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Chromosome compaction is essential for reliable transmission of genetic information. Experiments suggest that ~1000-fold compaction is driven by condensin complexes that extrude chromatin loops, by progressively collecting chromatin fiber from one or both sides of the complex to form a growing loop. Theory indicates that symmetric two-sided loop extrusion can achieve such compaction, but recent single-molecule studies (Golfier et al., 2020) observed diverse dynamics of condensins that perform one-sided, symmetric two-sided, and asymmetric two-sided extrusion. We use simulations and theory to determine how these molecular properties lead to chromosome compaction. High compaction can be achieved if even a small fraction of condensins have two essential properties: a long residence time and the ability to perform two-sided (not necessarily symmetric) extrusion. In mixtures of condensins I and II, coupling two-sided extrusion and stable chromatin binding by condensin II promotes compaction. These results provide missing connections between single-molecule observations and chromosome-scale organization. [ABSTRACT FROM AUTHOR]

Chromosomes are organized as chromatin loops that promote segregation, enhancer-promoter interactions, and other genomic functions. Loops were hypothesized to form by 'loop extrusion,' by which structural maintenance of chromosomes (SMC) complexes, such as condensin and cohesin, bind to chromatin, reel it in, and extrude it as a loop. However, such exotic motor activity had never been observed. Following an explosion of indirect evidence, recent single-molecule experiments directly imaged DNA loop extrusion by condensin and cohesin in vitro. These experiments observe rapid (kb/s) extrusion that requires ATP hydrolysis and stalls under pN forces. Surprisingly, condensin extrudes loops asymmetrically, challenging previous models. Extrusion by cohesin is symmetric but requires the protein Nipbl. We discuss how SMC complexes may perform their functions on chromatin in vivo. [ABSTRACT FROM AUTHOR]

Motivation Most existing coverage-based (epi)genomic datasets are one-dimensional, but newer technologies probing interactions (physical, genetic, etc.) produce quantitative maps with two-dimensional genomic coordinate systems. Storage and computational costs mount sharply with data resolution when such maps are stored in dense form. Hence, there is a pressing need to develop data storage strategies that handle the full range of useful resolutions in multidimensional genomic datasets by taking advantage of their sparse nature, while supporting efficient compression and providing fast random access to facilitate development of scalable algorithms for data analysis. Results We developed a file format called cooler, based on a sparse data model, that can support genomically labeled matrices at any resolution. It has the flexibility to accommodate various descriptions of the data axes (genomic coordinates, tracks and bin annotations), resolutions, data density patterns and metadata. Cooler is based on HDF5 and is supported by a Python library and command line suite to create, read, inspect and manipulate cooler data collections. The format has been adopted as a standard by the NIH 4D Nucleome Consortium. Availability and implementation Cooler is cross-platform, BSD-licensed and can be installed from the Python package index or the bioconda repository. The source code is maintained on Github at https://github.com/mirnylab/cooler. Supplementary information Supplementary data are available at Bioinformatics online. [ABSTRACT FROM AUTHOR]

The spatial organization of chromosomes has long been connected to their polymeric nature and is believed to be important for their biological functions, including the control of interactions between genomic elements, the maintenance of genetic information, and the compaction and safe transfer of chromosomes to cellular progeny. chromosome conformation capture techniques, particularly Hi-C, have provided a comprehensive picture of spatial chromosome organization and revealed new features and elements of chromosome folding. Furthermore, recent advances in microscopy have made it possible to obtain distance maps for extensive regions of chromosomes (Bintu et al. , 2018; Nir et al. , 2018 [ 2•• , 3 ]), providing information complementary to, and in excellent agreement with, Hi-C maps. Not only has the resolution of both techniques advanced significantly, but new perturbation data generated in the last two years have led to the identification of molecular mechanisms behind large-scale genome organization. Two major mechanisms that have been proposed to govern chromosome organization are (i) the active (ATP-dependent) process of loop extrusion by Structural Maintenance of Chromosomes (SMC) complexes, and (ii) the spatial compartmentalization of the genome, which is likely mediated by affinity interactions between heterochromatic regions (Falk et al., 2019 [ 76•• ]) rather than by ATP-dependent processes. Here, we review existing evidence that these two processes operate together to fold chromosomes in interphase and that loop extrusion alone drives mitotic compaction. We discuss possible implications of these mechanisms for chromosome function. [ABSTRACT FROM AUTHOR]

Structural biology has traditionally focused on the structures of proteins, short nucleic acids, small molecules, and their complexes. However, it is now widely recognized that the 3D organization of chromosomes should also be included in this list, despite significant differences in scale and complexity of organization. Here we highlight some notable similarities between the folding processes that shape proteins and chromosomes. Both biomolecules are folded by two types of processes: the affinity-mediated interactions, and by active (ATP-dependent) processes. Both chromosome and proteins in vivo can have partially unstructured and non-equilibrium ensembles with yet to be understood functional roles. By analyzing these biological systems in parallel, we can uncover universal principles of biomolecular organization that transcend specific biopolymers. • Interphase chromosomes are not nearly as structured as folded proteins, yet chromosomes are not unstructured either. • Proteins and chromosomes are folded by affinities between their elements and by active processes. • Active processes of folding are loop extrusion in chromosomes, and co-translational and chaperon-mediated in proteins. • Active processes not only facilitate folding but also generate folding patterns by placing pause site along the sequence. • Functional roles of these activity-driven and not fully structured organizations of both biopolymers are yet to be understood. [ABSTRACT FROM AUTHOR]

