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Accurate duplication and separation of long linear genomic DNA molecules is associated with a number of purely mechanical problems. SMC complexes are key components of the cellular machinery that ensures decatenation of sister chromosomes and compaction of genomic DNA during division. Cohesin, one of the essential eukaryotic SMC complexes, has a typical ring structure with intersubunit pore through which DNA molecules can be threaded. Capacity of cohesin for such topological entrapment of DNA is crucial for the phenomenon of post-replicative association of sister chromatids better known as cohesion. Recently, it became apparent that cohesin and other SMC complexes are, in fact, motor proteins with a very peculiar movement pattern leading to formation of DNA loops. This specific process has been called loop extrusion. Extrusion underlies multiple functions of cohesin beyond cohesion, but molecular mechanism of the process remains a mystery. In this review, we summarized the data on molecular architecture of cohesin, effect of ATP hydrolysis cycle on this architecture, and known modes of cohesin-DNA interactions. Many of the seemingly disparate facts presented here will probably be incorporated in a unified mechanistic model of loop extrusion in the not-so-distant future.
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Cohesinas , ADN , Animales , Humanos , Adenosina Trifosfato/metabolismo , Adenosina Trifosfato/química , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/química , Cromátides/metabolismo , Cromátides/química , Proteínas Cromosómicas no Histona/metabolismo , Proteínas Cromosómicas no Histona/química , Cohesinas/química , Cohesinas/metabolismo , ADN/metabolismo , ADN/químicaRESUMEN
The most prominent representatives of multisubunit SMC complexes, cohesin and condensin, are best known as structural components of mitotic chromosomes. It turned out that these complexes, as well as their bacterial homologues, are molecular motors, the ATP-dependent movement of these complexes along DNA threads leads to the formation of DNA loops. In recent years, we have witnessed an avalanche-like accumulation of data on the process of SMC dependent DNA looping, also known as loop extrusion. This review briefly summarizes the current understanding of the place and role of cohesin-dependent extrusion in cell physiology and presents a number of models describing the potential molecular mechanism of extrusion in a most compelling way. We conclude the review with a discussion of how the capacity of cohesin to extrude DNA loops may be mechanistically linked to its involvement in sister chromatid cohesion.
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Fenómenos Fisiológicos Celulares , Cohesinas , Animales , Humanos , Adenosina Trifosfatasas/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/química , Cromátides/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , Proteínas Cromosómicas no Histona/química , Cohesinas/metabolismo , ADN/metabolismo , ADN/química , Proteínas de Unión al ADN/metabolismo , Complejos Multiproteicos/metabolismo , Complejos Multiproteicos/químicaRESUMEN
Solutions of polyelectrolytes consisting of polycations and polyanions in equal proportions were studied in the present work. Due to the physical cross-links formed by the charged groups, physical gels were formed in such systems. The mechanical properties and structure of the obtained gels depending on the charge arrangement along the backbone and the dimensionless Bjerrum length λ were investigated. The response of the systems to a uniaxial affine deformation was studied first. It was found that the systems can be divided into three groups depending on the charge arrangement: showing an almost elastic response; showing a viscoelastic response with a very long relaxation time; and showing a weak viscoelastic response with a short relaxation time. Interestingly, no stable aggregates were formed in the systems with the charges located on spacers, probably because of the increased mobility of the charges in such systems. The obtained stress relaxation curves had different functional forms, indicating that the relaxation has at least two characteristic times, which are different for different systems. In order to understand the molecular nature of the observed mechanical response, the temporal evolution of the network structure of a system showing a viscoelastic response with a very long relaxation time was studied; the aggregates were found to be dynamic, which leads to the relaxation of the "subchains" conformation.
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Proximity ligation approaches, which are widely used to study the spatial organization of the genome, also make it possible to reveal patterns of RNA-DNA interactions. Here, we use RedC, an RNA-DNA proximity ligation approach, to assess the distribution of major RNA types along the genomes of E. coli, B. subtilis, and thermophilic archaeon T. adornatum. We find that (i) messenger RNAs preferentially interact with their cognate genes and the genes located downstream in the same operon, which is consistent with polycistronic transcription; (ii) ribosomal RNAs preferentially interact with active protein-coding genes in both bacteria and archaea, indicating co-transcriptional translation; and (iii) 6S noncoding RNA, a negative regulator of bacterial transcription, is depleted from active genes in E. coli and B. subtilis. We conclude that the RedC data provide a rich resource for studying both transcription dynamics and the function of noncoding RNAs in microbial organisms.
