D architecture and expression of the CFTR gene.

Genomics, 114(3):110350 pii:S0888-7543(22)00095-7 [Epub ahead of print].

Robust protocols to examine 3D chromatin structure have greatly advanced knowledge of gene regulatory mechanisms. Here we focus on the cystic fibrosis transmembrane conductance regulator (CFTR) gene, which provides a paradigm for validating models of gene regulation built upon genome-wide analysis. We examine the mechanisms by which multiple cis-regulatory elements (CREs) at the CFTR gene coordinate its expression in intestinal epithelial cells. Using CRISPR/Cas9 to remove CREs, individually and in tandem, followed by assays of gene expression and higher-order chromatin structure (4C-seq), we reveal the cross-talk and dependency of two cell-specific intronic enhancers. The results suggest a mechanism whereby the locus responds when CREs are lost, which may involve activating transcription factors such as FOXA2. Also, by removing the 5' topologically-associating domain (TAD) boundary, we illustrate its impact on CFTR gene expression and architecture. These data suggest a multi-layered regulatory hierarchy that is highly sensitive to perturbations.

RevDate: 2022-05-12
CmpDate: 2022-05-10

Paranjapye A, NandyMazumdar M, A Harris (2022)

Krüppel-Like Factor 5 Regulates CFTR Expression Through Repression by Maintaining Chromatin Architecture Coupled with Direct Enhancer Activation.

Journal of molecular biology, 434(10):167561.

Single cell RNA-sequencing has accurately identified cell types within the human airway that express the Cystic Fibrosis Transmembrane Conductance regulator (CFTR) gene. Low abundance CFTR transcripts are seen in many secretory cells, while high levels are restricted to rare pulmonary ionocytes. Here we focus on the mechanisms coordinating basal CFTR expression in the secretory compartment. Cell-selective regulation of CFTR is achieved within its invariant topologically associating domain by the recruitment of cis-regulatory elements (CREs). CRE activity is coordinated by cell-type-selective transcription factors. One such factor, Krüppel-Like Factor 5 (KLF5), profoundly represses CFTR transcript and protein in primary human airway epithelial cells and airway cell lines. Here we reveal the mechanism of action of KLF5 upon the CFTR gene. We find that depletion or ablation of KLF5 from airway epithelial cells changes higher order chromatin structure at the CFTR locus. Critical looping interactions that are required for normal gene expression are altered, the H3K27ac active chromatin mark is redistributed, and CTCF occupancy is modified. However, mutation of a single KLF5 binding site within a pivotal airway cell CRE abolishes CFTR expression. Hence, KLF5 has both direct activating and indirect repressive effects, which together coordinate CFTR expression in the airway.

RevDate: 2022-04-18
CmpDate: 2022-04-08

Girdhar K, Hoffman GE, Bendl J, et al (2022)

Chromatin domain alterations linked to 3D genome organization in a large cohort of schizophrenia and bipolar disorder brains.

Nature neuroscience, 25(4):474-483.

Chromosomal organization, scaling from the 147-base pair (bp) nucleosome to megabase-ranging domains encompassing multiple transcriptional units, including heritability loci for psychiatric traits, remains largely unexplored in the human brain. In this study, we constructed promoter- and enhancer-enriched nucleosomal histone modification landscapes for adult prefrontal cortex from H3-lysine 27 acetylation and H3-lysine 4 trimethylation profiles, generated from 388 controls and 351 individuals diagnosed with schizophrenia (SCZ) or bipolar disorder (BD) (n = 739). We mapped thousands of cis-regulatory domains (CRDs), revealing fine-grained, 104-106-bp chromosomal organization, firmly integrated into Hi-C topologically associating domain stratification by open/repressive chromosomal environments and nuclear topography. Large clusters of hyper-acetylated CRDs were enriched for SCZ heritability, with prominent representation of regulatory sequences governing fetal development and glutamatergic neuron signaling. Therefore, SCZ and BD brains show coordinated dysregulation of risk-associated regulatory sequences assembled into kilobase- to megabase-scaling chromosomal domains.

RevDate: 2022-04-27
CmpDate: 2022-04-27

Zhao C, Liu T, Z Wang (2022)

Functional Similarities of Protein-Coding Genes in Topologically Associating Domains and Spatially-Proximate Genomic Regions.

Genes, 13(3):.

Topologically associating domains (TADs) are the structural and functional units of the genome. However, the functions of protein-coding genes existing in the same or different TADs have not been fully investigated. We compared the functional similarities of protein-coding genes existing in the same TAD and between different TADs, and also in the same gap region (the region between two consecutive TADs) and between different gap regions. We found that the protein-coding genes from the same TAD or gap region are more likely to share similar protein functions, and this trend is more obvious with TADs than the gap regions. We further created two types of gene-gene spatial interaction networks: the first type is based on Hi-C contacts, whereas the second type is based on both Hi-C contacts and the relationship of being in the same TAD. A graph auto-encoder was applied to learn the network topology, reconstruct the two types of networks, and predict the functions of the central genes/nodes based on the functions of the neighboring genes/nodes. It was found that better performance was achieved with the second type of network. Furthermore, we detected long-range spatially-interactive regions based on Hi-C contacts and calculated the functional similarities of the gene pairs from these regions.

RevDate: 2022-05-02
CmpDate: 2022-05-02

Xia Y, Liu X, Mu W, et al (2022)

Capturing 3D Chromatin Maps of Human Primary Monocytes: Insights From High-Resolution Hi-C.

Frontiers in immunology, 13:837336.

Although the variation in chromatin architecture during adaptive immune responses has been thoroughly investigated, the 3D landscape of innate immunity is still unknown. Herein, chromatin regulation and heterogeneity among human primary monocytes were investigated. Peripheral blood was collected from two healthy persons and two patients with systemic lupus erythematosus (SLE), and CD14+ monocytes were selected to perform Hi-C, RNA-seq, ATAC-seq and ChIP-seq analyses. Raw data from the THP1 cell line Hi-C library were used for comparison. For each sample, we constructed three Hi-C libraries and obtained approximately 3 billion paired-end reads in total. Resolution analysis showed that more than 80% of bins presented depths greater than 1000 at a 5 kb resolution. The constructed high-resolution chromatin interaction maps presented similar landscapes in the four individuals, which showed significant divergence from the THP1 cell line chromatin structure. The variability in chromatin interactions around HLA-D genes in the HLA complex region was notable within individuals. We further found that the CD16-encoding gene (FCGR3A) is located at a variable topologically associating domain (TAD) boundary and that chromatin loop dynamics might modulate CD16 expression. Our results indicate both the stability and variability of high-resolution chromatin interaction maps among human primary monocytes. This work sheds light on the potential mechanisms by which the complex interplay of epigenetics and spatial 3D architecture regulates chromatin in innate immunity.

RevDate: 2022-05-04
CmpDate: 2022-05-04

Sanders JT, Golloshi R, Das P, et al (2022)

Loops, topologically associating domains, compartments, and territories are elastic and robust to dramatic nuclear volume swelling.

Scientific reports, 12(1):4721.

Layers of genome organization are becoming increasingly better characterized, but less is known about how these structures respond to perturbation or shape changes. Low-salt swelling of isolated chromatin fibers or nuclei has been used for decades to investigate the structural properties of chromatin. But, visible changes in chromatin appearance have not been linked to known building blocks of genome structure or features along the genome sequence. We combine low-salt swelling of isolated nuclei with genome-wide chromosome conformation capture (Hi-C) and imaging approaches to probe the effects of chromatin extension genome-wide. Photoconverted patterns on nuclei during expansion and contraction indicate that global genome structure is preserved after dramatic nuclear volume swelling, suggesting a highly elastic chromosome topology. Hi-C experiments before, during, and after nuclear swelling show changes in average contact probabilities at short length scales, reflecting the extension of the local chromatin fiber. But, surprisingly, during this large increase in nuclear volume, there is a striking maintenance of loops, TADs, active and inactive compartments, and chromosome territories. Subtle differences after expansion are observed, suggesting that the local chromatin state, protein interactions, and location in the nucleus can affect how strongly a given structure is maintained under stress. From these observations, we propose that genome topology is robust to extension of the chromatin fiber and isotropic shape change, and that this elasticity may be beneficial in physiological circumstances of changes in nuclear size and volume.

RevDate: 2022-03-22

Zhang J, Liu P, He M, et al (2022)

Reorganization of 3D genome architecture across wild boar and Bama pig adipose tissues.

Journal of animal science and biotechnology, 13(1):32.

BACKGROUND: A growing body of evidence has revealed that the mammalian genome is organized into hierarchical layers that are closely correlated with and may even be causally linked with variations in gene expression. Recent studies have characterized chromatin organization in various porcine tissues and cell types and compared them among species and during the early development of pigs. However, how chromatin organization differs among pig breeds is poorly understood.

RESULTS: In this study, we investigated the 3D genome organization and performed transcriptome characterization of two adipose depots (upper layer of backfat [ULB] and greater omentum [GOM]) in wild boars and Bama pigs; the latter is a typical indigenous pig in China. We found that over 95% of the A/B compartments and topologically associating domains (TADs) are stable between wild boars and Bama pigs. In contrast, more than 70% of promoter-enhancer interactions (PEIs) are dynamic and widespread, involving over a thousand genes. Alterations in chromatin structure are associated with changes in the expression of genes that are involved in widespread biological functions such as basic cellular functions, endocrine function, energy metabolism and the immune response. Approximately 95% and 97% of the genes associated with reorganized A/B compartments and PEIs in the two pig breeds differed between GOM and ULB, respectively.

CONCLUSIONS: We reported 3D genome organization in adipose depots from different pig breeds. In a comparison of Bama pigs and wild boar, large-scale compartments and TADs were mostly conserved, while fine-scale PEIs were extensively reorganized. The chromatin architecture in these two pig breeds was reorganized in an adipose depot-specific manner. These results contribute to determining the regulatory mechanism of phenotypic differences between Bama pigs and wild boar.

RevDate: 2022-04-30

Hicks P, O Oluwadare (2022)

HiCARN: Resolution Enhancement of Hi-C Data Using Cascading Residual Networks.

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

MOTIVATION: High throughput chromosome conformation capture (Hi-C) contact matrices are used to predict three-dimensional (3D) chromatin structures in eukaryotic cells. High resolution Hi-C data are less available than low resolution Hi-C data due to sequencing costs,but provide greater insight into the intricate details of 3D chromatin structures such as enhancer-promoter interactions and sub-domains. To provide a cost-effective solution to high resolution Hi-C data collection, deep learning models are used to predict high resolution Hi-C matrices from existing low-resolution matrices across multiple cell types.

RESULTS: Here, we present two Cascading Residual Networks called HiCARN-1 and HiCARN-2, a convolutional neural network and a generative adversarial network, that use a novel framework of cascading connections throughout the network for Hi-C contact matrix prediction from low-resolution data. Shown by image evaluation and Hi-C reproducibility metrics, both HiCARN models, overall, outperform state-of-the-art Hi-C resolution enhancement algorithms in predictive accuracy for both human and mouse 1/16, 1/32, 1/64, and 1/100 downsampled high-resolution Hi-C data. Also, validation by extracting topologically associating domains (TADs), chromosome 3D structure, and chromatin loop predictions from the enhanced data shows that HiCARN can proficiently reconstruct biologically significant regions.