Proper expression of genes requires communication with their regulatory elements that can be located elsewhere along the chromosome. The physics of chromatin fibers imposes a range of constraints on such communication. The molecular and biophysical mechanisms by which chromosomal communication is established, or prevented, have become a topic of intense study, and important roles for the spatial organization of chromosomes are being discovered. Here we present a view of the interphase 3D genome characterized by extensive physical compartmentalization and insulation on the one hand and facilitated long-range interactions on the other. We propose the existence of topological machines dedicated to set up and to exploit a 3D genome organization to both promote and censor communication along and between chromosomes. [ABSTRACT FROM AUTHOR]

Cancer progression is an example of a rapid adaptive process where evolving new traits is essential for survival and requires a high mutation rate. Precancerous cells acquire a few key mutations that drive rapid population growth and carcinogenesis. Cancer genomics demonstrates that these few driver mutations occur alongside thousands of random passenger mutations--a natural consequence of cancer's elevated mutation rate. Some passengers are deleterious to cancer cells, yet have been largely ignored in cancer research. In population genetics, however, the accumulation of mildly deleterious mutations has been shown to cause population meltdown. Here we develop a stochastic population model where beneficial drivers engage in a tug-of-war with frequent mildly deleterious passengers. These passengers present a barrier to cancer progression describable by a critical population size, below which most lesions fail to progress, and a critical mutation rate, above which cancers melt down. We find support for this model in cancer age-incidence and cancer genomics data that also allow us to estimate the fitness advantage of drivers and fitness costs of passengers. We identify two regimes of adaptive evolutionary dynamics and use these regimes to understand successes and failures of different treatment strategies. A tumor's load of deleterious passengers can explain previously paradoxical treatment outcomes and suggest that it could potentially serve as a biomarker of response to mutagenic therapies. The collective deleterious effect of passengers is currently an unexploited therapeutic target. We discuss how their effects might be exacerbated by current and future therapies. [ABSTRACT FROM AUTHOR]

Advances in microscopy and genomic techniques have provided new insight into spatial chromatin organization inside of the nucleus. In particular, chromosome conformation capture data has highlighted the relevance of polymer physics for high-order chromatin organization. In this context, we review basic polymer states, discuss how an appropriate polymer model can be determined from experimental data, and examine the success and limitations of various polymer models of higher-order interphase chromatin organization. By taking into account topological constraints acting on the chromatin fiber, recently developed polymer models of interphase chromatin can reproduce the observed scaling of distances between genomic loci, chromosomal territories, and probabilities of contacts between loci measured by chromosome conformation capture methods. Polymer models provide a framework for the interpretation of experimental data as ensembles of conformations rather than collections of loops, and will be crucial for untangling functional implications of chromosomal organization. [Copyright &y& Elsevier]

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. [ABSTRACT FROM AUTHOR]

Coordinated regulation of gene expression relies on transcription factors (TFs) binding to specific DNA sites. Our large-scale information–theoretical analysis of >950 TF-binding motifs demonstrates that prokaryotes and eukaryotes use strikingly different strategies to target TFs to specific genome locations. Although bacterial TFs can recognize a specific DNA site in the genomic background, eukaryotic TFs exhibit widespread, nonfunctional binding and require clustering of sites to achieve specificity. We find support for this mechanism in a range of experimental studies and in our evolutionary analysis of DNA-binding domains. Our systematic characterization of binding motifs provides a quantitative assessment of the differences in transcription regulation in prokaryotes and eukaryotes. [Copyright &y& Elsevier]

Our investigation of knotted structures in the Protein Data Bank reveals the most complicated knot discovered to date. We suggest that the occurrence of this knot in a human ubiquitin hydrolase might be related to the role of the enzyme in protein degradation. While knots are usually preserved among homologues, we also identify an exception in a transcarbamylase. This allows us to exemplify the function of knots in proteins and to suggest how they may have been created. [ABSTRACT FROM AUTHOR]

Proteins, nucleic acids, and small molecules form a dense network of molecular interactions in a cell. Molecules are nodes of this network, and the interactions between them are edges. The architecture of molecular networks can reveal important principles of cellular organization and function, similarly to the way that protein structure tells us about the function and organization of a protein. Computational analysis of molecular networks has been primarily concerned with node degree [Wagner, A. & Fell, D. A. (2001) Proc. R. Soc. London Ser. B 268, 1803-1810; Jeong, H., Tombor, B., Albert, R., Oltvai, Z. N. & Barabasi, A. L. (2000) Nature 407, 651-654] or degree correlation [Maslov, S. & Sneppen, K. (2002) Science 296, 910-913], and hence focused on single/two-body properties of these networks. Here, by analyzing the multibody structure of the network of protein-protein interactions, we discovered molecular modules that are densely connected within themselves but sparsely connected with the rest of the network. Comparison with experimental data and functional annotation of genes showed two types of modules: (1) protein complexes (splicing machinery, transcription factors, etc.) and (ill dynamic functional units (signaling cascades, cell-cycle regulation, etc.). Discovered modules are highly statistically significant, as is evident from comparison with random graphs, and are robust to noise in the data. Our results provide strong support for the network modularity principle introduced by Hartwell et al. [Hartwell, L. H., Hopfield, J. J., Leibler, S. & Murray, A. W. (1999) Nature 402, C47-C52], suggesting that found modules constitute the "building blocks" of molecular networks. [ABSTRACT FROM AUTHOR]

It has been conjectured that evolution exerted pressure to preserve amino acids bearing thermodynamic, kinetic, and functional roles. In this letter we show that the physical requirement to maintain protein stability gives rise to a sequence conservatism pattern that is in remarkable agreement with that found in natural proteins. Based on the physical properties of amino acids, we propose a model of evolution that explains conserved amino acids across protein families sharing the same fold. [Copyright &y& Elsevier]

