The sequence of a genome is insufficient to understand all genomic processes carried out in the cell nucleus. To achieve this, the knowledge of its three-dimensional architecture is necessary. ...Advances in genomic technologies and the development of new analytical methods, such as Chromosome Conformation Capture (3C) and its derivatives, provide unprecedented insights in the spatial organization of genomes. Here we present TADbit, a computational framework to analyze and model the chromatin fiber in three dimensions. Our package takes as input the sequencing reads of 3C-based experiments and performs the following main tasks: (i) pre-process the reads, (ii) map the reads to a reference genome, (iii) filter and normalize the interaction data, (iv) analyze the resulting interaction matrices, (v) build 3D models of selected genomic domains, and (vi) analyze the resulting models to characterize their structural properties. To illustrate the use of TADbit, we automatically modeled 50 genomic domains from the fly genome revealing differential structural features of the previously defined chromatin colors, establishing a link between the conformation of the genome and the local chromatin composition. TADbit provides three-dimensional models built from 3C-based experiments, which are ready for visualization and for characterizing their relation to gene expression and epigenetic states. TADbit is an open-source Python library available for download from https://github.com/3DGenomes/tadbit.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Chromosomes are large polymer molecules composed of nucleotides. In some species, such as humans, this polymer can sum up to meters long and still be properly folded within the nuclear space of few ...microns in size. The exact mechanisms of how the meters long DNA is folded into the nucleus, as well as how the regulatory machinery can access it, is to a large extend still a mystery. However, and thanks to newly developed molecular, genomic and computational approaches based on the Chromosome Conformation Capture (3C) technology, we are now obtaining insight on how genomes are spatially organized. Here we review a new family of computational approaches that aim at using 3C-based data to obtain spatial restraints for modeling genomes and genomic domains.
The three-dimensional (3D) architecture of a genome determines the spatial localization of regulatory elements and the genes they regulate. Thus, elucidating the 3D structure of a genome may result ...in significant insights about how genes are regulated. The current state-of-the art in experimental methods, including light microscopy and cell/molecular biology, are now able to provide detailed information on the position of genes and their interacting partners. However, such methods by themselves are not able to determine the high-resolution 3D structure of genomes or genomic domains. Here we describe a computational module of the Integrative Modeling Platform (IMP, http://www.integrativemodeling.org) that uses chromosome conformation capture data to determine the 3D architecture of genomic domains and entire genomes at unprecedented resolutions. This approach, through the visualization of looping interactions between distal regulatory elements, allows characterizing global chromatin features and their relation to gene expression. We illustrate our work by outlining the determination of the 3D architecture of the α-globin domain in the human genome.
DNA-binding proteins are central regulators of chromosome organization; however, in genome-reduced bacteria their diversity is largely diminished. Whether the chromosomes of such bacteria adopt ...defined three-dimensional structures remains unexplored. Here we combine Hi-C and super-resolution microscopy to determine the structure of the Mycoplasma pneumoniae chromosome at a 10 kb resolution. We find a defined structure, with a global symmetry between two arms that connect opposite poles, one bearing the chromosomal Ori and the other the midpoint. Analysis of local structures at a 3 kb resolution indicates that the chromosome is organized into domains ranging from 15 to 33 kb. We provide evidence that genes within the same domain tend to be co-regulated, suggesting that chromosome organization influences transcriptional regulation, and that supercoiling regulates local organization. This study extends the current understanding of bacterial genome organization and demonstrates that a defined chromosomal structure is a universal feature of living systems.
We have determined the three-dimensional (3D) architecture of the
Caulobacter crescentus genome by combining genome-wide chromatin interaction detection, live-cell imaging, and computational ...modeling. Using chromosome conformation capture carbon copy (5C), we derive ∼13 kb resolution 3D models of the
Caulobacter genome. The resulting models illustrate that the genome is ellipsoidal with periodically arranged arms. The
parS sites, a pair of short contiguous sequence elements known to be involved in chromosome segregation, are positioned at one pole, where they anchor the chromosome to the cell and contribute to the formation of a compact chromatin conformation. Repositioning these elements resulted in rotations of the chromosome that changed the subcellular positions of most genes. Such rotations did not lead to large-scale changes in gene expression, indicating that genome folding does not strongly affect gene regulation. Collectively, our data suggest that genome folding is globally dictated by the
parS sites and chromosome segregation.
► Chromatin interaction mapping and modeling elucidate
Caulobacter genome structure ► The genome is ellipsoidal with periodically arranged arms ► The
parS region shapes whole genome structure and affects chromatin compaction ► The
parS region is the only genomic region stably attached to the cell envelope
The human genome is segmented into topologically associating domains (TADs), but the role of this conserved organization during transient changes in gene expression is not known. Here we describe the ...distribution of progestin-induced chromatin modifications and changes in transcriptional activity over TADs in T47D breast cancer cells. Using ChIP-seq (chromatin immunoprecipitation combined with high-throughput sequencing), Hi-C (chromosome capture followed by high-throughput sequencing), and three-dimensional (3D) modeling techniques, we found that the borders of the ∼ 2000 TADs in these cells are largely maintained after hormone treatment and that up to 20% of the TADs could be considered as discrete regulatory units where the majority of the genes are either transcriptionally activated or repressed in a coordinated fashion. The epigenetic signatures of the TADs are homogeneously modified by hormones in correlation with the transcriptional changes. Hormone-induced changes in gene activity and chromatin remodeling are accompanied by differential structural changes for activated and repressed TADs, as reflected by specific and opposite changes in the strength of intra-TAD interactions within responsive TADs. Indeed, 3D modeling of the Hi-C data suggested that the structure of TADs was modified upon treatment. The differential responses of TADs to progestins and estrogens suggest that TADs could function as "regulons" to enable spatially proximal genes to be coordinately transcribed in response to hormones.
