Microrheology for Hi-C Data Reveals the Spectrum of the Dynamic 3D Genome Organization

The one-dimensional information of genomic DNA is hierarchically packed inside the eukaryotic cell nucleus and organized in a three-dimensional (3D) space. Genome-wide chromosome conformation capture (Hi-C) methods have uncovered the 3D genome organization and revealed multiscale chromatin domains of compartments and topologically associating domains (TADs). Moreover, single-nucleosome live-cell imaging experiments have revealed the dynamic organization of chromatin domains caused by stochastic thermal fluctuations. However, the mechanism underlying the dynamic regulation of such hierarchical and structural chromatin units within the microscale thermal medium remains unclear. Microrheology is a way to measure dynamic viscoelastic properties coupling between thermal microenvironment and mechanical response. Here, we propose a new, to our knowledge, microrheology for Hi-C data to analyze the dynamic compliance property as a measure of rigidness and flexibility of genomic regions along with the time evolution. Our method allows the conversion of an Hi-C matrix into the spectrum of the dynamic rheological property along the genomic coordinate of a single chromosome. To demonstrate the power of the technique, we analyzed Hi-C data during the neural differentiation of mouse embryonic stem cells. We found that TAD boundaries behave as more rigid nodes than the intra-TAD regions. The spectrum clearly shows the dynamic viscoelasticity of chromatin domain formation at different timescales. Furthermore, we characterized the appearance of synchronous and liquid-like intercompartment interactions in differentiated cells. Together, our microrheology data derived from Hi-C data provide physical insights into the dynamics of the 3D genome organization.     

Minimalistic 3D chromatin models: Sparse interactions in single cells drive the chromatin fold and form many-body units

Computational three-dimensional chromatin modeling has helped uncover principles of genome organization. Here, we discuss methods for modeling three-dimensional chromatin structures, with focus on a minimalistic polymer model which inverts population Hi-C into single-cell conformations. Utilizing only basic physical properties, this model reveals that a few specific Hi-C interactions can fold chromatin into conformations consistent with single-cell imaging, Dip-C, and FISH measurements. Aggregated single-cell chromatin conformations also reproduce Hi-C frequencies. This approach allows quantification of structural heterogeneity and discovery of many-body interaction units and has revealed additional insights, including (1) topologically associating domains as a byproduct of folding driven by specific interactions, (2) cell subpopulations with different structural scaffolds are developmental stage dependent, and (3) the functional landscape of many-body units within enhancer-rich regions. We also discuss these findings in relation to the genome structure–function relationship.     

Uncovering the statistical physics of 3D chromosomal organization using data-driven modeling

In recent years, much effort has been devoted to understanding the three-dimensional (3D) organization of the genome and how genomic structure mediates nuclear function. The development of experimental techniques that combine DNA proximity ligation with high-throughput sequencing, such as Hi-C, have substantially improved our knowledge about chromatin organization. Numerous experimental advancements, not only utilizing DNA proximity ligation but also high-resolution genome imaging (DNA tracing), have required theoretical modeling to determine the structural ensembles consistent with such data. These 3D polymer models of the genome provide an understanding of the physical mechanisms governing genome architecture. Here, we present an overview of the recent advances in modeling the ensemble of 3D chromosomal structures by employing the maximum entropy approach combined with polymer physics. Particularly, we discuss the minimal chromatin model (MiChroM) along with the “maximum entropy genomic annotations from biomarkers associated with structural ensembles” (MEGABASE) model, which have been remarkably successful in the accurate modeling of chromosomes consistent with both Hi-C and DNA-tracing data.     

Developing bioimaging and quantitative methods to study 3D genome

The recent advances in chromosome configuration capture (3C)-based series molecular methods and optical super-resolution (SR) techniques offer powerful tools to investigate three dimensional (3D) genomic structure in prokaryotic and eukaryotic cell nucleus. In this review, we focus on the progress during the last decade in this exciting field. Here we at first introduce briefly genome organization at chromosome, domain and sub-domain level, respectively; then we provide a short introduction to various super-resolution microscopy techniques which can be employed to detect genome 3D structure. We also reviewed the progress of quantitative and visualization tools to evaluate and visualize chromatin interactions in 3D genome derived from Hi-C data. We end up with the discussion that imaging methods and 3C-based molecular methods are not mutually exclusive - - - - actually they are complemental to each other and can be combined together to study 3D genome organization.     

Genomic views of chromatin

With the availability of complete genome sequences for a number of organisms, a major challenge has become to understand how chromatin and its epigenetic modifications regulate genome function. High-throughput microarray and sequencing technologies are being combined with biochemical and immunological enrichment methods to obtain genome-scale views of chromatin in a variety of organisms. The data pinpoint novel, genomic elements and expansive chromatin domains, and offer insight into the functions of histone modifications. In parallel, state-of-the-art imaging techniques are being used to investigate higher-order chromatin organization, and are beginning to bridge our understanding of chromatin biology with that of chromosome structure.     