Concepts of orthology and paralogy are become increasingly important as whole-genome comparison allows their identification in complete genomes. Functional specificity of proteins is assumed to be conserved among orthologs and is different among paralogs. We used this assumption to identify residues which determine specificity of protein–DNA and protein–ligand recognition. Finding such residues is crucial for understanding mechanisms of molecular recognition and for rational protein and drug design. Assuming conservation of specificity among orthologs and different specificity of paralogs, we identify residues that correlate with this grouping by specificity. The method is taking advantage of complete genomes to find multiple orthologs and paralogs. The central part of this method is a procedure to compute statistical significance of the predictions. The procedure is based on a simple statistical model of protein evolution. When applied to a large family of bacterial transcription factors, our method identified 12 residues that are presumed to determine the protein–DNA and protein–ligand recognition specificity. Structural analysis of the proteins and available experimental results strongly support our predictions. Our results suggest new experiments aimed at rational re-design of specificity in bacterial transcription factors by a minimal number of mutations. [Copyright &y& Elsevier]

This review focuses on recent advances in understanding protein folding kinetics in the context of nucleation theory. We present basic concepts such as nucleation, folding nucleus, and transition state ensemble and then discuss recent advances and challenges in theoretical understanding of several key aspects of protein folding kinetics. We cover recent topology-based approaches as well as evolutionary studies and molecular dynamics approaches to determine protein folding nucleus and analyze other aspects of folding kinetics. Finally, we briefly discuss successful all-atom Monte-Carlo simulations of protein folding and conclude with a brief outlook for the future. [ABSTRACT FROM AUTHOR]

Estimates the statistical significance of protein structure prediction by sequence-structure alignment. Introduction of a single parameter epsilon that serves as a universal measure determining the probability that the best alignment is a native-like analog; Comparison with gapless threading; Value distribution.

A lattice model with side chains was used to investigate protein folding with computer simulations. In this model, we rigorously demonstrate the existence of a specific folding nucleus. This nucleus contains specific interactions not present in the native state that, when weakened, slow folding but do not change protein stability. Such a decoupling of folding kinetics from thermodynamics has been observed experimentally for real proteins. From our results, we conclude that specific non-native interactions in the transition state would give rise to φ-values that are negative or larger than unity. Furthermore, we demonstrate that residue Ile 34 in src SH3, which has been shown to be kinetically, but not thermodynamically, important, is universally conserved in proteins with the SH3 fold. This is a clear example of evolution optimizing the folding rate of a protein independent of its stability and function. [ABSTRACT FROM AUTHOR]

Provides information on a study of the sequences of fast-folding model proteins (48 residues long on a cubic lattice), which were generated by an evolution-like selection toward fast folding. Methodology used to conduct the study; Results of the study.

The article reports on the effect of Brownian motion in limiting the rates of chemical reactions in a cell. It has been indicated by the author professor Marian Smoluchowski has set the diffusion limit of reaction rates by calculating the relationship between the rates of chemical reactions and the diffusion coefficient of the participating molecules. In addition, the Stokes-Einstein equation indicated that diffusion coefficients vary depending on the size of the molecules.

DNA methylation (DNAme) is a key epigenetic mark that regulates critical biological processes maintaining overall genome stability. Given its pleiotropic function, studies of DNAme dynamics are crucial, but currently available tools to interfere with DNAme have limitations and major cytotoxic side effects. Here, we present cell models that allow inducible and reversible DNAme modulation through DNMT1 depletion. By dynamically assessing whole genome and locus-specific effects of induced passive demethylation through cell divisions, we reveal a cooperative activity between DNMT1 and DNMT3B, but not of DNMT3A, to maintain and control DNAme. We show that gradual loss of DNAme is accompanied by progressive and reversible changes in heterochromatin, compartmentalization, and peripheral localization. DNA methylation loss coincides with a gradual reduction of cell fitness due to G1 arrest, with minor levels of mitotic failure. Altogether, this system allows DNMTs and DNA methylation studies with fine temporal resolution, which may help to reveal the etiologic link between DNAme dysfunction and human disease. [ABSTRACT FROM AUTHOR]

The cell nucleus is a remarkably well-organized organelle with membraneless but distinct compartments of various functions. The largest of them, euchromatin and heterochromatin, are spatially segregated in such a way that the transcriptionally active genome occupies the nuclear interior, whereas silent genomic loci are preferentially associated with the nuclear envelope. This rule is broken by rod photoreceptor cells of nocturnal mammals, in which the two major compartments have inverted positions. The inversion and dense compaction of heterochromatin converts these nuclei into microlenses that focus light and facilitate nocturnal vision. As is often the case in biology, when a mutation helps to understand normal processes and structures, inverted nuclei have served as a tool to unravel general principles of nuclear organization, including mechanisms of heterochromatin tethering to the nuclear envelope, autonomous behavior of small genomic segments, and euchromatin–heterochromatin segregation. The segregation of the eukaryotic genome into active euchromatic and silent heterochromatic nuclear compartments is crucial for the realization of nuclear processes. In most cells euchromatin occupies the nuclear interior, while heterochromatin is found at the nuclear and nucleolar peripheries. Nuclei of rod photoreceptor cells of nocturnal mammals represent the only known unique exception and have an inverted arrangement of the nuclear compartments, with heterochromatin concentrated in the nuclear center and acting as a microlens focusing light, thus facilitating light propagation with reduced photon scattering. Inverted rod nuclei, with their perfectly segregated and concentrically arranged compartments, serve as a model to understand general principles of genome organization such as heterochromatin tethering at the nuclear periphery, compartment recognition, and mechanisms of genome segregation. [ABSTRACT FROM AUTHOR]

The article offers information on the mechanism of epigenetic memory in cells, particularly focusing on the interplay between three-dimensional (3D) genome folding and the dynamics of chromatin marks. Topics include the challenge of maintaining stable epigenetic memory amidst the dynamic nature of individual marks; the role of 3D genome folding in mark spreading; and the development of a theoretical model to explore the dynamics of chromatin and marks through the cell cycle.