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Escherichia coli , Regulación Bacteriana de la Expresión Génica , Escherichia coli/genética , ADN , Bacterias/genética , OperónRESUMEN
In this work, the question of the influence of the counterion size on the self-assembly in melts of diblock copolymers with one charged block was studied using coarse-grained molecular dynamics simulations. It was assumed that the blocks were fully compatible, i.e., the Flory-Huggins parameter χ between them was equal to 0. Due to the presence of correlation attraction (electrostatic cohesion) between the charged species, the systems with all types of counterions underwent transitions to ordered states, forming various morphologies, including lamellae, perforated lamellae, and hexagonally packed cylinders. Phase diagrams were constructed by varying the chain composition fc and locating the order-disorder transition positions in terms of the electrostatic strength parameter λ (dimensionless Bjerrum length). Despite having a rather large ion size mismatch, the systems with smaller counterions demonstrated an even better tendency to form microphase separated states than the systems with larger ones. It was found that the differences between the phase diagrams of the systems with different counterions can be roughly rationalized by using coordinates (volume fraction of the charged block φc-modified interaction parameter λ*). The latter parameter assumes that the electrostatic energy is simply inversely proportional to the characteristic distance between the ions of different signs. Such an approach appeared to be rather effective and allowed the diagrams obtained for different counterion sizes to almost coincide. The results of this work suggest that the counterion size can be used as a tool to control the system morphology as well as the effective incompatibility between the blocks.
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The copolymer sequence can be considered as a new tool to shape the resulting system properties on demand. This perspective is devoted to copolymers with "partially segregated" (or nonblocky) sequences. Such copolymers include gradient copolymers and copolymers with random sequences as well as copolymers with precisely controlled sequences. We overview recent developments in the synthesis of these systems as well as new findings regarding their properties, in particular, self-assembly in solutions and in melts. An emphasis is put on how the microscopic behavior of polymer chains is influenced by the chain sequences. In addition to that, a novel class of approaches allowing one to efficiently tackle the problem of copolymer chain sequence designâdata driven methods (artificial intelligence and machine learning)âis discussed.
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In the current work, atom transfer radical polymerization-induced self-assembly (ATRP PISA) phase diagrams were obtained by the means of dissipative particle dynamics simulations. A fast algorithm for determining the equilibrium morphology of block copolymer aggregates was developed. Our goal was to assess how the chemical nature of ATRP affects the self-assembly of diblock copolymers in the course of PISA. We discovered that the chain growth termination via recombination played a key role in determining the ATRP PISA phase diagrams. In particular, ATRP with turned off recombination yielded a PISA phase diagram very similar to that obtained for a simple ideal living polymerization process. However, an increase in the recombination probability led to a significant change of the phase diagram: the transition between cylindrical micelles and vesicles was strongly shifted, and a dependence of the aggregate morphology on the concentration was observed. We speculate that this effect occurred due to the simultaneous action of two factors: the triblock copolymer architecture of the terminated chains and the dispersity of the solvophobic blocks. We showed that these two factors affected the phase diagram weakly if they acted separately; however, their combination, which naturally occurs during ATRP, affected the ATRP PISA phase diagram strongly. We suggest that the recombination reaction is a key factor leading to the complexity of experimental PISA phase diagrams.
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Topoisomerase inhibitors are widely used in cancer chemotherapy. However, one of the potential long-term adverse effects of such therapy is acute leukemia. A key feature of such therapy-induced acute myeloid leukemia (t-AML) is recurrent chromosomal translocations involving AML1 (RUNX1) or MLL (KMT2A) genes. The formation of chromosomal translocation depends on the spatial proximity of translocation partners and the mobility of the DNA ends. It is unclear which of these two factors might be decisive for recurrent t-AML translocations. Here, we used fluorescence in situ hybridization (FISH) and chromosome conformation capture followed by sequencing (4C-seq) to investigate double-strand DNA break formation and the mobility of broken ends upon etoposide treatment, as well as contacts between translocation partner genes. We detected the separation of the parts of the broken AML1 gene, as well as the increased mobility of these separated parts. 4C-seq analysis showed no evident contacts of AML1 and MLL with loci, implicated in recurrent t-AML translocations, either before or after etoposide treatment. We suggest that separation of the break ends and their increased non-targeted mobility-but not spatial predisposition of the rearrangement partners-plays a major role in the formation of these translocations.