A Scalable Computational Approach for Simulating Complexes of Multiple Chromosomes

Significant efforts have been recently made to obtain the three-dimensional (3D) structure of the genome with the goal of understanding how structures may affect gene regulation and expression. Chromosome conformational capture techniques such as Hi-C, have been key in uncovering the quantitative information needed to determine chromatin organization. Complementing these experimental tools, co-polymers theoretical methods are necessary to determine the ensemble of three-dimensional structures associated to the experimental data provided by Hi-C maps. Going beyond just structural information, these theoretical advances also start to provide an understanding of the underlying mechanisms governing genome assembly and function. Recent theoretical work, however, has been focused on single chromosome structures, missing the fact that, in the full nucleus, interactions between chromosomes play a central role in their organization. To overcome this limitation, MiChroM (Minimal Chromatin Model) has been modified to become capable of performing these multi-chromosome simulations. It has been upgraded into a fast and scalable software version, which is able to perform chromosome simulations using GPUs via OpenMM Python API, called Open-MiChroM. To validate the efficiency of this new version, analyses for GM12878 individual autosomes were performed and compared to earlier studies. This validation was followed by multi-chain simulations including the four largest human chromosomes (C1-C4). These simulations demonstrated the full power of this new approach. Comparison to Hi-C data shows that these multiple chromosome interactions are essential for a more accurate agreement with experimental results. Without any changes to the original MiChroM potential, it is now possible to predict experimentally observed inter-chromosome contacts. This scalability of Open-MiChroM allow for more audacious investigations, looking at interactions of multiple chains as well as moving towards higher resolution chromosomes models.     

Banding Pattern of Polytene Chromosomes as a Representation of Universal Principles of Chromatin Organization into Topological Domains

Drosophila polytene chromosomes are widely used as a model of eukaryotic interphase chromosomes. The most noticeable feature of polytene chromosome is transverse banding associated with alternation of dense stripes (dark or black bands) and light diffuse areas that encompass alternating less compact gray bands and interbands visible with an electron microscope. In recent years, several approaches have been developed to predict location of morphological structures of polytene chromosomes based on the distribution of proteins on the molecular map of Drosophila genome. Comparison of these structures with the results of analysis of the three-dimensional chromatin organization by the Hi-C method indicates that the morphology of polytene chromosomes represents direct visualization of the interphase nucleus spatial organization into topological domains. Compact black bands correspond to the extended topological domains of inactive chromatin, while interbands are the barriers between the adjacent domains. Here, we discuss the prospects of using polytene chromosomes to study mechanisms of spatial organization of interphase chromosomes, as well as their dynamics and evolution.     

Large-scale reconstruction of 3D structures of human chromosomes from chromosomal contact data

Chromosomes are not positioned randomly within a nucleus, but instead, they adopt preferred spatial conformations to facilitate necessary long-range gene–gene interactions and regulations. Thus, obtaining the 3D shape of chromosomes of a genome is critical for understanding how the genome folds, functions and how its genes interact and are regulated. Here, we describe a method to reconstruct preferred 3D structures of individual chromosomes of the human genome from chromosomal contact data generated by the Hi-C chromosome conformation capturing technique. A novel parameterized objective function was designed for modeling chromosome structures, which was optimized by a gradient descent method to generate chromosomal structural models that could satisfy as many intra-chromosomal contacts as possible. We applied the objective function and the corresponding optimization method to two Hi-C chromosomal data sets of both a healthy and a cancerous human B-cell to construct 3D models of individual chromosomes at resolutions of 1 MB and 200 KB, respectively. The parameters used with the method were calibrated according to an independent fluorescence in situ hybridization experimental data. The structural models generated by our method could satisfy a high percentage of contacts (pairs of loci in interaction) and non-contacts (pairs of loci not in interaction) and were compatible with the known two-compartment organization of human chromatin structures. Furthermore, structural models generated at different resolutions and from randomly permuted data sets were consistent.     

MaxHiC: A robust background correction model to identify biologically relevant chromatin interactions in Hi-C and capture Hi-C experiments

Hi-C is a genome-wide chromosome conformation capture technology that detects interactions between pairs of genomic regions and exploits higher order chromatin structures. Conceptually Hi-C data counts interaction frequencies between every position in the genome and every other position. Biologically functional interactions are expected to occur more frequently than transient background and artefactual interactions. To identify biologically relevant interactions, several background models that take biases such as distance, GC content and mappability into account have been proposed. Here we introduce MaxHiC, a background correction tool that deals with these complex biases and robustly identifies statistically significant interactions in both Hi-C and capture Hi-C experiments. MaxHiC uses a negative binomial distribution model and a maximum likelihood technique to correct biases in both Hi-C and capture Hi-C libraries. We systematically benchmark MaxHiC against major Hi-C background correction tools including Hi-C significant interaction callers (SIC) and Hi-C loop callers using published Hi-C, capture Hi-C, and Micro-C datasets. Our results demonstrate that 1) Interacting regions identified by MaxHiC have significantly greater levels of overlap with known regulatory features (e.g. active chromatin histone marks, CTCF binding sites, DNase sensitivity) and also disease-associated genome-wide association SNPs than those identified by currently existing models, 2) the pairs of interacting regions are more likely to be linked by eQTL pairs and 3) more likely to link known regulatory features including known functional enhancer-promoter pairs validated by CRISPRi than any of the existing methods. We also demonstrate that interactions between different genomic region types have distinct distance distributions only revealed by MaxHiC. MaxHiC is publicly available as a python package for the analysis of Hi-C, capture Hi-C and Micro-C data.     