The article discusses a genomic approach to analyse chromosomal organization and interactions in individual cells with the help of the chromosome conformation capture (3C) technique. It mentions that the technique produces maps of averaged genome-wide interactions in chromatin that helps to analyse chromosome organization. It states that analysing T helper cells inside the chromosome offers high level of cell-to-cell variability and also mentions high-resolution microscopy.

Recently, Chromosome Conformation Capture (3C) based experiments have highlighted the importance of computational models for the study of chromosome organization. In this review, we propose that current computational models can be grouped into roughly four classes, with two classes of data-driven models: consensus structures and data-driven ensembles, and two classes of de novo models: structural ensembles and mechanistic ensembles. Finally, we highlight specific questions mechanistic ensembles can address. [ABSTRACT FROM AUTHOR]

Our understanding of the physical principles organizing the genome in the nucleus is limited by the lack of tools to directly exert and measure forces on interphase chromosomes in vivo and probe their material nature. Here, we introduce an approach to actively manipulate a genomic locus using controlled magnetic forces inside the nucleus of a living human cell. We observed viscoelastic displacements over micrometers within minutes in response to near-piconewton forces, which are consistent with a Rouse polymer model. Our results highlight the fluidity of chromatin, with a moderate contribution of the surrounding material, revealing minor roles for cross-links and topological effects and challenging the view that interphase chromatin is a gel-like material. Our technology opens avenues for future research in areas from chromosome mechanics to genome functions. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Chromosomes must be highly compacted and organized within cells, but how this is achieved in vivo remains poorly understood. We report the use of chromosome conformation capture coupled with deep sequencing (Hi-C) to map the structure of bacterial chromosomes. Analysis of Hi-C data and polymer modeling indicates that the Caulobacter crescentus chromosome consists of multiple, largely independent spatial domains that are probably composed of supercoiled plectonemes arrayed into a bottle brush-like fiber. These domains are stable throughout the cell cycle and are reestablished concomitantly with DNA replication. We provide evidence that domain boundaries are established by highly expressed genes and the formation of plectoneme-free regions, whereas the histone-like protein HU and SMC (structural maintenance of chromosomes) promote short-range compaction and the colinearity of chromosomal arms, respectively. Collectively, our results reveal general principles for the organization and structure of chromosomes in vivo. [ABSTRACT FROM AUTHOR]

The topological state of chromosomes determines their mechanical properties, dynamics, and function. Recent work indicated that interphase chromosomes are largely free of entanglements. Here, we use Hi-C, polymer simulations, and multi-contact 3C and find that, by contrast, mitotic chromosomes are self-entangled. We explore how a mitotic self-entangled state is converted into an unentangled interphase state during mitotic exit. Most mitotic entanglements are removed during anaphase/telophase, with remaining ones removed during early G1, in a topoisomerase-II-dependent process. Polymer models suggest a two-stage disentanglement pathway: first, decondensation of mitotic chromosomes with remaining condensin loops produces entropic forces that bias topoisomerase II activity toward decatenation. At the second stage, the loops are released, and the formation of new entanglements is prevented by lower topoisomerase II activity, allowing the establishment of unentangled and territorial G1 chromosomes. When mitotic entanglements are not removed in experiments and models, a normal interphase state cannot be acquired. [Display omitted] • Mitotic chromatids are self-entangled, whereas interphase chromosomes are not • Disentanglement during mitotic exit involves regulated topoisomerase II activity • Decondensation drives topoisomerase-II-dependent decatenation of mitotic loops • Removal of mitotic entanglements is required to establish a normal interphase nucleus Recent work indicated that interphase chromosomes are largely free of entanglements. Here, Hildebrand and Polovnikov et al. combine experiments with computer simulations to show that mitotic chromosomes are self-entangled. The results suggest a robust biophysical mechanism of chromosome disentanglement during mitotic exit that relies on regulated activity of topoisomerase II. [ABSTRACT FROM AUTHOR]

How DNA is organized in three dimensions inside the cell nucleus and how this affects the ways in which cells access, read and interpret genetic information are among the longest standing questions in cell biology. Using newly developed molecular, genomic and computational approaches based on the chromosome conformation capture technology (such as 3C, 4C, 5C and Hi-C), the spatial organization of genomes is being explored at unprecedented resolution. Interpreting the increasingly large chromatin interaction data sets is now posing novel challenges. Here we describe several types of statistical and computational approaches that have recently been developed to analyse chromatin interaction data. [ABSTRACT FROM AUTHOR]