AVAILABILITY: HiCARN can be accessed and utilized as an open-sourced software at: and is also available as a containerized application that can be run on any platform.

SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

RevDate: 2022-04-01

Galan S, Serra F, MA Marti-Renom (2022)

Identification of chromatin loops from Hi-C interaction matrices by CTCF-CTCF topology classification.

NAR genomics and bioinformatics, 4(1):lqac021.

Genome-wide profiling of long-range interactions has revealed that the CCCTC-Binding factor (CTCF) often anchors chromatin loops and is enriched at boundaries of the so-called Topologically Associating Domains, which suggests that CTCF is essential in the 3D organization of chromatin. However, the systematic topological classification of pairwise CTCF-CTCF interactions has not been yet explored. Here, we developed a computational pipeline able to classify all CTCF-CTCF pairs according to their chromatin interactions from Hi-C experiments. The interaction profiles of all CTCF-CTCF pairs were further structurally clustered using self-organizing feature maps and their functionality characterized by their epigenetic states. The resulting clusters were then input to a convolutional neural network aiming at the de novo detecting chromatin loops from Hi-C interaction matrices. Our new method, called LOOPbit, is able to automatically detect significant interactions with a higher proportion of enhancer-promoter loops compared to other callers. Our highly specific loop caller adds a new layer of detail to the link between chromatin structure and function.

RevDate: 2022-03-11
CmpDate: 2022-03-04

Mourad R (2022)

TADreg: a versatile regression framework for TAD identification, differential analysis and rearranged 3D genome prediction.

BMC bioinformatics, 23(1):82.

BACKGROUND/AIM: In higher eukaryotes, the three-dimensional (3D) organization of the genome is intimately related to numerous key biological functions including gene expression, DNA repair and DNA replication regulations. Alteration of 3D organization, in particular topologically associating domains (TADs), is detrimental to the organism and can give rise to a broad range of diseases such as cancers.

METHODS: Here, we propose a versatile regression framework which not only identifies TADs in a fast and accurate manner, but also detects differential TAD borders across conditions for which few methods exist, and predicts 3D genome reorganization after chromosomal rearrangement. Moreover, the framework is biologically meaningful, has an intuitive interpretation and is easy to visualize.

RESULT AND CONCLUSION: The novel regression ranks among top TAD callers. Moreover, it identifies new features of the genome we called TAD facilitators, and that are enriched with specific transcription factors. It also unveils the importance of cell-type specific transcription factors in establishing novel TAD borders during neuronal differentiation. Lastly, it compares favorably with the state-of-the-art method for predicting rearranged 3D genome.

RevDate: 2022-03-04

Pommier Y, Nussenzweig A, Takeda S, et al (2022)

Human topoisomerases and their roles in genome stability and organization.

Nature reviews. Molecular cell biology [Epub ahead of print].

Human topoisomerases comprise a family of six enzymes: two type IB (TOP1 and mitochondrial TOP1 (TOP1MT), two type IIA (TOP2A and TOP2B) and two type IA (TOP3A and TOP3B) topoisomerases. In this Review, we discuss their biochemistry and their roles in transcription, DNA replication and chromatin remodelling, and highlight the recent progress made in understanding TOP3A and TOP3B. Because of recent advances in elucidating the high-order organization of the genome through chromatin loops and topologically associating domains (TADs), we integrate the functions of topoisomerases with genome organization. We also discuss the physiological and pathological formation of irreversible topoisomerase cleavage complexes (TOPccs) as they generate topoisomerase DNA-protein crosslinks (TOP-DPCs) coupled with DNA breaks. We discuss the expanding number of redundant pathways that repair TOP-DPCs, and the defects in those pathways, which are increasingly recognized as source of genomic damage leading to neurological diseases and cancer.

RevDate: 2022-05-07
CmpDate: 2022-04-12

Franke M, Daly AF, Palmeira L, et al (2022)

Duplications disrupt chromatin architecture and rewire GPR101-enhancer communication in X-linked acrogigantism.

American journal of human genetics, 109(4):553-570.

X-linked acrogigantism (X-LAG) is the most severe form of pituitary gigantism and is characterized by aggressive growth hormone (GH)-secreting pituitary tumors that occur in early childhood. X-LAG is associated with chromosome Xq26.3 duplications (the X-LAG locus typically includes VGLL1, CD40LG, ARHGEF6, RBMX, and GPR101) that lead to massive pituitary tumoral expression of GPR101, a novel regulator of GH secretion. The mechanism by which the duplications lead to marked pituitary misexpression of GPR101 alone was previously unclear. Using Hi-C and 4C-seq, we characterized the normal chromatin structure at the X-LAG locus. We showed that GPR101 is located within a topologically associating domain (TAD) delineated by a tissue-invariant border that separates it from centromeric genes and regulatory sequences. Next, using 4C-seq with GPR101, RBMX, and VGLL1 viewpoints, we showed that the duplications in multiple X-LAG-affected individuals led to ectopic interactions that crossed the invariant TAD border, indicating the existence of a similar and consistent mechanism of neo-TAD formation in X-LAG. We then identified several pituitary active cis-regulatory elements (CREs) within the neo-TAD and demonstrated in vitro that one of them significantly enhanced reporter gene expression. At the same time, we showed that the GPR101 promoter permits the incorporation of new regulatory information. Our results indicate that X-LAG is a TADopathy of the endocrine system in which Xq26.3 duplications disrupt the local chromatin architecture forming a neo-TAD. Rewiring GPR101-enhancer interaction within the new regulatory unit is likely to cause the high levels of aberrant expression of GPR101 in pituitary tumors caused by X-LAG.

RevDate: 2022-04-11
CmpDate: 2022-04-11

Wu H, Zhang P, Ai Z, et al (2022)

StackTADB: a stacking-based ensemble learning model for predicting the boundaries of topologically associating domains (TADs) accurately in fruit flies.

Briefings in bioinformatics, 23(2):.

Chromosome is composed of many distinct chromatin domains, referred to variably as topological domains or topologically associating domains (TADs). The domains are stable across different cell types and highly conserved across species, thus these chromatin domains have been considered as the basic units of chromosome folding and regarded as an important secondary structure in chromosome organization. However, the identification of TAD boundaries is still a great challenge due to the high cost and low resolution of Hi-C data or experiments. In this study, we propose a novel ensemble learning framework, termed as StackTADB, for predicting the boundaries of TADs. StackTADB integrates four base classifiers including Random Forest, Logistic Regression, K-NearestNeighbor and Support Vector Machine. From the analysis of a series of examinations on the data set in the previous study, it is concluded that StackTADB has optimal performance in six metrics, AUC, Accuracy, MCC, Precision, Recall and F1 score, and it is superior to the existing methods. In addition, the comparison of the performance of multiple features shows that Kmers-based features play an essential role in predicting TADs boundaries of fruit flies, and we also apply the SHapley Additive exPlanations (SHAP) framework to interpret the predictions of StackTADB to identify the reason why Kmers-based features are vital. The experimental results show that the subsequences matching the BEAF-32 motif play a crucial role in predicting the boundaries of TADs. The source code is freely available at and the webserver of StackTADB is freely available at

RevDate: 2022-05-10
CmpDate: 2022-05-02

Osman N, Shawky AE, M Brylinski (2022)

Exploring the effects of genetic variation on gene regulation in cancer in the context of 3D genome structure.

BMC genomic data, 23(1):13.

BACKGROUND: Numerous genome-wide association studies (GWAS) conducted to date revealed genetic variants associated with various diseases, including breast and prostate cancers. Despite the availability of these large-scale data, relatively few variants have been functionally characterized, mainly because the majority of single-nucleotide polymorphisms (SNPs) map to the non-coding regions of the human genome. The functional characterization of these non-coding variants and the identification of their target genes remain challenging.

RESULTS: In this communication, we explore the potential functional mechanisms of non-coding SNPs by integrating GWAS with the high-resolution chromosome conformation capture (Hi-C) data for breast and prostate cancers. We show that more genetic variants map to regulatory elements through the 3D genome structure than the 1D linear genome lacking physical chromatin interactions. Importantly, the association of enhancers, transcription factors, and their target genes with breast and prostate cancers tends to be higher when these regulatory elements are mapped to high-risk SNPs through spatial interactions compared to simply using a linear proximity. Finally, we demonstrate that topologically associating domains (TADs) carrying high-risk SNPs also contain gene regulatory elements whose association with cancer is generally higher than those belonging to control TADs containing no high-risk variants.

CONCLUSIONS: Our results suggest that many SNPs may contribute to the cancer development by affecting the expression of certain tumor-related genes through long-range chromatin interactions with gene regulatory elements. Integrating large-scale genetic datasets with the 3D genome structure offers an attractive and unique approach to systematically investigate the functional mechanisms of genetic variants in disease risk and progression.

RevDate: 2022-05-04

Ilyin AA, Kononkova AD, Golova AV, et al (2022)

Comparison of genome architecture at two stages of male germline cell differentiation in Drosophila.

Nucleic acids research, 50(6):3203-3225.

Eukaryotic chromosomes are spatially segregated into topologically associating domains (TADs). Some TADs are attached to the nuclear lamina (NL) through lamina-associated domains (LADs). Here, we identified LADs and TADs at two stages of Drosophila spermatogenesis - in bamΔ86 mutant testes which is the commonly used model of spermatogonia (SpG) and in larval testes mainly filled with spermatocytes (SpCs). We found that initiation of SpC-specific transcription correlates with promoters' detachment from the NL and with local spatial insulation of adjacent regions. However, this insulation does not result in the partitioning of inactive TADs into sub-TADs. We also revealed an increased contact frequency between SpC-specific genes in SpCs implying their de novo gathering into transcription factories. In addition, we uncovered the specific X chromosome organization in the male germline. In SpG and SpCs, a single X chromosome is stronger associated with the NL than autosomes. Nevertheless, active chromatin regions in the X chromosome interact with each other more frequently than in autosomes. Moreover, despite the absence of dosage compensation complex in the male germline, randomly inserted SpG-specific reporter is expressed higher in the X chromosome than in autosomes, thus evidencing that non-canonical dosage compensation operates in SpG.

RevDate: 2022-05-12
CmpDate: 2022-02-25

Ortabozkoyun H, Huang PY, Cho H, et al (2022)

CRISPR and biochemical screens identify MAZ as a cofactor in CTCF-mediated insulation at Hox clusters.

Nature genetics, 54(2):202-212.

CCCTC-binding factor (CTCF) is critical to three-dimensional genome organization. Upon differentiation, CTCF insulates active and repressed genes within Hox gene clusters. We conducted a genome-wide CRISPR knockout (KO) screen to identify genes required for CTCF-boundary activity at the HoxA cluster, complemented by biochemical approaches. Among the candidates, we identified Myc-associated zinc-finger protein (MAZ) as a cofactor in CTCF insulation. MAZ colocalizes with CTCF at chromatin borders and, similar to CTCF, interacts with the cohesin subunit RAD21. MAZ KO disrupts gene expression and local contacts within topologically associating domains. Similar to CTCF motif deletions, MAZ motif deletions lead to derepression of posterior Hox genes immediately after CTCF boundaries upon differentiation, giving rise to homeotic transformations in mouse. Thus, MAZ is a factor contributing to appropriate insulation, gene expression and genomic architecture during development.

RevDate: 2022-02-16
CmpDate: 2022-02-14

Chang JM, Weng YF, Chang WT, et al (2022)

HiCmapTools: a tool to access HiC contact maps.