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Leucemia Mieloide Aguda , Translocación Genética , ADN , Roturas del ADN de Doble Cadena , Etopósido/efectos adversos , Humanos , Hibridación Fluorescente in Situ , Leucemia Mieloide Aguda/genética , Inhibidores de Topoisomerasa II/efectos adversosRESUMEN
Correction for 'Effect of network topology and crosslinker reactivity on microgel structure and ordering at liquid-liquid interface' by Rustam A. Gumerov et al., Soft Matter, 2022, 18, 3738-3747, https://doi.org/10.1039/D2SM00269H.
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Polymer microgels synthesized in silico were studied at a liquid-liquid interface via mesoscopic computer simulations and compared to microgels with ideal (diamond-like) structure. The effect of crosslinkers reactivity ratio on the single particle morphology at the interface and monolayer behavior was examined. It was demonstrated that single particles deform into an explicit core-corona morphology when adsorbed at the interface. An increase in the crosslinker reactivity ratio decreased both the deformation ratio and the ratio between the core and corona sizes. Meanwhile, the compression of microgel monolayers revealed the existence of five distinct interparticle contact regimes, which have been observed experimentally in the literature. The crosslinker reactivity ratio appeared to define the compression range in these regimes and the sharpness of the transition between them. In particular, the higher the crosslinker reactivity ratio, the smaller the corona, and in turn, the narrower the range of the intermediate regime comprising both core-core and corona-corona contacts. The obtained results demonstrate that the more realistic model of microgels synthesized via precipitation polymerization allows for a more accurate prediction of the properties of the microgels at a liquid-liquid interface in comparison to the conventional diamond-like lattice model.
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In this work, we studied the equilibrium structures formed by a single (AB)k multiblock copolymer chain. Within our model, the interactions between the A-type beads were repulsive and the B-type beads could form pairwise reversible bonds with each other (BB-bonds). Our goal was to investigate how the formation of pairwise reversible bonds between the A-type beads and the B-type beads (AB-bonds) affected the structure of the chain. We observed the formation of well-studied intramolecular micelles when the AB-bonds were absent; however, the chain folding changed dramatically when the formation of the AB-bonds was introduced. In this case, the multiblock copolymer formed a globule, which had a unique heterogeneous checkerboard-like distribution of the contact density. We discovered that contacts of beads of different types (i.e., AB-contacts) occurred much more frequently than contacts of beads of the same type (i.e., AA- and BB-contacts) in these structures. This effect can be explained by a simple model of chemical equilibrium in a two-component fluid of reversibly interacting particles, which can be solved exactly. This novel type of folding can serve as a basic model for any (AB)k multiblock copolymer chain with a non-vanishing attraction between A and B blocks.
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Nuclear noncoding RNAs (ncRNAs) are key regulators of gene expression and chromatin organization. The progress in studying nuclear ncRNAs depends on the ability to identify the genome-wide spectrum of contacts of ncRNAs with chromatin. To address this question, a panel of RNA-DNA proximity ligation techniques has been developed. However, neither of these techniques examines proteins involved in RNA-chromatin interactions. Here, we introduce RedChIP, a technique combining RNA-DNA proximity ligation and chromatin immunoprecipitation for identifying RNA-chromatin interactions mediated by a particular protein. Using antibodies against architectural protein CTCF and the EZH2 subunit of the Polycomb repressive complex 2, we identify a spectrum of cis- and trans-acting ncRNAs enriched at Polycomb- and CTCF-binding sites in human cells, which may be involved in Polycomb-mediated gene repression and CTCF-dependent chromatin looping. By providing a protein-centric view of RNA-DNA interactions, RedChIP represents an important tool for studies of nuclear ncRNAs.