Chromosomal translocations are frequent features of cancer genomes that contribute to disease progression. These rearrangements result from formation and illegitimate repair of DNA double-strand breaks (DSBs), a process that requires spatial colocalization of chromosomal breakpoints. The "contact first" hypothesis suggests that translocation partners colocalize in the nuclei of normal cells, prior to rearrangement. It is unclear, however, the extent to which spatial interactions based on three-dimensional genome architecture contribute to chromosomal rearrangements in human disease. Here we intersect Hi-C maps of three-dimensional chromosome conformation with collections of 1,533 chromosomal translocations from cancer and germline genomes. We show that many translocation-prone pairs of regions genome-wide, including the cancer translocation partners BCR-ABL and MYC-IGH, display elevated Hi-C contact frequencies in normal human cells. Considering tissue specificity, we find that translocation breakpoints reported in human hematologic malignancies have higher Hi-C contact frequencies in lymphoid cells than those reported in sarcomas and epithelial tumors. However, translocations from multiple tissue types show significant correlation with Hi-C contact frequencies, suggesting that both tissue-specific and universal features of chromatin structure contribute to chromosomal alterations. Our results demonstrate that three-dimensional genome architecture shapes the landscape of rearrangements directly observed in human disease and establish Hi-C as a key method for dissecting these effects. [ABSTRACT FROM AUTHOR]

A ubiquitous building block of signaling pathways is a cycle of covalent modification (e.g., phosphorylation and dephosphorylation in MAPK cascades). Our paper explores the kind of information processing and filtering that can be accomplished by this simple biochemical circuit. Signaling cycles are particularly known for exhibiting a highly sigmoidal (ultrasensitive) input-output characteristic in a certain steady-state regime. Here, we systematically study the cycle's steady-state behavior and its response to time-varying stimuli. We demonstrate that the cycle can actually operate in four different regimes, each with its specific input-output characteristics. These results are obtained using the total quasi-steady-state approximation, which is more generally valid than the typically used Michaelis-Menten approximation for enzymatic reactions. We invoke experimental data that suggest the possibility of signaling cycles operating in one of the new regimes. We then consider the cycle's dynamic behavior, which has so far been relatively neglected. We demonstrate that the intrinsic architecture of the cycles makes them act-in all four regimes-as tunable low-pass filters, filtering out high-frequency fluctuations or noise in signals and environmental cues. Moreover, the cutoff frequency can be adjusted by the cell. Numerical simulations show that our analytical results hold well even for noise of large amplitude. We suggest that noise filtering and tunability make signaling cycles versatile components of more elaborate cell-signaling pathways. [ABSTRACT FROM AUTHOR]

The binding between a PK and its target is highly specific, despite the fact that many different PKs exhibit significant sequence and structure homology. There must be, then, specificity-determining residues (SDRs) that enable different PKs to recognize their unique substrate. Here we use and further develop a computational procedure to discover putative SDRs (PSDRs) in protein families, whereby a family of homologous proteins is split into orthologous proteins, which are assumed to have the same specificity, and paralogous proteins, which have different specificities. We reason that PSDRs must be similar among orthologs, whereas they must necessarily be different among paralogs. Our statistical procedure and evolutionary model identifies such residues by discriminating a functional signal from a phylogenetic one. As case studies we investigate the prokaryotic two-component system and the eukaryotic AGC (i.e., cAMP-dependent PK, cGMP-dependent PK, and PKC) PKs. Without using experimental data, we predict PSDRs in prokaryotic and eukaryotic PKs, and suggest precise mutations that may convert the specificity of one PK to another. We compare our predictions with current experimental results and obtain considerable agreement with them. Our analysis unifies much of existing data on PK specificity. Finally, we find PSDRs that are outside the active site. Based on our results, as well as structural and biochemical characterizations of eukaryotic PKs, we propose the testable hypothesis of "specificity via differential activation" as a way for the cell to control kinase specificity. [ABSTRACT FROM AUTHOR]

To separate replicated sister chromatids during mitosis, eukaryotes and prokaryotes have structural maintenance of chromosome (SMC) condensin complexes that were recently shown to organize chromosomes by a process known as DNA loop extrusion. In rapidly dividing bacterial cells, the process of separating sister chromatids occurs concomitantly with ongoing transcription. How transcription interferes with the condensin loop-extrusion process is largely unexplored, but recent experiments have shown that sites of high transcription may directionally affect condensin loop extrusion. We quantitatively investigate different mechanisms of interaction between condensin and elongating RNA polymerases (RNAPs) and find that RNAPs are likely steric barriers that can push and interact with condensins. Supported by chromosome conformation capture and chromatin immunoprecipitation for cells after transcription inhibition and RNAP degradation, we argue that translocating condensins must bypass transcribing RNAPs within ~1 to 2 s of an encounter at rRNA genes and within ~10 s at proteincoding genes. Thus, while individual RNAPs have little effect on the progress of loop extrusion, long, highly transcribed operons can significantly impede the extrusion process. Our data and quantitative models further suggest that bacterial condensin loop extrusion occurs by 2 independent, uncoupled motor activities; the motors translocate on DNA in opposing directions and function together to enlarge chromosomal loops, each independently bypassing steric barriers in their path. Our study provides a quantitative link between transcription and 3D genome organization and proposes a mechanism of interactions between SMC complexes and elongating transcription machinery relevant from bacteria to higher eukaryotes. [ABSTRACT FROM AUTHOR]

Mammalian chromatin is spatially organized at many scales showing two prominent features in interphase: (i) alternating regions (1-10 Mb) of active and inactive chromatin that spatially segregate into different compartments, and (ii) domains (<1 Mb), that is, regions that preferentially interact internally [topologically associating domains (TADs)] and are central to gene regulation. There is growing evidence that TADs are formed by active extrusion of chromatin loops by cohesin, whereas compartmentalization is established according to local chromatin states. Here, we use polymer simulations to examine how loop extrusion and compartmental segregation work collectively and potentially interfere in shaping global chromosome organization. A model with differential attraction between euchromatin and heterochromatin leads to phase separation and reproduces compartmentalization as observed in Hi-C. Loop extrusion, essential for TAD formation, in turn, interferes with compartmentalization. Our integrated model faithfully reproduces Hi-C data from puzzling experimental observations where altering loop extrusion also led to changes in compartmentalization. Specifically, depletion of chromatin-associated cohesin reduced TADs and revealed finer compartments, while increased processivity of cohesin strengthened large TADs and reduced compartmentalization; and depletion of the TAD boundary protein CTCF weakened TADs while leaving compartments unaffected. We reveal that these experimental perturbations are special cases of a general polymer phenomenon of active mixing by loop extrusion. Our results suggest that chromatin organization on the megabase scale emerges from competition of nonequilibrium active loop extrusion and epigenetically defined compartment structure. [ABSTRACT FROM AUTHOR]