BMC bioinformatics, 23(1):64.

BACKGROUND: With the development of HiC technology, more and more HiC sequencing data have been produced. Although there are dozens of packages that can turn sequencing data into contact maps, there is no appropriate tool to query contact maps in order to extract biological information from HiC datasets.

RESULTS: We present HiCmapTools, a tool for biologists to efficiently calculate and analyze HiC maps. The complete program provides multi-query modes and analysis tools. We have validated its utility on two real biological questions: TAD loop and TAD intra-density.

CONCLUSIONS: HiCmapTools supports seven access options so that biologists can quantify contact frequency of the interest sites. The tool has been implemented in C++ and R and is freely available at and documented at .

RevDate: 2022-03-10
CmpDate: 2022-03-07

Owens DDG, Anselmi G, Oudelaar AM, et al (2022)

Dynamic Runx1 chromatin boundaries affect gene expression in hematopoietic development.

Nature communications, 13(1):773.

The transcription factor RUNX1 is a critical regulator of developmental hematopoiesis and is frequently disrupted in leukemia. Runx1 is a large, complex gene that is expressed from two alternative promoters under the spatiotemporal control of multiple hematopoietic enhancers. To dissect the dynamic regulation of Runx1 in hematopoietic development, we analyzed its three-dimensional chromatin conformation in mouse embryonic stem cell (ESC) differentiation cultures. Runx1 resides in a 1.1 Mb topologically associating domain (TAD) demarcated by convergent CTCF motifs. As ESCs differentiate to mesoderm, chromatin accessibility, Runx1 enhancer-promoter (E-P) interactions, and CTCF-CTCF interactions increase in the TAD, along with initiation of Runx1 expression from the P2 promoter. Differentiation to hematopoietic progenitor cells is associated with the formation of tissue-specific sub-TADs over Runx1, a shift in E-P interactions, P1 promoter demethylation, and robust expression from both Runx1 promoters. Deletion of promoter-proximal CTCF sites at the sub-TAD boundaries has no obvious effects on E-P interactions but leads to partial loss of domain structure, mildly affects gene expression, and delays hematopoietic development. Together, our analysis of gene regulation at a large multi-promoter developmental gene reveals that dynamic sub-TAD chromatin boundaries play a role in establishing TAD structure and coordinated gene expression.

RevDate: 2022-04-05
CmpDate: 2022-04-04

Pei L, Huang X, Liu Z, et al (2022)

Dynamic 3D genome architecture of cotton fiber reveals subgenome-coordinated chromatin topology for 4-staged single-cell differentiation.

Genome biology, 23(1):45.

BACKGROUND: Despite remarkable advances in our knowledge of epigenetically mediated transcriptional programming of cell differentiation in plants, little is known about chromatin topology and its functional implications in this process.

RESULTS: To interrogate its significance, we establish the dynamic three-dimensional (3D) genome architecture of the allotetraploid cotton fiber, representing a typical single cell undergoing staged development in plants. We show that the subgenome-relayed switching of the chromatin compartment from active to inactive is coupled with the silencing of developmentally repressed genes, pinpointing subgenome-coordinated contribution to fiber development. We identify 10,571 topologically associating domain-like (TAD-like) structures, of which 25.6% are specifically organized in different stages and 75.23% are subject to partition or fusion between two subgenomes. Notably, dissolution of intricate TAD-like structure cliques showing long-range interactions represents a prominent characteristic at the later developmental stage. Dynamic chromatin loops are found to mediate the rewiring of gene regulatory networks that exhibit a significant difference between the two subgenomes, implicating expression bias of homologous genes.

CONCLUSIONS: This study sheds light on the spatial-temporal asymmetric chromatin structures of two subgenomes in the cotton fiber and offers a new insight into the regulatory orchestration of cell differentiation in plants.

RevDate: 2022-02-11
CmpDate: 2022-02-11

Batut PJ, Bing XY, Sisco Z, et al (2022)

Genome organization controls transcriptional dynamics during development.

Science (New York, N.Y.), 375(6580):566-570.

Past studies offer contradictory claims for the role of genome organization in the regulation of gene activity. Here, we show through high-resolution chromosome conformation analysis that the Drosophila genome is organized by two independent classes of regulatory sequences, tethering elements and insulators. Quantitative live imaging and targeted genome editing demonstrate that this two-tiered organization is critical for the precise temporal dynamics of Hox gene transcription during development. Tethering elements mediate long-range enhancer-promoter interactions and foster fast activation kinetics. Conversely, the boundaries of topologically associating domains (TADs) prevent spurious interactions with enhancers and silencers located in neighboring TADs. These two levels of genome organization operate independently of one another to ensure precision of transcriptional dynamics and the reliability of complex patterning processes.

RevDate: 2022-02-01

Liu K, Li H, Li Y, et al (2022)

A comparison of topologically associating domain callers based on Hi-C data.

IEEE/ACM transactions on computational biology and bioinformatics, PP: [Epub ahead of print].

Topologically associating domains (TADs) are local chromatin interaction domains, which have been shown to play an important role in gene expression regulation. TADs were originally discovered in the investigation of 3D genome organization based on High-throughput Chromosome Conformation Capture (Hi-C) data. Continuous considerable efforts have been dedicated to developing methods for detecting TADs from Hi-C data. Different computational methods for TADs identification vary in their assumptions and criteria in calling TADs. As a consequence, the TADs called by these methods differ in their similarities and biological features they are enriched in. In this work, we performed a systematic comparison of twenty-four TAD callers. We first compared the TADs and gaps between adjacent TADs across different methods, resolutions, and sequencing depths. We then assessed the quality of TADs and TAD boundaries according to three criteria: the decay of contact frequencies over the genomic distance, enrichment and depletion of regulatory elements around TAD boundaries, and reproducibility of TADs and TAD boundaries in replicate samples. Last, due to the lack of a gold standard for TADs, we also evaluated the performance of the methods on synthetic datasets.

RevDate: 2022-02-01

Chu X, J Wang (2022)

Dynamics and Pathways of Chromosome Structural Organizations during Cell Transdifferentiation.

JACS Au, 2(1):116-127.

Direct conversion of one differentiated cell type into another is defined as cell transdifferentiation. In avoidance of forming pluripotency, cell transdifferentiation can reduce the potential risk of tumorigenicity, thus offering significant advantages over cell reprogramming in clinical applications. Until now, the mechanism of cell transdifferentiation is still largely unknown. It has been well recognized that cell transdifferentiation is determined by the underlying gene expression regulation, which relies on the accurate adaptation of the chromosome structure. To dissect the transdifferentiation at the molecular level, we develop a nonequilibrium landscape-switching model to investigate the chromosome structural dynamics during the state transitions between the human fibroblast and neuron cells. We uncover the high irreversibility of the transdifferentiation at the local chromosome structural ranges, where the topologically associating domains form. In contrast, the pathways in the two opposite directions of the transdifferentiation projected onto the chromosome compartment profiles are highly overlapped, indicating that the reversibility vanishes at the long-range chromosome structures. By calculating the contact strengths in the chromosome at the states along the paths, we observe strengthening contacts in compartment A concomitant with weakening contacts in compartment B at the early stages of the transdifferentiation. This further leads to adapting contacts toward the ones at the embryonic stem cell. In light of the intimate structure-function relationship at the chromosomal level, we suggest an increase of "stemness" during the transdifferentiation. In addition, we find that the neuron progenitor cell (NPC), a cell developmental state, is located on the transdifferentiation pathways projected onto the long-range chromosome contacts. The findings are consistent with the previous single-cell RNA sequencing experiment, where the NPC-like cell states were observed during the direct conversion of the fibroblast to neuron cells. Thus, we offer a promising microscopic and physical approach to study the cell transdifferentiation mechanism from the chromosome structural perspective.

RevDate: 2022-05-05
CmpDate: 2022-05-05

Long HS, Greenaway S, Powell G, et al (2022)

Making sense of the linear genome, gene function and TADs.

Epigenetics & chromatin, 15(1):4.

BACKGROUND: Topologically associating domains (TADs) are thought to act as functional units in the genome. TADs co-localise genes and their regulatory elements as well as forming the unit of genome switching between active and inactive compartments. This has led to the speculation that genes which are required for similar processes may fall within the same TADs, allowing them to share regulatory programs and efficiently switch between chromatin compartments. However, evidence to link genes within TADs to the same regulatory program is limited.

RESULTS: We investigated the functional similarity of genes which fall within the same TAD. To do this we developed a TAD randomisation algorithm to generate sets of "random TADs" to act as null distributions. We found that while pairs of paralogous genes are enriched in TADs overall, they are largely depleted in TADs with CCCTC-binding factor (CTCF) ChIP-seq peaks at both boundaries. By assessing gene constraint as a proxy for functional importance we found that genes which singly occupy a TAD have greater functional importance than genes which share a TAD, and these genes are enriched for developmental processes. We found little evidence that pairs of genes in CTCF bound TADs are more likely to be co-expressed or share functional annotations than can be explained by their linear proximity alone.

CONCLUSIONS: These results suggest that algorithmically defined TADs consist of two functionally different groups, those which are bound by CTCF and those which are not. We detected no association between genes sharing the same CTCF TADs and increased co-expression or functional similarity, other than that explained by linear genome proximity. We do, however, find that functionally important genes are more likely to fall within a TAD on their own suggesting that TADs play an important role in the insulation of these genes.

RevDate: 2022-05-03
CmpDate: 2022-05-03

Lybaek H, Robson M, de Leeuw N, et al (2022)

LRFN5 locus structure is associated with autism and influenced by the sex of the individual and locus conversions.

Autism research : official journal of the International Society for Autism Research, 15(3):421-433.

LRFN5 is a regulator of synaptic development and the only gene in a 5.4 Mb mammalian-specific conserved topologically associating domain (TAD); the LRFN5 locus. An association between locus structural changes and developmental delay (DD) and/or autism was suggested by several cases in DECIPHER and own records. More significantly, we found that maternal inheritance of a specific LRFN5 locus haplotype segregated with an identical type of autism in distantly related males. This autism-susceptibility haplotype had a specific TAD pattern. We also found a male/female quantitative difference in the amount histone-3-lysine-9-associated chromatin around the LRFN5 gene itself (p < 0.01), possibly related to the male-restricted autism susceptibility. To better understand locus behavior, the prevalence of a 60 kb deletion polymorphism was investigated. Surprisingly, in three cohorts of individuals with DD (n = 8757), the number of deletion heterozygotes was 20%-26% lower than expected from Hardy-Weinberg equilibrium. This suggests allelic interaction, also because the conversions from heterozygosity to wild-type or deletion homozygosity were of equal magnitudes. Remarkably, in a control group of medical students (n = 1416), such conversions were three times more common (p = 0.00001), suggesting a regulatory role of this allelic interaction. Taken together, LRFN5 regulation appears unusually complex, and LRFN5 dysregulation could be an epigenetic cause of autism. LAY SUMMARY: LRFN5 is involved with communication between brain cells. The gene sits alone in a huge genomic niche, called the LRFN5 locus, of complex structure and high mammalian conservation. We have found that a specific locus structure increases autism susceptibility in males, but we do not yet know how common this epigenetic cause of autism is. It is, however, a cause that potentially could explain why higher-functioning autism is more common in males than females.

