Dramatic chromosome motion is a characteristic of mid-prophase of meiosis that is observed across broadly divergent eukaryotic phyla. Although the specific mechanisms underlying chromosome motions ...vary among organisms studied to date, the outcome is similar in all cases: vigorous back-and-forth movement (as fast as ∼1 μm/sec for budding yeast), led by chromosome ends (or near-end regions), and directed by cytoskeletal components via direct association through the nuclear envelope. The exact role(s) of these movements remains unknown, although an idea gaining currency is that movement serves as a stringency factor, eliminating unwanted inter-chromosomal associations or entanglements that have arisen as part of the homolog pairing process and, potentially, unwanted associations of chromatin with the nuclear envelope. Turbulent chromosome movements observed during bipolar orientation of chromosomes for segregation could also serve similar roles during mitosis. Recent advances shed light on the contribution of protein complexes involved in the meiotic movements in chromosome dynamics during the mitotic program.
Higher-order chromosome folding and segregation are tightly regulated in all domains of life. In bacteria, details on nucleoid organization regulatory mechanisms and function remain poorly ...characterized, especially in non-model species. Here, we investigate the role of DNA-partitioning protein ParB and SMC condensin complexes in the actinobacterium Corynebacterium glutamicum. Chromosome conformation capture reveals SMC-mediated long-range interactions around ten centromere-like parS sites clustered at the replication origin (oriC). At least one oriC-proximal parS site is necessary for reliable chromosome segregation. We use chromatin immunoprecipitation and photoactivated single-molecule localization microscopy to show the formation of distinct, parS-dependent ParB-nucleoprotein subclusters. We further show that SMC/ScpAB complexes, loaded via ParB at parS sites, mediate chromosomal inter-arm contacts (as previously shown in Bacillus subtilis). However, the MukBEF-like SMC complex MksBEFG does not contribute to chromosomal DNA-folding; instead, this complex is involved in plasmid maintenance and interacts with the polar oriC-tethering factor DivIVA. Our results complement current models of ParB-SMC/ScpAB crosstalk and show that some condensin complexes evolved functions that are apparently uncoupled from chromosome folding.
Bacteriophages play important roles in regulating the intestinal human microbiota composition, dynamics, and homeostasis, and characterizing their bacterial hosts is needed to understand their ...impact. We applied a metagenomic Hi-C approach on 10 healthy human gut samples to unveil a large infection network encompassing more than 6000 interactions bridging a metagenomic assembled genomes (MAGs) and a phage sequence, allowing to study in situ phage-host ratio. Whereas three-quarters of these sequences likely correspond to dormant prophages, 5% exhibit a much higher coverage than their associated MAG, representing potentially actively replicating phages. We detected 17 sequences of members of the crAss-like phage family, whose hosts diversity remained until recently relatively elusive. For each of them, a unique bacterial host was identified, all belonging to different genus of Bacteroidetes. Therefore, metaHiC deciphers infection network of microbial population with a high specificity paving the way to dynamic analysis of mobile genetic elements in complex ecosystems.
As in eukaryotes, bacterial genomes are not randomly folded. Bacterial genetic information is generally carried on a circular chromosome with a single origin of replication from which two replication ...forks proceed bidirectionally toward the opposite terminus region. Here, we investigate the higher-order architecture of the Escherichia coli genome, showing its partition into two structurally distinct entities by a complex and intertwined network of contacts: the replication terminus (ter) region and the rest of the chromosome. Outside of ter, the condensin MukBEF and the ubiquitous nucleoid-associated protein (NAP) HU promote DNA contacts in the megabase range. Within ter, the MatP protein prevents MukBEF activity, and contacts are restricted to ∼280 kb, creating a domain with distinct structural properties. We also show how other NAPs contribute to nucleoid organization, such as H-NS, which restricts short-range interactions. Combined, these results reveal the contributions of major evolutionarily conserved proteins in a bacterial chromosome organization.
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•Hi-C approach confirms the segmentation of the E. coli chromosome into macrodomains•The multilevel organization is mediated by ubiquitous nucleoid-associated proteins•Condensin and histone-like HU protein promote long-range contacts in chromosome arms•The protein MatP insulates the ter region from the structuring action of condensin
Contacts within the E. coli chromosome effectively divide it into functionally distinct structural regions.
Chromatin organization has been increasingly studied in relation with its important influence on DNA-related metabolic processes such as replication or regulation of gene expression. Since its ...original design ten years ago, capture of chromosome conformation (3C) has become an essential tool to investigate the overall conformation of chromosomes. It relies on the capture of long-range trans and cis interactions of chromosomal segments whose relative proportions in the final bank reflect their frequencies of interactions, hence their spatial proximity in a population of cells. The recent coupling of 3C with deep sequencing approaches now allows the generation of high resolution genome-wide chromosomal contact maps. Different protocols have been used to generate such maps in various organisms. This includes mammals, drosophila and yeast. The massive amount of raw data generated by the genomic 3C has to be carefully processed to alleviate the various biases and byproducts generated by the experiments. Our study aims at proposing a simple normalization procedure to minimize the influence of these unwanted but inevitable events on the final results.