Fertilization triggers assembly of higher-order chromatin structure from a condensed maternal and a naïve paternal genome to generate a totipotent embryo. Chromatin loops and domains have been detected in mouse zygotes by single-nucleus Hi-C (snHi-C), but not bulk Hi-C. It is therefore unclear when and how embryonic chromatin conformations are assembled. Here, we investigated whether a mechanism of cohesin-dependent loop extrusion generates higher-order chromatin structures within the one-cell embryo. Using snHi-C of mouse knockout embryos, we demonstrate that the zygotic genome folds into loops and domains that critically depend on Scc1-cohesin and that are regulated in size and linear density by Wapl. Remarkably, we discovered distinct effects on maternal and paternal chromatin loop sizes, likely reflecting differences in loop extrusion dynamics and epigenetic reprogramming. Dynamic polymer models of chromosomes reproduce changes in snHi-C, suggesting a mechanism where cohesin locally compacts chromatin by active loop extrusion, whose processivity is controlled by Wapl. Our simulations and experimental data provide evidence that cohesin-dependent loop extrusion organizes mammalian genomes over multiple scales from the one-cell embryo onward. [ABSTRACT FROM AUTHOR]

Predicting and constraining RNA virus evolution require understanding the molecular factors that define the mutational landscape accessible to these pathogens. RNA viruses typically have high mutation rates, resulting in frequent production of protein variants with compromised biophysical properties. Their evolution is necessarily constrained by the consequent challenge to protein folding and function. We hypothesized that host proteostasis mechanisms may be significant determinants of the fitness of viral protein variants, serving as a critical force shaping viral evolution. Here, we test that hypothesis by propagating influenza in host cells displaying chemically-controlled, divergent proteostasis environments. We find that both the nature of selection on the influenza genome and the accessibility of specific mutational trajectories are significantly impacted by host proteostasis. These findings provide new insights into features of host-pathogen interactions that shape viral evolution, and into the potential design of host proteostasis-targeted antiviral therapeutics that are refractory to resistance. [ABSTRACT FROM AUTHOR]

Summary The cell fate decision between interferon-producing plasmacytoid DC (pDC) and antigen-presenting classical DC (cDC) is controlled by the E protein transcription factor TCF4 (E2-2). We report that TCF4 comprises two transcriptional isoforms, both of which are required for optimal pDC development in vitro. The long Tcf4 isoform is expressed specifically in pDCs, and its deletion in mice impaired pDCs development and led to the expansion of non-canonical CD8 + cDCs. The expression of Tcf4 commenced in progenitors and was further upregulated in pDCs, correlating with stage-specific activity of multiple enhancer elements. A conserved enhancer downstream of Tcf4 was required for its upregulation during pDC differentiation, revealing a positive feedback loop. The expression of Tcf4 and the resulting pDC differentiation were selectively sensitive to the inhibition of enhancer-binding BET protein activity. Thus, lineage-specifying function of E proteins is facilitated by lineage-specific isoform expression and by BET-dependent feedback regulation through distal regulatory elements. [ABSTRACT FROM AUTHOR]

Summary Hematopoietic stem cells (HSCs) sustain long-term reconstitution of hematopoiesis in transplantation recipients, yet their role in the endogenous steady-state hematopoiesis remains unclear. In particular, recent studies suggested that HSCs provide a relatively minor contribution to immune cell development in adults. We directed transgene expression in a fraction of HSCs that maintained reconstituting activity during serial transplantations. Inducible genetic labeling showed that transgene-expressing HSCs gave rise to other phenotypic HSCs, confirming their top position in the differentiation hierarchy. The labeled HSCs rapidly contributed to committed progenitors of all lineages and to mature myeloid cells and lymphocytes, but not to B-1a cells or tissue macrophages. Importantly, labeled HSCs gave rise to more than two-thirds of all myeloid cells and platelets in adult mice, and this contribution could be accelerated by an induced interferon response. Thus, classically defined HSCs maintain immune cell development in the steady state and during systemic cytokine responses. [ABSTRACT FROM AUTHOR]

The mechanism by which chromatids and chromosomes are segregated during mitosis and meiosis is a major puzzle of biology and biophysics. Using polymer simulations of chromosome dynamics, we show that a single mechanism of loop extrusion by condensins can robustly compact, segregate and disentangle chromosomes, arriving at individualized chromatids with morphology observed in vivo. Our model resolves the paradox of topological simplification concomitant with chromosome 'condensation', and explains how enzymes a few nanometers in size are able to control chromosome geometry and topology at micron length scales. We suggest that loop extrusion is a universal mechanism of genome folding that mediates functional interactions during interphase and compacts chromosomes during mitosis. [ABSTRACT FROM AUTHOR]