RevDate: 2022-01-29

Wang H, Cui B, Sun H, et al (2021)

Aberrant GATA2 Activation in Pediatric B-Cell Acute Lymphoblastic Leukemia.

Frontiers in pediatrics, 9:795529.

GATA2 is a transcription factor that is critical for the generation and survival of hematopoietic stem cells (HSCs). It also plays an important role in the regulation of myeloid differentiation. Accordingly, GATA2 expression is restricted to HSCs and hematopoietic progenitors as well as early erythroid cells and megakaryocytic cells. Here we identified aberrant GATA2 expression in B-cell acute lymphoblastic leukemia (B-ALL) by analyzing transcriptome sequencing data obtained from St. Jude Cloud. Differentially expressed genes upon GATA2 activation showed significantly myeloid-like transcription signature. Further analysis identified several tumor-associated genes as targets of GATA2 activation including BAG3 and EPOR. In addition, the correlation between KMT2A-USP2 fusion and GATA2 activation not only indicates a potential trans-activating mechanism of GATA2 but also suggests that GATA2 is a target of KMT2A-USP2. Furthermore, by integrating whole-genome and transcriptome sequencing data, we showed that GATA2 is also cis activated. A somatic focal deletion located in the GATA2 neighborhood that disrupts the boundaries of topologically associating domains was identified in one B-ALL patient with GATA2 activation. These evidences support the hypothesis that GATA2 could be involved in leukemogenesis of B-ALL and can be transcriptionally activated through multiple mechanisms. The findings of aberrant activation of GATA2 and its molecular function extend our understanding of transcriptional factor dysregulation in B-ALL.

RevDate: 2022-05-03
CmpDate: 2022-05-03

Ing-Simmons E, Rigau M, JM Vaquerizas (2022)

Emerging mechanisms and dynamics of three-dimensional genome organisation at zygotic genome activation.

Current opinion in cell biology, 74:37-46.

The genome of an early embryo undergoes significant remodelling at the epigenetic, transcriptional, and structural levels. New technological developments have made it possible to study 3D genome organisation in the zygote and early embryo of many different species. Recent studies in human embryos, zebrafish, medaka, and Xenopus have revealed that, similar to previous results in mouse and Drosophila, the zygotic genome is unstructured prior to zygotic genome activation. While these studies show that topologically associating domains are established coincident with zygotic genome activation across species, other 3D genome structures have more varied timing. Here, we review recent studies examining the timing and mechanisms of establishment of 3D genome organisation in the early embryo, and discuss similarities and differences between species. Investigating the establishment of 3D chromatin conformation in early embryos has the potential to reveal novel mechanisms of 3D genome organisation.

RevDate: 2022-01-21

Tsagiopoulou M, Pechlivanis N, Maniou MC, et al (2022)

InterTADs: integration of multi-omics data on topologically associated domains, application to chronic lymphocytic leukemia.

NAR genomics and bioinformatics, 4(1):lqab121.

The integration of multi-omics data can greatly facilitate the advancement of research in Life Sciences by highlighting new interactions. However, there is currently no widespread procedure for meaningful multi-omics data integration. Here, we present a robust framework, called InterTADs, for integrating multi-omics data derived from the same sample, and considering the chromatin configuration of the genome, i.e. the topologically associating domains (TADs). Following the integration process, statistical analysis highlights the differences between the groups of interest (normal versus cancer cells) relating to (i) independent and (ii) integrated events through TADs. Finally, enrichment analysis using KEGG database, Gene Ontology and transcription factor binding sites and visualization approaches are available. We applied InterTADs to multi-omics datasets from 135 patients with chronic lymphocytic leukemia (CLL) and found that the integration through TADs resulted in a dramatic reduction of heterogeneity compared to individual events. Significant differences for individual events and on TADs level were identified between patients differing in the somatic hypermutation status of the clonotypic immunoglobulin genes, the core biological stratifier in CLL, attesting to the biomedical relevance of InterTADs. In conclusion, our approach suggests a new perspective towards analyzing multi-omics data, by offering reasonable execution time, biological benchmarking and potentially contributing to pattern discovery through TADs.

RevDate: 2022-02-15
CmpDate: 2022-02-15

Li CC, Zhang G, Du J, et al (2022)

Pre-configuring chromatin architecture with histone modifications guides hematopoietic stem cell formation in mouse embryos.

Nature communications, 13(1):346.

The gene activity underlying cell differentiation is regulated by a diverse set of transcription factors (TFs), histone modifications, chromatin structures and more. Although definitive hematopoietic stem cells (HSCs) are known to emerge via endothelial-to-hematopoietic transition (EHT), how the multi-layered epigenome is sequentially unfolded in a small portion of endothelial cells (ECs) transitioning into the hematopoietic fate remains elusive. With optimized low-input itChIP-seq and Hi-C assays, we performed multi-omics dissection of the HSC ontogeny trajectory across early arterial ECs (eAECs), hemogenic endothelial cells (HECs), pre-HSCs and long-term HSCs (LT-HSCs) in mouse embryos. Interestingly, HSC regulatory regions are already pre-configurated with active histone modifications as early as eAECs, preceding chromatin looping dynamics within topologically associating domains. Chromatin looping structures between enhancers and promoters only become gradually strengthened over time. Notably, RUNX1, a master TF for hematopoiesis, enriched at half of these loops is observed early from eAECs through pre-HSCs but its enrichment further increases in HSCs. RUNX1 and co-TFs together constitute a central, progressively intensified enhancer-promoter interactions. Thus, our study provides a framework to decipher how temporal epigenomic configurations fulfill cell lineage specification during development.

RevDate: 2022-02-01

Miura H, I Hiratani (2022)

Cell cycle dynamics and developmental dynamics of the 3D genome: toward linking the two timescales.

Current opinion in genetics & development, 73:101898 pii:S0959-437X(21)00143-X [Epub ahead of print].

In the mammalian cell nucleus, chromosomes are folded differently in interphase and mitosis. Interphase chromosomes are relatively decondensed and display at least two unique layers of higher-order organization: topologically associating domains (TADs) and cell-type-specific A/B compartments, which correlate well with early/late DNA replication timing (RT). In mitosis, these structures rapidly disappear but are gradually reconstructed during G1 phase, coincident with the establishment of the RT program. However, these structures also change dynamically during cell differentiation and reprogramming, and yet we are surprisingly ignorant about the relationship between their cell cycle dynamics and developmental dynamics. In this review, we summarize the recent findings on this topic, discuss how these two processes might be coordinated with each other and its potential significance.

RevDate: 2022-03-16
CmpDate: 2022-03-16

Jablonski KP, Carron L, Mozziconacci J, et al (2022)

Contribution of 3D genome topological domains to genetic risk of cancers: a genome-wide computational study.

Human genomics, 16(1):2.

BACKGROUND: Genome-wide association studies have identified statistical associations between various diseases, including cancers, and a large number of single-nucleotide polymorphisms (SNPs). However, they provide no direct explanation of the mechanisms underlying the association. Based on the recent discovery that changes in three-dimensional genome organization may have functional consequences on gene regulation favoring diseases, we investigated systematically the genome-wide distribution of disease-associated SNPs with respect to a specific feature of 3D genome organization: topologically associating domains (TADs) and their borders.

RESULTS: For each of 449 diseases, we tested whether the associated SNPs are present in TAD borders more often than observed by chance, where chance (i.e., the null model in statistical terms) corresponds to the same number of pointwise loci drawn at random either in the entire genome, or in the entire set of disease-associated SNPs listed in the GWAS catalog. Our analysis shows that a fraction of diseases displays such a preferential localization of their risk loci. Moreover, cancers are relatively more frequent among these diseases, and this predominance is generally enhanced when considering only intergenic SNPs. The structure of SNP-based diseasome networks confirms that localization of risk loci in TAD borders differs between cancers and non-cancer diseases. Furthermore, different TAD border enrichments are observed in embryonic stem cells and differentiated cells, consistent with changes in topological domains along embryogenesis and delineating their contribution to disease risk.

CONCLUSIONS: Our results suggest that, for certain diseases, part of the genetic risk lies in a local genetic variation affecting the genome partitioning in topologically insulated domains. Investigating this possible contribution to genetic risk is particularly relevant in cancers. This study thus opens a way of interpreting genome-wide association studies, by distinguishing two types of disease-associated SNPs: one with an effect on an individual gene, the other acting in interplay with 3D genome organization.

RevDate: 2022-01-25
CmpDate: 2022-01-25

Li D, Ning C, Zhang J, et al (2022)

Dynamic transcriptome and chromatin architecture in granulosa cells during chicken folliculogenesis.

Nature communications, 13(1):131.

Folliculogenesis is a complex biological process involving a central oocyte and its surrounding somatic cells. Three-dimensional chromatin architecture is an important transcription regulator; however, little is known about its dynamics and role in transcriptional regulation of granulosa cells during chicken folliculogenesis. We investigate the transcriptomic dynamics of chicken granulosa cells over ten follicular stages and assess the chromatin architecture dynamics and how it influences gene expression in granulosa cells at three key stages: the prehierarchical small white follicles, the first largest preovulatory follicles, and the postovulatory follicles. Our results demonstrate the consistency between the global reprogramming of chromatin architecture and the transcriptomic divergence during folliculogenesis, providing ample evidence for compartmentalization rearrangement, variable organization of topologically associating domains, and rewiring of the long-range interaction between promoter and enhancers. These results provide key insights into avian reproductive biology and provide a foundational dataset for the future in-depth functional characterization of granulosa cells.

RevDate: 2022-03-02
CmpDate: 2022-01-12

Tian GG, Zhao X, Hou C, et al (2022)

Integrative analysis of the 3D genome structure reveals that CTCF maintains the properties of mouse female germline stem cells.

Cellular and molecular life sciences : CMLS, 79(1):22.

The three-dimensional configuration of the genome ensures cell type-specific gene expression profiles by placing genes and regulatory elements in close spatial proximity. Here, we used in situ high-throughput chromosome conformation (in situ Hi-C), RNA sequencing (RNA-seq) and chromatin immunoprecipitation sequencing (ChIP-seq) to characterize the high-order chromatin structure signature of female germline stem cells (FGSCs) and identify its regulating key factor based on the data-driven of multiple omics data. By comparison with pluripotent stem cells (PSCs), adult stem cells (ASCs), and somatic cells at three major levels of chromatin architecture, A/B compartments, topologically associating domains, and chromatin loops, the chromatin architecture of FGSCs was most similar to that of other ASCs and largely different from that of PSCs and somatic cells. After integrative analysis of the three-dimensional chromatin structure, active compartment-associating loops (aCALs) were identified as a signature of high-order chromatin organization in FGSCs, which revealed that CCCTC-binding factor was a major factor to maintain the properties of FGSCs through regulation of aCALs. We found FGSCs belong to ASCs at chromatin structure level and characterized aCALs as the high-order chromatin structure signature of FGSCs. Furthermore, CTCF was identified to play a key role in regulating aCALS to maintain the biological functions of FGSCs. These data provide a valuable resource for future studies of the features of chromatin organization in mammalian stem cells and further understanding of the fundamental characteristics of FGSCs.

RevDate: 2022-05-11
CmpDate: 2022-04-29

Zheng Y, Zhang L, Jin L, et al (2022)

Unraveling three-dimensional chromatin structural dynamics during spermatogonial differentiation.

The Journal of biological chemistry, 298(2):101559.