细胞终末分化过程中三维基因组结构与功能调控的分子机制

真核细胞中的染色质DNA高度折叠形成复杂的三维结构,其空间组织方式对精准调控基因的表达和细胞发挥正常功能都起着重要的作用。细胞终末分化成熟过程中形态及基因表达谱常发生显著改变,同时伴随着明显的基因组三维结构变化。本文在简单介绍三维基因组多层次组织结构(染色质领域、A/B区室、拓扑相关结构域和成环构象等)基础上,重点综述了细胞终末分化过程中三维基因组结构变化与功能调控方面的研究进展,并探讨了当前三维基因组研究在解析细胞分化成熟过程时存在的问题和前景。     

Spatial organization of interphase chromosomes and the role of chromatin fibril dynamics in the positioning of genome elements

Many studies are devoted to the analysis of interphase chromosome architecture due to the evidence of the functional-dependent spatial organization of the genome. These studies are based on classical cytological methods, as well as on biochemical approaches (3C, 4C, 5C, Hi-C), which allow one to detect long-range interactions between fragments of chromatin fibril, including the genome-wide interactions. In this review, we discuss the results of these projects, which allow us to explain the functional basis of nucleus multilevel compartmentalization and to identify the principles of high-level chromatin organization. Special attention is paid to the enhancer-promoter interactions, which are important for the regulation of gene expression. In this regard, we provide a new interpretation to the model of an active chromatin hub and to the alternative model of an active chromatin compartment, which was proposed during reconsideration of some steps of the 3C procedure.     

Comparison of the three-dimensional organization of sperm and fibroblast genomes using the Hi-C approach

BackgroundThe three-dimensional organization of the genome is tightly connected to its biological function. The Hi-C approach was recently introduced as a method that can be used to identify higher-order chromatin interactions genome-wide. The aim of this study was to determine genome-wide chromatin interaction frequencies using the Hi-C approach in mouse sperm cells and embryonic fibroblasts.ResultsThe obtained data demonstrate that the three-dimensional genome organizations of sperm and fibroblast cells show a high degree of similarity both with each other and with the previously described mouse embryonic stem cells. Both A- and B-compartments and topologically associated domains are present in spermatozoa and fibroblasts. Nevertheless, sperm cells and fibroblasts exhibit statistically significant differences between each other in the contact probabilities of defined loci. Tight packaging of the sperm genome results in an enrichment of long-range contacts compared with the fibroblasts. However, only 30% of the differences in the number of contacts are based on differences in the densities of their genome packages; the main source of the differences is the gain or loss of contacts that are specific for defined genome regions. We find that the dependence of the contact probability on genomic distance for sperm is close to the dependence predicted for the fractal globular folding of chromatin.ConclusionsOverall, we can conclude that the three-dimensional structure of the genome is passed through generations without being dramatically changed in sperm cells.

Electronic supplementary material

The online version of this article (doi:10.1186/s13059-015-0642-0) contains supplementary material, which is available to authorized users.     

Bridging the resolution gap in structural modeling of 3D genome organization

Over the last decade, and especially after the advent of fluorescent in situ hybridization imaging and chromosome conformation capture methods, the availability of experimental data on genome three-dimensional organization has dramatically increased. We now have access to unprecedented details of how genomes organize within the interphase nucleus. Development of new computational approaches to leverage this data has already resulted in the first three-dimensional structures of genomic domains and genomes. Such approaches expand our knowledge of the chromatin folding principles, which has been classically studied using polymer physics and molecular simulations. Our outlook describes computational approaches for integrating experimental data with polymer physics, thereby bridging the resolution gap for structural determination of genomes and genomic domains.     

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

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.Subject terms: Embryonic stem cells, Reprogramming, Stem-cell differentiation     

Chromatin architectural alterations due to null mutation of a major CG methylase in rice

Associations between 3D chromatin architectures and epigenetic modifications have been characterized in animals.However,any impact of DNA methylation on chromatin architecture in plants is understudied,which is confined to Arabidopsis thaliana.Because plant species differ in genome size,composition,and overall chromatin packing,it is unclear to what extent findings from A.thaliana hold in other species.Moreover,the incomplete chromatin architectural profiles and the low-resolution high-throughpu...