Three-dimensional (3D) organization of genomes affects critical cellular processes such as transcription, replication, and deoxyribo nucleic acid (DNA) repair. While previous studies have investigated the natural role, the 3D organization plays in limiting a possible set of genomic rearrangements following DNA repair, the influence of specific organizational principles on this process, particularly over longer evolutionary time scales, remains relatively unexplored. In budding yeast S.cerevisiae, chromosomes are organized into a Rabl-like configuration, with clustered centromeres and telomeres tethered to the nuclear periphery. Hi-C data for S.cerevisiae show that a consequence of this Rabl-like organization is that regions equally distant from centromeres are more frequently in contact with each other, between arms of both the same and different chromosomes. Here, we detect rearrangement events in Saccharomyces species using an automatic approach, and observe increased rearrangement frequency between regions with higher contact frequencies. Together, our results underscore how specific principles of 3D chromosomal organization can influence evolutionary events. [ABSTRACT FROM AUTHOR]

Summary Mammalian interphase chromosomes interact with the nuclear lamina (NL) through hundreds of large lamina-associated domains (LADs). We report a method to map NL contacts genome-wide in single human cells. Analysis of nearly 400 maps reveals a core architecture consisting of gene-poor LADs that contact the NL with high cell-to-cell consistency, interspersed by LADs with more variable NL interactions. The variable contacts tend to be cell-type specific and are more sensitive to changes in genome ploidy than the consistent contacts. Single-cell maps indicate that NL contacts involve multivalent interactions over hundreds of kilobases. Moreover, we observe extensive intra-chromosomal coordination of NL contacts, even over tens of megabases. Such coordinated loci exhibit preferential interactions as detected by Hi-C. Finally, the consistency of NL contacts is inversely linked to gene activity in single cells and correlates positively with the heterochromatic histone modification H3K9me3. These results highlight fundamental principles of single-cell chromatin organization. Video Abstract [ABSTRACT FROM AUTHOR]

Eukaryotic genomes are folded into three-dimensional structures, such as self-associating topological domains, the borders of which are enriched in cohesin and CCCTC-binding factor (CTCF) required for long-range interactions. How local chromatin interactions govern higher-order folding of chromatin fibres and the function of cohesin in this process remain poorly understood. Here we perform genome-wide chromatin conformation capture (Hi-C) analysis to explore the high-resolution organization of the Schizosaccharomyces pombe genome, which despite its small size exhibits fundamental features found in other eukaryotes. Our analyses of wild-type and mutant strains reveal key elements of chromosome architecture and genome organization. On chromosome arms, small regions of chromatin locally interact to form 'globules'. This feature requires a function of cohesin distinct from its role in sister chromatid cohesion. Cohesin is enriched at globule boundaries and its loss causes disruption of local globule structures and global chromosome territories. By contrast, heterochromatin, which loads cohesin at specific sites including pericentromeric and subtelomeric domains, is dispensable for globule formation but nevertheless affects genome organization. We show that heterochromatin mediates chromatin fibre compaction at centromeres and promotes prominent inter-arm interactions within centromere-proximal regions, providing structural constraints crucial for proper genome organization. Loss of heterochromatin relaxes constraints on chromosomes, causing an increase in intra- and inter-chromosomal interactions. Together, our analyses uncover fundamental genome folding principles that drive higher-order chromosome organization crucial for coordinating nuclear functions. [ABSTRACT FROM AUTHOR]

The classic model of eukaryotic gene expression requires direct spatial contact between a distal enhancer and a proximal promoter. Recent Chromosome Conformation Capture (3C) studies show that enhancers and promoters are embedded in a complex network of looping interactions. Here we use a polymer model of chromatin fiber to investigate whether, and to what extent, looping interactions between elements in the vicinity of an enhancer-promoter pair can influence their contact frequency. Our equilibrium polymer simulations show that a chromatin loop, formed by elements flanking either an enhancer or a promoter, suppresses enhancer-promoter interactions, working as an insulator. A loop formed by elements located in the region between an enhancer and a promoter, on the contrary, facilitates their interactions. We find that different mechanisms underlie insulation and facilitation; insulation occurs due to steric exclusion by the loop, and is a global effect, while facilitation occurs due to an effective shortening of the enhancer-promoter genomic distance, and is a local effect. Consistently, we find that these effects manifest quite differently for in silico 3C and microscopy. Our results show that looping interactions that do not directly involve an enhancer-promoter pair can nevertheless significantly modulate their interactions. This phenomenon is analogous to allosteric regulation in proteins, where a conformational change triggered by binding of a regulatory molecule to one site affects the state of another site. [ABSTRACT FROM AUTHOR]

miR-10b is silenced in normal neuroglial cells of the brain but commonly activated in glioma, where it assumes an essential tumor-promoting role. We demonstrate that the entire miR-10b-hosting HOXD locus is activated in glioma via the cis -acting mechanism involving 3D chromatin reorganization and CTCF-cohesin-mediated looping. This mechanism requires two interacting lncRNAs, HOXD-AS2 and LINC01116, one associated with HOXD3/HOXD4/miR-10b promoter and another with the remote enhancer. Knockdown of either lncRNA in glioma cells alters CTCF and cohesin binding, abolishes chromatin looping, inhibits the expression of all genes within HOXD locus, and leads to glioma cell death. Conversely, in cortical astrocytes, enhancer activation is sufficient for HOXD/miR-10b locus reorganization, gene derepression, and neoplastic cell transformation. LINC01116 RNA is essential for this process. Our results demonstrate the interplay of two lncRNAs in the chromatin folding and concordant regulation of miR-10b and multiple HOXD genes normally silenced in astrocytes and triggering the neoplastic glial transformation. [Display omitted] • Chromatin topology of the HOXD locus is altered between the brain and glioblastoma • lncRNAs regulate CTCF/cohesin-dependent loop formation and HOXD gene expression • Activation of LINC01116 enhancer RNA leads to astrocyte transformation • The transformation depends on the loop formation and miR-10b derepression Deforzh et al. investigated a common mechanism of HOXD/miR-10b genes' derepression in glioblastoma and revealed the coordinated activity of two lncRNAs, HOXD-embedded HOXD-AS2 and distant enhancer-associated LINC01116, in CTCF/cohesin binding, chromatin topology, and astrocyte transformation. The work shed light on the molecular mechanisms of gliomagenesis. [ABSTRACT FROM AUTHOR]