Spermatogonial stem cells (SSCs) are able to undergo both self-renewal and differentiation. Unlike self-renewal, which replenishes the SSC and progenitor pool, differentiation is an irreversible process committing cells to meiosis. Although the preparations for meiotic events in differentiating spermatogonia (Di-SG) are likely to be accompanied by alterations in chromatin structure, the three-dimensional chromatin architectural differences between SSCs and Di-SG, and the higher-order chromatin dynamics during spermatogonial differentiation, have not been systematically investigated. Here, we performed in situ high-throughput chromosome conformation capture, RNA-seq, and chromatin immunoprecipitation-sequencing analyses on porcine undifferentiated spermatogonia (which consist of SSCs and progenitors) and Di-SG. We identified that Di-SG exhibited less compact chromatin structural organization, weakened compartmentalization, and diminished topologically associating domains in comparison with undifferentiated spermatogonia, suggesting that diminished higher-order chromatin architecture in meiotic cells, as shown by recent reports, might be preprogrammed in Di-SG. Our data also revealed that A/B compartments, representing open or closed chromatin regions respectively, and topologically associating domains were related to dynamic gene expression during spermatogonial differentiation. Furthermore, we unraveled the contribution of promoter-enhancer interactions to premeiotic transcriptional regulation, which has not been accomplished in previous studies due to limited cell input and resolution. Together, our study uncovered the three-dimensional chromatin structure of SSCs/progenitors and Di-SG, as well as the interplay between higher-order chromatin architecture and dynamic gene expression during spermatogonial differentiation. These findings provide novel insights into the mechanisms for SSC self-renewal and differentiation and have implications for diagnosis and treatment of male sub-/infertility.

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

Salari H, Di Stefano M, D Jost (2022)

Spatial organization of chromosomes leads to heterogeneous chromatin motion and drives the liquid- or gel-like dynamical behavior of chromatin.

Genome research, 32(1):28-43.

Chromosome organization and dynamics are involved in regulating many fundamental processes such as gene transcription and DNA repair. Experiments unveiled that chromatin motion is highly heterogeneous inside cell nuclei, ranging from a liquid-like, mobile state to a gel-like, rigid regime. Using polymer modeling, we investigate how these different physical states and dynamical heterogeneities may emerge from the same structural mechanisms. We found that the formation of topologically associating domains (TADs) is a key driver of chromatin motion heterogeneity. In particular, we showed that the local degree of compaction of the TAD regulates the transition from a weakly compact, fluid state of chromatin to a more compact, gel state exhibiting anomalous diffusion and coherent motion. Our work provides a comprehensive study of chromosome dynamics and a unified view of chromatin motion enabling interpretation of the wide variety of dynamical behaviors observed experimentally across different biological conditions, suggesting that the "liquid" or "solid" state of chromatin are in fact two sides of the same coin.

RevDate: 2021-12-28

Kumar S, Kaur S, Seem K, et al (2021)

Understanding 3D Genome Organization and Its Effect on Transcriptional Gene Regulation Under Environmental Stress in Plant: A Chromatin Perspective.

Frontiers in cell and developmental biology, 9:774719.

The genome of a eukaryotic organism is comprised of a supra-molecular complex of chromatin fibers and intricately folded three-dimensional (3D) structures. Chromosomal interactions and topological changes in response to the developmental and/or environmental stimuli affect gene expression. Chromatin architecture plays important roles in DNA replication, gene expression, and genome integrity. Higher-order chromatin organizations like chromosome territories (CTs), A/B compartments, topologically associating domains (TADs), and chromatin loops vary among cells, tissues, and species depending on the developmental stage and/or environmental conditions (4D genomics). Every chromosome occupies a separate territory in the interphase nucleus and forms the top layer of hierarchical structure (CTs) in most of the eukaryotes. While the A and B compartments are associated with active (euchromatic) and inactive (heterochromatic) chromatin, respectively, having well-defined genomic/epigenomic features, TADs are the structural units of chromatin. Chromatin architecture like TADs as well as the local interactions between promoter and regulatory elements correlates with the chromatin activity, which alters during environmental stresses due to relocalization of the architectural proteins. Moreover, chromatin looping brings the gene and regulatory elements in close proximity for interactions. The intricate relationship between nucleotide sequence and chromatin architecture requires a more comprehensive understanding to unravel the genome organization and genetic plasticity. During the last decade, advances in chromatin conformation capture techniques for unravelling 3D genome organizations have improved our understanding of genome biology. However, the recent advances, such as Hi-C and ChIA-PET, have substantially increased the resolution, throughput as well our interest in analysing genome organizations. The present review provides an overview of the historical and contemporary perspectives of chromosome conformation capture technologies, their applications in functional genomics, and the constraints in predicting 3D genome organization. We also discuss the future perspectives of understanding high-order chromatin organizations in deciphering transcriptional regulation of gene expression under environmental stress (4D genomics). These might help design the climate-smart crop to meet the ever-growing demands of food, feed, and fodder.

RevDate: 2021-12-27
CmpDate: 2021-12-27

Conte M, Fiorillo L, Annunziatella C, et al (2021)

Dynamic and equilibrium properties of finite-size polymer models of chromosome folding.

Physical review. E, 104(5-1):054402.

Novel technologies are revealing that chromosomes have a complex three-dimensional organization within the cell nucleus that serves functional purposes. Models from polymer physics have been developed to quantitively understand the molecular principles controlling their structure and folding mechanisms. Here, by using massive molecular-dynamics simulations we show that classical scaling laws combined with finite-size effects of a simple polymer model can effectively explain the scaling behavior that chromatin exhibits at the topologically associating domains level, as revealed by experimental observations. Model results are then validated against recently published high-resolution in situ Hi-C data.

RevDate: 2022-03-24
CmpDate: 2022-03-10

Huang N, Seow WQ, Appert A, et al (2022)

Accessible Region Conformation Capture (ARC-C) gives high-resolution insights into genome architecture and regulation.

Genome research, 32(2):357-366.

Nuclear organization and chromatin interactions are important for genome function, yet determining chromatin connections at high resolution remains a major challenge. To address this, we developed Accessible Region Conformation Capture (ARC-C), which profiles interactions between regulatory elements genome-wide without a capture step. Applied to Caenorhabditis elegans, ARC-C identifies approximately 15,000 significant interactions between regulatory elements at 500-bp resolution. Of 105 TFs or chromatin regulators tested, we find that the binding sites of 60 are enriched for interacting with each other, making them candidates for mediating interactions. These include cohesin and condensin II. Applying ARC-C to a mutant of transcription factor BLMP-1 detected changes in interactions between its targets. ARC-C simultaneously profiles domain-level architecture, and we observe that C. elegans chromatin domains defined by either active or repressive modifications form topologically associating domains (TADs) that interact with A/B (active/inactive) compartment-like structure. Furthermore, we discover that inactive compartment interactions are dependent on H3K9 methylation. ARC-C is a powerful new tool to interrogate genome architecture and regulatory interactions at high resolution.

RevDate: 2021-12-17

Tian W, Wang Z, Wang D, et al (2021)

Chromatin Interaction Responds to Breast Muscle Development and Intramuscular Fat Deposition Between Chinese Indigenous Chicken and Fast-Growing Broiler.

Frontiers in cell and developmental biology, 9:782268.

Skeletal muscle development and intramuscular fat (IMF) content, which positively contribute to meat production and quality, are regulated by precisely orchestrated processes. However, changes in three-dimensional chromatin structure and interaction, a newly emerged mediator of gene expression, during the skeletal muscle development and IMF deposition have remained unclear. In the present study, we analyzed the differences in muscle development and IMF content between one-day-old commercial Arbor Acres broiler (AA) and Chinese indigenous Lushi blue-shelled-egg chicken (LS) and performed Hi-C analysis on their breast muscles. Our results indicated that significantly higher IMF content, however remarkably lower muscle fiber diameter was detected in breast muscle of LS chicken compared to that of AA broiler. The chromatin intra-interaction was prior to inter-interaction in both AA and LS chicken, and chromatin inter-interaction was heavily focused on the small and gene-rich chromosomes. For genomic compartmentalization, no significant difference in the number of B type compartments was found, but AA had more A type compartments versus LS. The A/B compartment switching of AA versus LS showed more A to B switching than B to A switching. There were no significant differences in the average sizes and distributions of topologically associating domains (TAD). Additionally, approximately 50% of TAD boundaries were overlapping. The reforming and disappearing events of TAD boundaries were identified between AA and LS chicken breast muscles. Among these, the HMGCR gene was located in the TAD-boundary regions in AA broilers, but in TAD-interior regions in LS chickens, and the IGF2BP3 gene was located in the AA-unique TAD boundaries. Both HMGCR and IGF2BP3 genes exhibited increased mRNA expression in one-day-old AA broiler breast muscles. It was demonstrated that the IGF2BP3 and HMGCR genes regulated by TAD boundary sliding were potential biomarkers for chicken breast muscle development and IMF deposition. Our data not only provide a valuable understanding of higher-order chromatin dynamics during muscle development and lipid accumulation but also reveal new insights into the regulatory mechanisms of muscle development and IMF deposition in chicken.

RevDate: 2022-03-22
CmpDate: 2021-12-22

San Martin R, Das P, Dos Reis Marques R, et al (2022)

Chromosome compartmentalization alterations in prostate cancer cell lines model disease progression.

The Journal of cell biology, 221(2):.

Prostate cancer aggressiveness and metastatic potential are influenced by gene expression and genomic aberrations, features that can be influenced by the 3D structure of chromosomes inside the nucleus. Using chromosome conformation capture (Hi-C), we conducted a systematic genome architecture comparison on a cohort of cell lines that model prostate cancer progression, from normal epithelium to bone metastasis. We describe spatial compartment identity (A-open versus B-closed) changes with progression in these cell lines and their relation to gene expression changes in both cell lines and patient samples. In particular, 48 gene clusters switch from the B to the A compartment, including androgen receptor, WNT5A, and CDK14. These switches are accompanied by changes in the structure, size, and boundaries of topologically associating domains (TADs). Further, compartment changes in chromosome 21 are exacerbated with progression and may explain, in part, the genesis of the TMPRSS2-ERG translocation. These results suggest that discrete 3D genome structure changes play a deleterious role in prostate cancer progression. .

RevDate: 2021-12-07

Cardozo Gizzi AM (2021)

A Shift in Paradigms: Spatial Genomics Approaches to Reveal Single-Cell Principles of Genome Organization.

Frontiers in genetics, 12:780822.

The genome tridimensional (3D) organization and its role towards the regulation of key cell processes such as transcription is currently a main question in biology. Interphase chromosomes are spatially segregated into "territories," epigenetically-defined large domains of chromatin that interact to form "compartments" with common transcriptional status, and insulator-flanked domains called "topologically associating domains" (TADs). Moreover, chromatin organizes around nuclear structures such as lamina, speckles, or the nucleolus to acquire a higher-order genome organization. Due to recent technological advances, the different hierarchies are being solved. Particularly, advances in microscopy technologies are shedding light on the genome structure at multiple levels. Intriguingly, more and more reports point to high variability and stochasticity at the single-cell level. However, the functional consequences of such variability in genome conformation are still unsolved. Here, I will discuss the implication of the cell-to-cell heterogeneity at the different scales in the context of newly developed imaging approaches, particularly multiplexed Fluorescence in situ hybridization methods that enabled "chromatin tracing." Extensions of these methods are now combining spatial information of dozens to thousands of genomic loci with the localization of nuclear features such as the nucleolus, nuclear speckles, or even histone modifications, creating the fast-moving field of "spatial genomics." As our view of genome organization shifts the focus from ensemble to single-cell, new insights to fundamental questions begin to emerge.