Restraint-based modeling of genomes has been recently explored with the advent of Chromosome Conformation Capture (3C-based) experiments. We previously developed a reconstruction method to resolve ...the 3D architecture of both prokaryotic and eukaryotic genomes using 3C-based data. These models were congruent with fluorescent imaging validation. However, the limits of such methods have not systematically been assessed. Here we propose the first evaluation of a mean-field restraint-based reconstruction of genomes by considering diverse chromosome architectures and different levels of data noise and structural variability. The results show that: first, current scoring functions for 3D reconstruction correlate with the accuracy of the models; second, reconstructed models are robust to noise but sensitive to structural variability; third, the local structure organization of genomes, such as Topologically Associating Domains, results in more accurate models; fourth, to a certain extent, the models capture the intrinsic structural variability in the input matrices and fifth, the accuracy of the models can be a priori predicted by analyzing the properties of the interaction matrices. In summary, our work provides a systematic analysis of the limitations of a mean-field restrain-based method, which could be taken into consideration in further development of methods as well as their applications.
Recent technological advances in the field of chromosome conformation capture are facilitating tremendous progress in the ability to map the three-dimensional (3D) organization of chromosomes at a ...resolution of several Kb and at the scale of complete genomes. Here we review progress in analyzing chromosome organization in human cells by building 3D models of chromatin based on comprehensive chromatin interaction datasets. We describe recent experiments that suggest that long-range interactions between active functional elements are sufficient to drive folding of local chromatin domains into compact globular states. We propose that chromatin globules are commonly formed along chromosomes, in a cell type specific pattern, as a result of frequent long-range interactions among active genes and nearby regulatory elements. Further, we speculate that increasingly longer range interactions can drive aggregation of groups of globular domains. This process would yield a compartmentalized chromosome conformation, consistent with recent observations obtained with genome-wide chromatin interaction mapping.
NCT04580069.
Total knee arthroplasty is associated with an elevated inflammatory response both at a local and systemic level. The main objective of this study is to demonstrate the effectiveness of ...lymphatic drainage and connective tissue techniques in modulating systemic inflammation. Another objective is to evaluate the existence, at baseline, of a correlation between the inflammation indices and the level of adherence to the Mediterranean diet.
34 patients were recruited, and divided into three groups. The control group followed the normal rehabilitation protocol. The other two groups were subjected, in addition to the standard treatment, to manual lymphatic drainage treatment or connective tissue techniques. The outcomes were recorded in three stages: upon entering the hospital, 1 week after entry and at follow-up 21 days after surgery.
The results of the study showed that both methods, compared with the standard treatment only, positively influenced the final outcomes. In regard to the systemic inflammation, lymphatic drainage and connective techniques showed equal efficacy and similar timing in modulating ESR, while they differ in how they affect CRP. With regard to the local inflammation, the effectiveness of both methods was confirmed with some differences in the location. Finally, analysis of the correlation between inflammatory T0 indices and adherence to the Mediterranean diet showed that patients with higher adhesion index have on average lower PCR, EDO and EDU values.
The post-surgical inflammatory pattern can be positively modified by the rehabilitation methods analyzed, albeit with different methodologies and timing.
The influence of the diet on inflammatory parameters, although less evident, seems to show encouraging results worth of further studies.
PA-824 is a promising drug candidate for the treatment of tuberculosis (TB). It is in phase II clinical trials as part of the first newly designed regimen containing multiple novel antituberculosis ...drugs (PA-824 in combination with moxifloxacin and pyrazinamide). However, given that the genes involved in resistance against PA-824 are not fully conserved in the Mycobacterium tuberculosis complex (MTBC), this regimen might not be equally effective against different MTBC genotypes. To investigate this question, we sequenced two PA-824 resistance genes (fgd1 Rv0407 and ddn Rv3547) in 65 MTBC strains representing major phylogenetic lineages. The MICs of representative strains were determined using the modified proportion method in the Bactec MGIT 960 system. Our analysis revealed single-nucleotide polymorphisms in both genes that were specific either for several genotypes or for individual strains, yet none of these mutations significantly affected the PA-824 MICs (≤0.25 μg/ml). These results were supported by in silico modeling of the mutations identified in Fgd1. In contrast, "Mycobacterium canettii" strains displayed a higher MIC of 8 μg/ml. In conclusion, we found a large genetic diversity in PA-824 resistance genes that did not lead to elevated PA-824 MICs. In contrast, M. canettii strains had MICs that were above the plasma concentrations of PA-824 documented so far in clinical trials. As M. canettii is also intrinsically resistant against pyrazinamide, new regimens containing PA-824 and pyrazinamide might not be effective in treating M. canettii infections. This finding has implications for the design of multiple ongoing clinical trials.
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