Job Dekker1 Mitotic chromosomes are among the most recognizable structures in the cell, yet for over a century their internal organization remains largely unsolved. We applied chromosome conformation capture methods, 5C and Hi-C, across the cell cycle and revealed two distinct three-dimensional folding states of the human genome. We show that the highly compartmentalized and cell type-specific organization described previously for nonsynchronous cells is restricted to interphase. In metaphase, we identified a homogenous folding state that is locus-independent, common to all chromosomes, and consistent among cell types, suggesting a general principle of metaphase chromosome organization. Using polymer simulations, we found that metaphase Hi-C data are inconsistent with classic hierarchical models and are instead best described by a linearly organized longitudinally compressed array of consecutive chromatin loops. [ABSTRACT FROM AUTHOR]

The article discusses the issues related to the effect of chromosomal make-up on the cell differentiation. It offers information on the utility of Hi-C method in providing a comprehensive whole-genome picture of physical contacts between distal loci. It also emphasizes the need of consistency in Hi-C data analysis and removal of experimental biases for chromosomal architecture study between cell types.

Cancer progression is driven by the accumulation of a small number of genetic alterations. However, these few driver alterations reside in a cancer genome alongside tens of thousands of additional mutations termed passengers. Passengers are widely believed to have no role in cancer, yet many passengers fall within protein-coding genes and other functional elements that can have potentially deleterious effects on cancer cells. Here we investigate the potential of moderately deleterious passengers to accumulate and alter the course of neoplastic progression. Our approach combines evolutionary simulations of cancer progression with an analysis of cancer sequencing data. From simulations, we find that passengers accumulate and largely evade natural selection during progression. Although individually weak, the collective burden of passengers alters the course of progression, leading to several oncological phenomena that are hard to explain with a traditional driver-centric view. We then tested the predictions of our model using cancer genomics data and confirmed that many passengers are likely damaging and have largely evaded negative selection. Finally, we use our model to explore cancer treatments that exploit the load of passengers by either (i) increasing the mutation rate or (ii) exacerbating their deleterious effects. Though both approaches lead to cancer regression, the latter is a more effective therapy. Our results suggest a unique framework for understanding cancer progression as a balance of driver and passenger mutations. [ABSTRACT FROM AUTHOR]

Proper timing of gene expression requires that transcription factors (TFs) efficiently locate and bind their target sites within a genome. Theoretical studies have long proposed that one-dimensional sliding along DNA while simultaneously reading its sequence can accelerate TF's location of target sites. Sliding by prokaryotic and eukaryotic TFs were subsequently observed. More recent theoretical investigations have argued that simultaneous reading and sliding is not possible for TFs without their possessing at least two DNA-binding modes. The tumor suppressor p53 has been shown to slide on DNA, and recent experiments have offered structural and single molecule support for a two-mode model for the protein. If the model is applicable to p53, then the requirement that TFs be able to read while sliding implies that noncognate sites will affect p53's mobility on DNA, which will thus be generally sequence-dependent. Here, we confirm this prediction with single-molecule microscopy measurements of p53's local diffusivity on noncognate DNA.We show how a two-mode model accurately predicts the variation in local diffusivity, while a single-mode model does not. We further determine that the best model of sequence-specific binding energy includes terms for "hemi-specific" binding, with one dimer of tetrameric p53 binding specifically to a half-site and the other binding nonspecifically to noncognate DNA. Our work provides evidence that the recognition by p53 of its targets and the timing thereof can depend on its noncognate binding properties and its ability to change between multiple modes of binding, in addition to the much better-studied effects of cognate-site binding. [ABSTRACT FROM AUTHOR]

Extracting biologically meaningful information from chromosomal interactions obtained with genome-wide chromosome conformation capture (3C) analyses requires the elimination of systematic biases. We present a computational pipeline that integrates a strategy to map sequencing reads with a data-driven method for iterative correction of biases, yielding genome-wide maps of relative contact probabilities. We validate this ICE (iterative correction and eigenvector decomposition) technique on published data obtained by the high-throughput 3C method Hi-C, and we demonstrate that eigenvector decomposition of the obtained maps provides insights into local chromatin states, global patterns of chromosomal interactions, and the conserved organization of human and mouse chromosomes. [ABSTRACT FROM AUTHOR]

The accumulation of data on structural variation in cancer genomes provides an opportunity to better understand the mechanisms of genomic alterations and the forces of selection that act upon these alterations in cancer. Here we test evidence supporting the influence of two major forces, spatial chromosome structure and purifying (or negative) selection, on the landscape of somatic copy-number alterations (SCNAs) in cancer. Using a maximum likelihood approach, we compare SCNA maps and three-dimensional genome architecture as determined by genome-wide chromosome conformation capture (HiC) and described by the proposed fractal-globule model. This analysis suggests that the distribution of chromosomal alterations in cancer is spatially related to three-dimensional genomic architecture and that purifying selection, as well as positive selection, influences SCNAs during somatic evolution of cancer cells. [ABSTRACT FROM AUTHOR]