RevDate: 2022-02-04
CmpDate: 2022-02-04

Cinque L, Micale L, Manara E, et al (2022)

A novel complex genomic rearrangement affecting the KCNJ2 regulatory region causes a variant of Cooks syndrome.

Human genetics, 141(2):217-227.

Cooks syndrome (CS) is an ultrarare limb malformation due to in tandem microduplications involving KCNJ2 and extending to the 5' regulatory element of SOX9. To date, six CS families were resolved at the molecular level. Subsequent studies explored the evolutionary and pathological complexities of the SOX9-KCNJ2/Sox9-Kcnj2 locus, and suggested a key role for the formation of novel topologically associating domain (TAD) by inter-TAD duplications in causing CS. Here, we report a unique case of CS associated with a de novo 1;17 translocation affecting the KCNJ2 locus. On chromosome 17, the breakpoint mapped between KCNJ16 and KCNJ2, and combined with a ~ 5 kb deletion in the 5' of KCNJ2. Based on available capture Hi-C data, the breakpoint on chromosome 17 separated KCNJ2 from a putative enhancer. Gene expression analysis demonstrated downregulation of KCNJ2 in both patient's blood cells and cultured skin fibroblasts. Our findings suggest that a complex rearrangement falling in the 5' of KCNJ2 may mimic the developmental consequences of in tandem duplications affecting the SOX9-KCNJ2/Sox9-Kcnj2 locus. This finding adds weight to the notion of an intricate role of gene regulatory regions and, presumably, the related three-dimensional chromatin structure in normal and abnormal human morphology.

RevDate: 2021-11-23

Maslova A, A Krasikova (2021)

FISH Going Meso-Scale: A Microscopic Search for Chromatin Domains.

Frontiers in cell and developmental biology, 9:753097.

The intimate relationships between genome structure and function direct efforts toward deciphering three-dimensional chromatin organization within the interphase nuclei at different genomic length scales. For decades, major insights into chromatin structure at the level of large-scale euchromatin and heterochromatin compartments, chromosome territories, and subchromosomal regions resulted from the evolution of light microscopy and fluorescence in situ hybridization. Studies of nanoscale nucleosomal chromatin organization benefited from a variety of electron microscopy techniques. Recent breakthroughs in the investigation of mesoscale chromatin structures have emerged from chromatin conformation capture methods (C-methods). Chromatin has been found to form hierarchical domains with high frequency of local interactions from loop domains to topologically associating domains and compartments. During the last decade, advances in super-resolution light microscopy made these levels of chromatin folding amenable for microscopic examination. Here we are reviewing recent developments in FISH-based approaches for detection, quantitative measurements, and validation of contact chromatin domains deduced from C-based data. We specifically focus on the design and application of Oligopaint probes, which marked the latest progress in the imaging of chromatin domains. Vivid examples of chromatin domain FISH-visualization by means of conventional, super-resolution light and electron microscopy in different model organisms are provided.

RevDate: 2022-03-25
CmpDate: 2022-03-25

Wu DY, Li X, Sun QR, et al (2021)

Defective chromatin architectures in embryonic stem cells derived from somatic cell nuclear transfer impair their differentiation potentials.

Cell death & disease, 12(12):1085.

Nuclear transfer embryonic stem cells (ntESCs) hold enormous promise for individual-specific regenerative medicine. However, the chromatin states of ntESCs remain poorly characterized. In this study, we employed ATAC-seq and Hi-C techniques to explore the chromatin accessibility and three-dimensional (3D) genome organization of ntESCs. The results show that the chromatin accessibility and genome structures of somatic cells are re-arranged to ESC-like states overall in ntESCs, including compartments, topologically associating domains (TADs) and chromatin loops. However, compared to fertilized ESCs (fESCs), ntESCs show some abnormal openness and structures that have not been reprogrammed completely, which impair the differentiation potential of ntESCs. The histone modification H3K9me3 may be involved in abnormal structures in ntESCs, including incorrect compartment switches and incomplete TAD rebuilding. Moreover, ntESCs and iPSCs show high similarity in 3D genome structures, while a few differences are detected due to different somatic cell origins and reprogramming mechanisms. Through systematic analyses, our study provides a global view of chromatin accessibility and 3D genome organization in ntESCs, which can further facilitate the understanding of the similarities and differences between ntESCs and fESCs.

RevDate: 2022-03-18
CmpDate: 2022-01-27

Cheng Y, Liu M, Hu M, et al (2021)

TAD-like single-cell domain structures exist on both active and inactive X chromosomes and persist under epigenetic perturbations.

Genome biology, 22(1):309.

BACKGROUND: Topologically associating domains (TADs) are important building blocks of three-dimensional genome architectures. The formation of TADs has been shown to depend on cohesin in a loop-extrusion mechanism. Recently, advances in an image-based spatial genomics technique known as chromatin tracing lead to the discovery of cohesin-independent TAD-like structures, also known as single-cell domains, which are highly variant self-interacting chromatin domains with boundaries that occasionally overlap with TAD boundaries but tend to differ among single cells and among single chromosome copies. Recent computational modeling studies suggest that epigenetic interactions may underlie the formation of the single-cell domains.

RESULTS: Here we use chromatin tracing to visualize in female human cells the fine-scale chromatin folding of inactive and active X chromosomes, which are known to have distinct global epigenetic landscapes and distinct population-averaged TAD profiles, with inactive X chromosomes largely devoid of TADs and cohesin. We show that both inactive and active X chromosomes possess highly variant single-cell domains across the same genomic region despite the fact that only active X chromosomes show clear TAD structures at the population level. These X chromosome single-cell domains exist in distinct cell lines. Perturbations of major epigenetic components and transcription mostly do not affect the frequency or strength of the single-cell domains. Increased chromatin compaction of inactive X chromosomes occurs at a length scale above that of the single-cell domains.

CONCLUSIONS: In sum, this study suggests that single-cell domains are genome architecture building blocks independent of the tested major epigenetic components.

RevDate: 2022-03-24
CmpDate: 2022-03-24

Yildirir G, Sperschneider J, Malar C M, et al (2022)

Long reads and Hi-C sequencing illuminate the two-compartment genome of the model arbuscular mycorrhizal symbiont Rhizophagus irregularis.

The New phytologist, 233(3):1097-1107.

Chromosome folding links genome structure with gene function by generating distinct nuclear compartments and topologically associating domains. In mammals, these undergo preferential interactions and regulate gene expression. However, their role in fungal genome biology is unclear. Here, we combine Nanopore (ONT) sequencing with chromatin conformation capture sequencing (Hi-C) to reveal chromosome and epigenetic diversity in a group of obligate plant symbionts: the arbuscular mycorrhizal fungi (AMF). We find that five phylogenetically distinct strains of the model AMF Rhizophagus irregularis carry 33 chromosomes with substantial within-species variability in size, as well as in gene and repeat content. Strain-specific Hi-C contact maps reveal a 'checkerboard' pattern that underline two dominant euchromatin (A) and heterochromatin (B) compartments. Each compartment differs in the level of gene transcription, regulation of candidate effectors and methylation frequencies. The A-compartment is more gene-dense and contains most core genes, while the B-compartment is more repeat-rich and has higher rates of chromosomal rearrangement. While the B-compartment is transcriptionally repressed, it has significantly more secreted proteins and in planta upregulated candidate effectors, suggesting a possible host-induced change in chromosome conformation. Overall, this study provides a fine-scale view into the genome biology and evolution of model plant symbionts, and opens avenues to study the epigenetic mechanisms that modify chromosome folding during host-microbe interactions.

RevDate: 2022-03-30

Stilianoudakis SC, Marshall MA, MG Dozmorov (2021)

preciseTAD: A transfer learning framework for 3D domain boundary prediction at base-pair resolution.

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

MOTIVATION: Chromosome conformation capture technologies (Hi-C) revealed extensive DNA folding into discrete 3D domains, such as Topologically Associating Domains and chromatin loops. The correct binding of CTCF and cohesin at domain boundaries is integral in maintaining the proper structure and function of these 3D domains. 3D domains have been mapped at the resolutions of 1 kilobase and above. However, it has not been possible to define their boundaries at the resolution of boundary-forming proteins.

RESULTS: To predict domain boundaries at base-pair resolution, we developed preciseTAD, an optimized transfer learning framework trained on high-resolution genome annotation data. In contrast to current TAD/loop callers, preciseTAD-predicted boundaries are strongly supported by experimental evidence. Importantly, this approach can accurately delineate boundaries in cells without Hi-C data. preciseTAD provides a powerful framework to improve our understanding of how genomic regulators are shaping the 3D structure of the genome at base-pair resolution.

AVAILABILITY: preciseTAD is an R/Bioconductor package available at

SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

RevDate: 2021-11-02

Liehr T (2021)

Molecular Cytogenetics in the Era of Chromosomics and Cytogenomic Approaches.

Frontiers in genetics, 12:720507.

Here the role of molecular cytogenetics in the context of yet available all other cytogenomic approaches is discussed. A short introduction how cytogenetics and molecular cytogenetics were established is followed by technical aspects of fluorescence in situ hybridization (FISH). The latter contains the methodology itself, the types of probe- and target-DNA, as well as probe sets. The main part deals with examples of modern FISH-applications, highlighting unique possibilities of the approach, like the possibility to study individual cells and even individual chromosomes. Different variants of FISH can be used to retrieve information on genomes from (almost) base pair to whole genomic level, as besides only second and third generation sequencing approaches can do. Here especially highlighted variations of FISH are molecular combing, chromosome orientation-FISH (CO-FISH), telomere-FISH, parental origin determination FISH (POD-FISH), FISH to resolve the nuclear architecture, multicolor-FISH (mFISH) approaches, among other applied in chromoanagenesis studies, Comet-FISH, and CRISPR-mediated FISH-applications. Overall, molecular cytogenetics is far from being outdated and actively involved in up-to-date diagnostics and research.

RevDate: 2022-05-12
CmpDate: 2022-04-04

Awotoye W, Comnick C, Pendleton C, et al (2022)

Genome-wide Gene-by-Sex Interaction Studies Identify Novel Nonsyndromic Orofacial Clefts Risk Locus.

Journal of dental research, 101(4):465-472.