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Factor de Unión a CCCTC/metabolismo , Proteínas del Grupo Polycomb/metabolismo , ARN no Traducido/metabolismo , Inmunoprecipitación de Cromatina , Proteínas de Unión al ADN/metabolismo , HumanosRESUMEN
In this work using dissipative particle dynamics simulations with explicit treatment of polar species we demonstrate that the molecular nature of dielectric media has a significant impact on swelling and collapse of a polyelectrolyte chain in a dilute solution. We show that the small-scale effects related to the presence of polar species lead to the intensification of the electrostatic interactions when the charges are close to each other and/or their density is high enough. As a result, the electrostatic strength , usually regarded as the main parameter governing the polyelectrolyte chain collapse, does not have a universal meaning: the value of λ at which the coil-to-globule transition occurs is found to be dependent on the specific fixed value of the solvent bulk permittivity ε while varying the monomer unit charge Q and vice versa. This effect is observed even when the backbone and the counterions have the same polarity as the solvent beads, i.e. no dielectric mismatch is present. The reason for such behavior is rationalized in terms of the "effective" dielectric permittivity εeff which depends on the volume fraction φ of charged units inside the polymer chain volume; using εeff instead of ε collapses all data onto one master curve describing the chain shrinking with λ. Furthermore, it is shown that a polar chain adopts less swollen conformations in the polyelectrolyte regime and collapses more easily compared to a non-polar chain.
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Cohesin is a key organizer of chromatin folding in eukaryotic cells. The two main activities of this ring-shaped protein complex are the maintenance of sister chromatid cohesion and the establishment of long-range DNA-DNA interactions through the process of loop extrusion. Although the basic principles of both cohesion and loop extrusion have been described, we still do not understand several crucial mechanistic details. One of such unresolved issues is the question of whether a cohesin ring topologically embraces DNA string(s) during loop extrusion. Here, we show that cohesin complexes residing on CTCF-occupied genomic sites in mammalian cells do not interact with DNA topologically. We assessed the stability of cohesin-dependent loops and cohesin association with chromatin in high-ionic-strength conditions in G1-synchronized HeLa cells. We found that increased salt concentration completely displaces cohesin from those genomic regions that correspond to CTCF-defined loop anchors. Unsurprisingly, CTCF-anchored cohesin loops also dissipate in these conditions. Because topologically engaged cohesin is considered to be salt resistant, our data corroborate a non-topological model of loop extrusion. We also propose a model of cohesin activity throughout the interphase, which essentially equates the termination of non-topological loop extrusion with topological loading of cohesin. This theoretical framework enables a parsimonious explanation of various seemingly contradictory experimental findings.
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Cromatina , Proteínas Cromosómicas no Histona , Animales , Proteínas de Ciclo Celular/genética , Proteínas Cromosómicas no Histona/genética , Células HeLa , Humanos , CohesinasRESUMEN
One of the most intriguing questions facing modern biology concerns how the genome directs the construction of cells, tissues, and whole organisms. It is tempting to suggest that the part of the genome that does not encode proteins contains architectural plans. We are still far from understanding how these plans work at the level of building tissues and the body as a whole. However, the results of recent studies demonstrate that at the cellular level, special non-coding RNAs serve as scaffolds for the construction of various intracellular structures. The term "architectural RNAs" was proposed to designate this subset of non-coding RNAs. In this review, we discuss the role of architectural RNAs in the construction of the cell nucleus and maintenance of the three-dimensional organization of the genome.
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Núcleo Celular/genética , Cromatina/química , Cromatina/genética , Genoma , ARN Largo no Codificante/genética , Animales , HumanosRESUMEN
HYPOTHESIS: Wormlike surfactant micelles (WLMs) are prospective as nanoreactors for micellar copolymerization of hydrophilic and hydrophobic monomers. Hydrophilic monomers can destroy WLMs. Large size and cylindrical shape of micelles can be preserved by high salt content favoring closer packing of surfactant heads. EXPERIMENTS: The effect of a water-soluble monomer (acrylamide) on the structure and rheological properties of giant WLMs of an anionic surfactant potassium oleate at different salt content was investigated by combined experimental (SANS, rheometry, fluorescence and NMR spectroscopy, tensiometry) and molecular dynamics simulations studies. FINDINGS: At low salt content, when WLMs are linear, acrylamide induces their shortening and transformation into spherical micelles as a result of its incorporation into the micellar corona, leading to the drop of viscosity. At high salt content providing branched WLMs, monomer first triggers their transition to long linear chains, which enhances the viscoelasticity, and then to rods. This is the first report showing that the effect of monomer on the rheological properties is quite different for linear and branched micelles. Using branched micelles allows preserving large WLMs at high water-soluble monomer content, which is favorable for their use as nanoreactors for synthesis of copolymers with high degree of blockiness, which give mechanically tough polymer gels.