Careful analysis of the raw data generated previously for budding yeast S. cerevisiae led to the identification of three main biases affecting the final datasets, including a previously unknown bias resulting from the circularization of DNA molecules. We then developed a simple normalization procedure to process the data and allow the generation of a normalized, highly contrasted, chromosomal contact map for S. cerevisiae. The same method was then extended to the first human genome contact map. Using the normalized data, we revisited the preferential interactions originally described between subsets of discrete chromosomal features. Notably, the detection of preferential interactions between tRNA in yeast and CTCF, PolII binding sites in human can vary with the normalization procedure used.
We quantitatively reanalyzed the genomic 3C data obtained for S. cerevisiae, identified some of the biases inherent to the technique and proposed a simple normalization procedure to analyse them. Such an approach can be easily generalized for genomic 3C experiments in other organisms. More experiments and analysis will be necessary to reach optimal resolution and accuracies of the maps generated through these approaches. Working with cell population presenting highest levels of homogeneity will prove useful in this regards.
Abstract
Chromosomes of all species studied so far display a variety of higher-order organisational features, such as self-interacting domains or loops. These structures, which are often associated ...to biological functions, form distinct, visible patterns on genome-wide contact maps generated by chromosome conformation capture approaches such as Hi-C. Here we present Chromosight, an algorithm inspired from computer vision that can detect patterns in contact maps. Chromosight has greater sensitivity than existing methods on synthetic simulated data, while being faster and applicable to any type of genomes, including bacteria, viruses, yeasts and mammals. Our method does not require any prior training dataset and works well with default parameters on data generated with various protocols.
Chromosomes of a broad range of species, from bacteria to mammals, are structured by large topological domains whose precise functional roles and regulatory mechanisms remain elusive. Here, we ...combine super-resolution microscopies and chromosome-capture technologies to unravel the higher-order organization of the Bacillus subtilis chromosome and its dynamic rearrangements during the cell cycle. We decipher the fine 3D architecture of the origin domain, revealing folding motifs regulated by condensin-like complexes. This organization, along with global folding throughout the genome, is present before replication, disrupted by active DNA replication, and re-established thereafter. Single-cell analysis revealed a strict correspondence between sub-cellular localization of origin domains and their condensation state. Our results suggest that the precise 3D folding pattern of the origin domain plays a role in the regulation of replication initiation, chromosome organization, and DNA segregation.
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•B. subtilis chromosome arms are fully zipped by condensin-like complexes•Architecture of the origin domain may play a role in the regulation of replication•The origin domain dynamically unfolds and refolds in concert with DNA replication•Origin folding requires condensin and the bacterial mitotic-like partition system
Marbouty et al. combine super-resolution microscopies and chromosome-capture technologies to unveil the higher-order organization of the Bacillus subtilis chromosome. They describe the dynamic rearrangements of the origin domain architecture during the cell cycle and dissect the molecular mechanisms and processes involved.
Homologous recombination repairs DNA double-strand breaks (DSB) using an intact dsDNA molecule as a template. It entails a homology search step, carried out along a conserved RecA/Rad51-ssDNA ...filament assembled on each DSB end. Whether, how and to what extent a DSB impacts chromatin folding, and how this (re)organization in turns influences the homology search process, remain ill-defined. Here we characterize two layers of spatial chromatin reorganization following DSB formation in Saccharomyces cerevisiae. Although cohesin folds chromosomes into cohesive arrays of ~20-kb-long chromatin loops as cells arrest in G2/M, the DSB-flanking regions interact locally in a resection- and 9-1-1 clamp-dependent manner, independently of cohesin, Mec1
, Rad52 and Rad51. This local structure blocks cohesin progression, constraining the DSB region at the base of a loop. Functionally, cohesin promotes DSB-dsDNA interactions and donor identification in cis, while inhibiting them in trans. This study identifies multiple direct and indirect ways by which cohesin regulates homology search during recombinational DNA repair.
Genomic analyses of microbial populations in their natural environment remain limited by the difficulty to assemble full genomes of individual species. Consequently, the chromosome organization of ...microorganisms has been investigated in a few model species, but the extent to which the features described can be generalized to other taxa remains unknown. Using controlled mixes of bacterial and yeast species, we developed meta3C, a metagenomic chromosome conformation capture approach that allows characterizing individual genomes and their average organization within a mix of organisms. Not only can meta3C be applied to species already sequenced, but a single meta3C library can be used for assembling, scaffolding and characterizing the tridimensional organization of unknown genomes. By applying meta3C to a semi-complex environmental sample, we confirmed its promising potential. Overall, this first meta3C study highlights the remarkable diversity of microorganisms chromosome organization, while providing an elegant and integrated approach to metagenomic analysis.
Genome-wide chromatin conformation capture assays provide formidable insights into the spatial organization of genomes. However, due to the complexity of the data structure, their integration in ...multi-omics workflows remains challenging. We present data structures, computational methods and visualization tools available in Bioconductor to investigate Hi-C, micro-C and other 3C-related data, in R. An online book ( https://bioconductor.org/books/OHCA/ ) further provides prospective end users with a number of workflows to process, import, analyze and visualize any type of chromosome conformation capture data.
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