Risk loci identified through genome-wide association studies have explained about 25% of the phenotypic variations in nonsyndromic orofacial clefts (nsOFCs) on the liability scale. Despite the notable sex differences in the incidences of the different cleft types, investigation of loci for sex-specific effects has been understudied. To explore the sex-specific effects in genetic etiology of nsOFCs, we conducted a genome-wide gene × sex (GxSex) interaction study in a sub-Saharan African orofacial cleft cohort. The sample included 1,019 nonsyndromic orofacial cleft cases (814 cleft lip with or without cleft palate and 205 cleft palate only) and 2,159 controls recruited from 3 sites (Ethiopia, Ghana, and Nigeria). An additive logistic model was used to examine the joint effects of the genotype and GxSex interaction. Furthermore, we examined loci with suggestive significance (P < 1E-5) in the additive model for the effect of the GxSex interaction only. We identified a novel risk locus on chromosome 8p22 with genome-wide significant joint and GxSex interaction effects (rs2720555, p2df = 1.16E-08, pGxSex = 1.49E-09, odds ratio [OR] = 0.44, 95% CI = 0.34 to 0.57). For males, the risk of cleft lip with or without cleft palate at this locus decreases with additional copies of the minor allele (p < 0.0001, OR = 0.60, 95% CI = 0.48 to 0.74), but the effect is reversed for females (p = 0.0004, OR = 1.36, 95% CI = 1.15 to 1.60). We replicated the female-specific effect of this locus in an independent cohort (p = 0.037, OR = 1.30, 95% CI = 1.02 to 1.65), but no significant effect was found for the males (p = 0.29, OR = 0.86, 95% CI = 0.65 to 1.14). This locus is in topologically associating domain with craniofacially expressed and enriched genes during embryonic development. Rare coding mutations of some of these genes were identified in nsOFC cohorts through whole exome sequencing analysis. Our study is additional proof that genome-wide GxSex interaction analysis provides an opportunity for novel findings of loci and genes that contribute to the risk of nsOFCs.

RevDate: 2021-10-23

Marti-Marimon M, Vialaneix N, Lahbib-Mansais Y, et al (2021)

Major Reorganization of Chromosome Conformation During Muscle Development in Pig.

Frontiers in genetics, 12:748239.

The spatial organization of the genome in the nucleus plays a crucial role in eukaryotic cell functions, yet little is known about chromatin structure variations during late fetal development in mammals. We performed in situ high-throughput chromosome conformation capture (Hi-C) sequencing of DNA from muscle samples of pig fetuses at two late stages of gestation. Comparative analysis of the resulting Hi-C interaction matrices between both groups showed widespread differences of different types. First, we discovered a complex landscape of stable and group-specific Topologically Associating Domains (TADs). Investigating the nuclear partition of the chromatin into transcriptionally active and inactive compartments, we observed a genome-wide fragmentation of these compartments between 90 and 110 days of gestation. Also, we identified and characterized the distribution of differential cis- and trans-pairwise interactions. In particular, trans-interactions at chromosome extremities revealed a mechanism of telomere clustering further confirmed by 3D Fluorescence in situ Hybridization (FISH). Altogether, we report major variations of the three-dimensional genome conformation during muscle development in pig, involving several levels of chromatin remodeling and structural regulation.

RevDate: 2021-10-09

Servetti M, Pisciotta L, Tassano E, et al (2021)

Neurodevelopmental Disorders in Patients With Complex Phenotypes and Potential Complex Genetic Basis Involving Non-Coding Genes, and Double CNVs.

Frontiers in genetics, 12:732002.

Neurodevelopmental disorders (NDDs) are a heterogeneous class of brain diseases, with a complex genetic basis estimated to account for up to 50% of cases. Nevertheless, genetic diagnostic yield is about 20%. Array-comparative genomic hybridization (array-CGH) is an established first-level diagnostic test able to detect pathogenic copy number variants (CNVs), however, most identified variants remain of uncertain significance (VUS). Failure of interpretation of VUSs may depend on various factors, including complexity of clinical phenotypes and inconsistency of genotype-phenotype correlations. Indeed, although most NDD-associated CNVs are de novo, transmission from unaffected parents to affected children of CNVs with high risk for NDDs has been observed. Moreover, variability of genetic components overlapped by CNVs, such as long non-coding genes, genomic regions with long-range effects, and additive effects of multiple CNVs can make CNV interpretation challenging. We report on 12 patients with complex phenotypes possibly explained by complex genetic mechanisms, including involvement of antisense genes and boundaries of topologically associating domains. Eight among the 12 patients carried two CNVs, either de novo or inherited, respectively, by each of their healthy parents, that could additively contribute to the patients' phenotype. CNVs overlapped either known NDD-associated or novel candidate genes (PTPRD, BUD13, GLRA3, MIR4465, ABHD4, and WSCD2). Bioinformatic enrichment analyses showed that genes overlapped by the co-occurring CNVs have synergistic roles in biological processes fundamental in neurodevelopment. Double CNVs could concur in producing deleterious effects, according to a two-hit model, thus explaining the patients' phenotypes and the incomplete penetrance, and variable expressivity, associated with the single variants. Overall, our findings could contribute to the knowledge on clinical and genetic diagnosis of complex forms of NDD.

RevDate: 2021-10-15
CmpDate: 2021-10-15

Esquivel-López A, Arzate-Mejía R, Pérez-Molina R, et al (2021)

In-Nucleus Hi-C in Drosophila Cells.

Journal of visualized experiments : JoVE.

The genome is organized into topologically associating domains (TADs) delimited by boundaries that isolate interactions between domains. In Drosophila, the mechanisms underlying TAD formation and boundaries are still under investigation. The application of the in-nucleus Hi-C method described here helped to dissect the function of architectural protein (AP)-binding sites at TAD boundaries isolating the Notch gene. Genetic modification of domain boundaries that cause loss of APs results in TAD fusion, transcriptional defects, and long-range topological alterations. These results provided evidence demonstrating the contribution of genetic elements to domain boundary formation and gene expression control in Drosophila. Here, the in-nucleus Hi-C method has been described in detail, which provides important checkpoints to assess the quality of the experiment along with the protocol. Also shown are the required numbers of sequencing reads and valid Hi-C pairs to analyze genomic interactions at different genomic scales. CRISPR/Cas9-mediated genetic editing of regulatory elements and high-resolution profiling of genomic interactions using this in-nucleus Hi-C protocol could be a powerful combination for the investigation of the structural function of genetic elements.

RevDate: 2022-01-13
CmpDate: 2022-01-13

Luo H, Li X, Tian GG, et al (2021)

Offspring production of ovarian organoids derived from spermatogonial stem cells by defined factors with chromatin reorganization.

Journal of advanced research, 33:81-98.

Introduction: Fate determination of germline stem cells remains poorly understood at the chromatin structure level.

Objectives: Our research hopes to develop successful offspring production of ovarian organoids derived from spermatogonial stem cells (SSCs) by defined factors.

Methods: The offspring production from oocytes transdifferentiated from mouse SSCs with tracking of transplanted SSCs in vivo, single cell whole exome sequencing, and in 3D cell culture reconstitution of the process of oogenesis derived from SSCs. The defined factors were screened with ovarian organoids. We uncovered extensive chromatin reorganization during SSC conversion into induced germline stem cells (iGSCs) using high throughput chromosome conformation.

Results: We demonstrate successful production of offspring from oocytes transdifferentiated from mouse spermatogonial stem cells (SSCs). Furthermore, we demonstrate direct induction of germline stem cells (iGSCs) differentiated into functional oocytes by transduction of H19, Stella, and Zfp57 and inactivation of Plzf in SSCs after screening with ovarian organoids. We uncovered extensive chromatin reorganization during SSC conversion into iGSCs, which was highly similar to female germline stem cells. We observed that although topologically associating domains were stable during SSC conversion, chromatin interactions changed in a striking manner, altering 35% of inactive and active chromosomal compartments throughout the genome.

Conclusion: We demonstrate successful offspring production of ovarian organoids derived from SSCs by defined factors with chromatin reorganization. These findings have important implications in various areas including mammalian gametogenesis, genetic and epigenetic reprogramming, biotechnology, and medicine.

RevDate: 2021-10-04

Grabowicz IE, Wilczyński B, Kamińska B, et al (2021)

Author Correction: The role of epigenetic modifications, long-range contacts, enhancers and topologically associating domains in the regulation of glioma grade-specific genes.

Scientific reports, 11(1):19628 pii:10.1038/s41598-021-99319-4.

RevDate: 2022-02-09
CmpDate: 2022-02-09

He L, Ding Y, Zhao Y, et al (2021)

CRISPR/Cas9/AAV9-mediated in vivo editing identifies MYC regulation of 3D genome in skeletal muscle stem cell.

Stem cell reports, 16(10):2442-2458.

Skeletal muscle satellite cells (SCs) are stem cells responsible for muscle development and regeneration. Although CRISPR/Cas9 has been widely used, its application in endogenous SCs remains elusive. Here, we generate mice expressing Cas9 in SCs and achieve robust editing in juvenile SCs at the postnatal stage through AAV9-mediated short guide RNA (sgRNA) delivery. Additionally, we reveal that quiescent SCs are resistant to CRISPR/Cas9-mediated editing. As a proof of concept, we demonstrate efficient editing of master transcription factor (TF) Myod1 locus using the CRISPR/Cas9/AAV9-sgRNA system in juvenile SCs. Application on two key TFs, MYC and BCL6, unveils distinct functions in SC activation and muscle regeneration. Particularly, we reveal that MYC orchestrates SC activation through regulating 3D genome architecture. Its depletion results in strengthening of the topologically associating domain boundaries thus may affect gene expression. Altogether, our study establishes a platform for editing endogenous SCs that can be harnessed to elucidate the functionality of key regulators governing SC activities.

RevDate: 2021-11-08
CmpDate: 2021-10-19

Franke M, De la Calle-Mustienes E, Neto A, et al (2021)

CTCF knockout in zebrafish induces alterations in regulatory landscapes and developmental gene expression.

Nature communications, 12(1):5415.

Coordinated chromatin interactions between enhancers and promoters are critical for gene regulation. The architectural protein CTCF mediates chromatin looping and is enriched at the boundaries of topologically associating domains (TADs), which are sub-megabase chromatin structures. In vitro CTCF depletion leads to a loss of TADs but has only limited effects over gene expression, challenging the concept that CTCF-mediated chromatin structures are a fundamental requirement for gene regulation. However, how CTCF and a perturbed chromatin structure impacts gene expression during development remains poorly understood. Here we link the loss of CTCF and gene regulation during patterning and organogenesis in a ctcf knockout zebrafish model. CTCF absence leads to loss of chromatin structure and affects the expression of thousands of genes, including many developmental regulators. Our results demonstrate the essential role of CTCF in providing the structural context for enhancer-promoter interactions, thus regulating developmental genes.

RevDate: 2021-12-17
CmpDate: 2021-12-17

MacPhillamy C, Pitchford WS, Alinejad-Rokny H, et al (2021)

Opportunity to improve livestock traits using 3D genomics.

Animal genetics, 52(6):785-798.

The advent of high-throughput chromosome conformation capture and sequencing (Hi-C) has enabled researchers to probe the 3D architecture of the mammalian genome in a genome-wide manner. Simultaneously, advances in epigenomic assays, such as chromatin immunoprecipitation and sequencing (ChIP-seq) and DNase-seq, have enabled researchers to study cis-regulatory interactions and chromatin accessibility across the same genome-wide scale. The use of these data has revealed many unique insights into gene regulation and disease pathomechanisms in several model organisms. With the advent of these high-throughput sequencing technologies, there has been an ever-increasing number of datasets available for study; however, this is often limited to model organisms. Livestock species play critical roles in the economies of developing and developed nations alike. Despite this, they are greatly underrepresented in the 3D genomics space; Hi-C and related technologies have the potential to revolutionise livestock breeding by enabling a more comprehensive understanding of how production traits are controlled. The growth in human and model organism Hi-C data has seen a surge in the availability of computational tools for use in 3D genomics, with some tools using machine learning techniques to predict features and improve dataset quality. In this review, we provide an overview of the 3D genome and discuss the status of 3D genomics in livestock before delving into advancing the field by drawing inspiration from research in human and mouse. We end by offering future directions for livestock research in the field of 3D genomics.