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Micelas , Tensoactivos , Estudios Prospectivos , Viscosidad , AguaRESUMEN
Liquid-liquid phase separation (LLPS) contributes to the spatial and functional segregation of molecular processes within the cell nucleus. However, the role played by LLPS in chromatin folding in living cells remains unclear. Here, using stochastic optical reconstruction microscopy (STORM) and Hi-C techniques, we studied the effects of 1,6-hexanediol (1,6-HD)-mediated LLPS disruption/modulation on higher-order chromatin organization in living cells. We found that 1,6-HD treatment caused the enlargement of nucleosome clutches and their more uniform distribution in the nuclear space. At a megabase-scale, chromatin underwent moderate but irreversible perturbations that resulted in the partial mixing of A and B compartments. The removal of 1,6-HD from the culture medium did not allow chromatin to acquire initial configurations, and resulted in more compact repressed chromatin than in untreated cells. 1,6-HD treatment also weakened enhancer-promoter interactions and TAD insulation but did not considerably affect CTCF-dependent loops. Our results suggest that 1,6-HD-sensitive LLPS plays a limited role in chromatin spatial organization by constraining its folding patterns and facilitating compartmentalization at different levels.
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Cromatina/química , Glicoles/farmacología , Cromatina/efectos de los fármacos , Elementos de Facilitación Genéticos/efectos de los fármacos , Genoma Humano , Células HeLa , Humanos , Microscopía , Regiones Promotoras Genéticas/efectos de los fármacosRESUMEN
Chromatin loops represent one of the major levels of hierarchical folding of the genome. Although the situation is evolving, current methods have various difficulties with the accurate mapping of loops even in mammalian Hi-C data, and most of them fail to identify chromatin loops in animal species with substantially different genome architecture. This paper presents the loop and significant contact annotation (LASCA) pipeline, which uses Weibull distribution-based modeling to effectively identify loops and enhancer-promoter interactions in Hi-C data from evolutionarily distant species: from yeast and worms to mammals. Available at: https://github.com/ArtemLuzhin/LASCA_pipeline .
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Cromatina/genética , Elementos de Facilitación Genéticos/genética , Genoma/genética , Regiones Promotoras Genéticas/genética , Animales , Cromosomas/genética , Genómica , Humanos , Mamíferos/genética , Anotación de Secuencia Molecular , Programas Informáticos , Levaduras/genéticaRESUMEN
In this Note, we study the total conservative force {instead of pure electrostatic force as it was carried out in the work by Gavrilov [J. Chem. Phys. 152, 164101 (2020)]} acting on two charges in a polar liquid using dissipative particle dynamics and coarse-grained molecular dynamics simulations. We show that such force (instead of the electrostatic force) obeys Coulomb's law at large distances between the charges. Apparently, the dielectric response of a polar liquid (at least, within such coarse-grained models) can be decomposed into two contributions: the reorientation of the dipoles (i.e., electrostatic contribution) and the density redistribution (i.e., volume interaction contribution).
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Mammalian and Drosophila genomes are partitioned into topologically associating domains (TADs). Although this partitioning has been reported to be functionally relevant, it is unclear whether TADs represent true physical units located at the same genomic positions in each cell nucleus or emerge as an average of numerous alternative chromatin folding patterns in a cell population. Here, we use a single-nucleus Hi-C technique to construct high-resolution Hi-C maps in individual Drosophila genomes. These maps demonstrate chromatin compartmentalization at the megabase scale and partitioning of the genome into non-hierarchical TADs at the scale of 100 kb, which closely resembles the TAD profile in the bulk in situ Hi-C data. Over 40% of TAD boundaries are conserved between individual nuclei and possess a high level of active epigenetic marks. Polymer simulations demonstrate that chromatin folding is best described by the random walk model within TADs and is most suitably approximated by a crumpled globule build of Gaussian blobs at longer distances. We observe prominent cell-to-cell variability in the long-range contacts between either active genome loci or between Polycomb-bound regions, suggesting an important contribution of stochastic processes to the formation of the Drosophila 3D genome.