RevDate: 2022-02-18
CmpDate: 2021-09-30

Melo US, Piard J, Fischer-Zirnsak B, et al (2021)

Complete lung agenesis caused by complex genomic rearrangements with neo-TAD formation at the SHH locus.

Human genetics, 140(10):1459-1469.

During human organogenesis, lung development is a timely and tightly regulated developmental process under the control of a large number of signaling molecules. Understanding how genetic variants can disturb normal lung development causing different lung malformations is a major goal for dissecting molecular mechanisms during embryogenesis. Here, through exome sequencing (ES), array CGH, genome sequencing (GS) and Hi-C, we aimed at elucidating the molecular basis of bilateral isolated lung agenesis in three fetuses born to a non-consanguineous family. We detected a complex genomic rearrangement containing duplicated, triplicated and deleted fragments involving the SHH locus in fetuses presenting complete agenesis of both lungs and near-complete agenesis of the trachea, diagnosed by ultrasound screening and confirmed at autopsy following termination. The rearrangement did not include SHH itself, but several regulatory elements for lung development, such as MACS1, a major SHH lung enhancer, and the neighboring genes MNX1 and NOM1. The rearrangement incorporated parts of two topologically associating domains (TADs) including their boundaries. Hi-C of cells from one of the affected fetuses showed the formation of two novel TADs each containing SHH enhancers and the MNX1 and NOM1 genes. Hi-C together with GS indicate that the new 3D conformation is likely causative for this condition by an inappropriate activation of MNX1 included in the neo-TADs by MACS1 enhancer, further highlighting the importance of the 3D chromatin conformation in human disease.

RevDate: 2022-01-29
CmpDate: 2022-01-18

Ge X, Frank-Bertoncelj M, Klein K, et al (2021)

Functional genomics atlas of synovial fibroblasts defining rheumatoid arthritis heritability.

Genome biology, 22(1):247.

BACKGROUND: Genome-wide association studies have reported more than 100 risk loci for rheumatoid arthritis (RA). These loci are shown to be enriched in immune cell-specific enhancers, but the analysis so far has excluded stromal cells, such as synovial fibroblasts (FLS), despite their crucial involvement in the pathogenesis of RA. Here we integrate DNA architecture, 3D chromatin interactions, DNA accessibility, and gene expression in FLS, B cells, and T cells with genetic fine mapping of RA loci.

RESULTS: We identify putative causal variants, enhancers, genes, and cell types for 30-60% of RA loci and demonstrate that FLS account for up to 24% of RA heritability. TNF stimulation of FLS alters the organization of topologically associating domains, chromatin state, and the expression of putative causal genes such as TNFAIP3 and IFNAR1. Several putative causal genes constitute RA-relevant functional networks in FLS with roles in cellular proliferation and activation. Finally, we demonstrate that risk variants can have joint-specific effects on target gene expression in RA FLS, which may contribute to the development of the characteristic pattern of joint involvement in RA.

CONCLUSION: Overall, our research provides the first direct evidence for a causal role of FLS in the genetic susceptibility for RA accounting for up to a quarter of RA heritability.

RevDate: 2022-03-23
CmpDate: 2021-11-29

Groves IJ, Drane ELA, Michalski M, et al (2021)

Short- and long-range cis interactions between integrated HPV genomes and cellular chromatin dysregulate host gene expression in early cervical carcinogenesis.

PLoS pathogens, 17(8):e1009875.

Development of cervical cancer is directly associated with integration of human papillomavirus (HPV) genomes into host chromosomes and subsequent modulation of HPV oncogene expression, which correlates with multi-layered epigenetic changes at the integrated HPV genomes. However, the process of integration itself and dysregulation of host gene expression at sites of integration in our model of HPV16 integrant clone natural selection has remained enigmatic. We now show, using a state-of-the-art 'HPV integrated site capture' (HISC) technique, that integration likely occurs through microhomology-mediated repair (MHMR) mechanisms via either a direct process, resulting in host sequence deletion (in our case, partially homozygously) or via a 'looping' mechanism by which flanking host regions become amplified. Furthermore, using our 'HPV16-specific Region Capture Hi-C' technique, we have determined that chromatin interactions between the integrated virus genome and host chromosomes, both at short- (<500 kbp) and long-range (>500 kbp), appear to drive local host gene dysregulation through the disruption of host:host interactions within (but not exceeding) host structures known as topologically associating domains (TADs). This mechanism of HPV-induced host gene expression modulation indicates that integration of virus genomes near to or within a 'cancer-causing gene' is not essential to influence their expression and that these modifications to genome interactions could have a major role in selection of HPV integrants at the early stage of cervical neoplastic progression.

RevDate: 2022-03-10
CmpDate: 2022-03-10

Chiliński M, Sengupta K, D Plewczynski (2022)

From DNA human sequence to the chromatin higher order organisation and its biological meaning: Using biomolecular interaction networks to understand the influence of structural variation on spatial genome organisation and its functional effect.

Seminars in cell & developmental biology, 121:171-185.

The three-dimensional structure of the human genome has been proven to have a significant functional impact on gene expression. The high-order spatial chromatin is organised first by looping mediated by multiple protein factors, and then it is further formed into larger structures of topologically associated domains (TADs) or chromatin contact domains (CCDs), followed by A/B compartments and finally the chromosomal territories (CTs). The genetic variation observed in human population influences the multi-scale structures, posing a question regarding the functional impact of structural variants reflected by the variability of the genes expression patterns. The current methods of evaluating the functional effect include eQTLs analysis which uses statistical testing of influence of variants on spatially close genes. Rarely, non-coding DNA sequence changes are evaluated by their impact on the biomolecular interaction network (BIN) reflecting the cellular interactome that can be analysed by the classical graph-theoretic algorithms. Therefore, in the second part of the review, we introduce the concept of BIN, i.e. a meta-network model of the complete molecular interactome developed by integrating various biological networks. The BIN meta-network model includes DNA-protein binding by the plethora of protein factors as well as chromatin interactions, therefore allowing connection of genomics with the downstream biomolecular processes present in a cell. As an illustration, we scrutinise the chromatin interactions mediated by the CTCF protein detected in a ChIA-PET experiment in the human lymphoblastoid cell line GM12878. In the corresponding BIN meta-network the DNA spatial proximity is represented as a graph model, combined with the Proteins-Interaction Network (PIN) of human proteome using the Gene Association Network (GAN). Furthermore, we enriched the BIN with the signalling and metabolic pathways and Gene Ontology (GO) terms to assert its functional context. Finally, we mapped the Single Nucleotide Polymorphisms (SNPs) from the GWAS studies and identified the chromatin mutational hot-spots associated with a significant enrichment of SNPs related to autoimmune diseases. Afterwards, we mapped Structural Variants (SVs) from healthy individuals of 1000 Genomes Project and identified an interesting example of the missing protein complex associated with protein Q6GYQ0 due to a deletion on chromosome 14. Such an analysis using the meta-network BIN model is therefore helpful in evaluating the influence of genetic variation on spatial organisation of the genome and its functional effect in a cell.

RevDate: 2022-04-21
CmpDate: 2022-04-21

Arrastia MV, Jachowicz JW, Ollikainen N, et al (2022)

Single-cell measurement of higher-order 3D genome organization with scSPRITE.

Nature biotechnology, 40(1):64-73.

Although three-dimensional (3D) genome organization is central to many aspects of nuclear function, it has been difficult to measure at the single-cell level. To address this, we developed 'single-cell split-pool recognition of interactions by tag extension' (scSPRITE). scSPRITE uses split-and-pool barcoding to tag DNA fragments in the same nucleus and their 3D spatial arrangement. Because scSPRITE measures multiway DNA contacts, it generates higher-resolution maps within an individual cell than can be achieved by proximity ligation. We applied scSPRITE to thousands of mouse embryonic stem cells and detected known genome structures, including chromosome territories, active and inactive compartments, and topologically associating domains (TADs) as well as long-range inter-chromosomal structures organized around various nuclear bodies. We observe that these structures exhibit different levels of heterogeneity across the population, with TADs representing dynamic units of genome organization across cells. We expect that scSPRITE will be a critical tool for studying genome structure within heterogeneous populations.


ESP Quick Facts

ESP Origins

In the early 1990's, Robert Robbins was a faculty member at Johns Hopkins, where he directed the informatics core of GDB — the human gene-mapping database of the international human genome project. To share papers with colleagues around the world, he set up a small paper-sharing section on his personal web page. This small project evolved into The Electronic Scholarly Publishing Project.

ESP Support

In 1995, Robbins became the VP/IT of the Fred Hutchinson Cancer Research Center in Seattle, WA. Soon after arriving in Seattle, Robbins secured funding, through the ELSI component of the US Human Genome Project, to create the original ESP.ORG web site, with the formal goal of providing free, world-wide access to the literature of classical genetics.

ESP Rationale

Although the methods of molecular biology can seem almost magical to the uninitiated, the original techniques of classical genetics are readily appreciated by one and all: cross individuals that differ in some inherited trait, collect all of the progeny, score their attributes, and propose mechanisms to explain the patterns of inheritance observed.

ESP Goal

In reading the early works of classical genetics, one is drawn, almost inexorably, into ever more complex models, until molecular explanations begin to seem both necessary and natural. At that point, the tools for understanding genome research are at hand. Assisting readers reach this point was the original goal of The Electronic Scholarly Publishing Project.

ESP Usage

Usage of the site grew rapidly and has remained high. Faculty began to use the site for their assigned readings. Other on-line publishers, ranging from The New York Times to Nature referenced ESP materials in their own publications. Nobel laureates (e.g., Joshua Lederberg) regularly used the site and even wrote to suggest changes and improvements.

ESP Content

When the site began, no journals were making their early content available in digital format. As a result, ESP was obliged to digitize classic literature before it could be made available. For many important papers — such as Mendel's original paper or the first genetic map — ESP had to produce entirely new typeset versions of the works, if they were to be available in a high-quality format.

ESP Help

Early support from the DOE component of the Human Genome Project was critically important for getting the ESP project on a firm foundation. Since that funding ended (nearly 20 years ago), the project has been operated as a purely volunteer effort. Anyone wishing to assist in these efforts should send an email to Robbins.

ESP Plans

With the development of methods for adding typeset side notes to PDF files, the ESP project now plans to add annotated versions of some classical papers to its holdings. We also plan to add new reference and pedagogical material. We have already started providing regularly updated, comprehensive bibliographies to the ESP.ORG site.

Electronic Scholarly Publishing
961 Red Tail Lane
Bellingham, WA 98226

E-mail: RJR8222 @

Papers in Classical Genetics

The ESP began as an effort to share a handful of key papers from the early days of classical genetics. Now the collection has grown to include hundreds of papers, in full-text format.

Digital Books

Along with papers on classical genetics, ESP offers a collection of full-text digital books, including many works by Darwin (and even a collection of poetry — Chicago Poems by Carl Sandburg).


ESP now offers a much improved and expanded collection of timelines, designed to give the user choice over subject matter and dates.


Biographical information about many key scientists.

Selected Bibliographies

Bibliographies on several topics of potential interest to the ESP community are now being automatically maintained and generated on the ESP site.

ESP Picks from Around the Web (updated 07 JUL 2018 )