Bacterial genomes have been shown to be partitioned into several kilobases long chromosomal domains that are topologically independent from each other, meaning that change of DNA superhelicity in one domain does not propagate to neighbors. Both in vivo and in vitro experiments have been performed to question the nature of the topological barriers at play, leading to several predictions on possible molecular actors. Here, we address the question of topological barriers using polymer models of supercoiled DNA chains. More specifically, we determine under which conditions DNA-bridging proteins may act as topological barriers. To this end, we developed a coarse-grained bead-and-spring model and investigated its properties through Brownian dynamics simulations. As a result, we find that DNA-bridging proteins must exert rather strong constraints on their binding sites: they must block the diffusion of the excess of twist through the two binding sites on the DNA molecule and, simultaneously, prevent the rotation of one DNA segment relative to the other one. Importantly, not all DNA-bridging proteins satisfy this second condition. For example, single bridges formed by proteins that bind DNA non-specifically, like H-NS dimers, are expected to fail with this respect. Our findings might also explain, in the case of specific DNA-bridging proteins like LacI, why multiple bridges are required to create stable independent topological domains. Strikingly, when the relative rotation of the DNA segments is not prevented, relaxation results in complex intrication of the two domains. Moreover, while the value of the torsional stress in each domain may vary, their differential is preserved. Our work also predicts that nucleoid associated proteins known to wrap DNA must form higher protein-DNA complexes to efficiently work as topological barriers.
Under supercoiling constraints, naked DNA, such as a large part of bacterial DNA, folds into braided structures called plectonemes. The double-helix can also undergo local structural transitions, leading to the formation of denaturation bubbles and other alternative structures. Various polymer models have been developed to capture these properties, with Monte-Carlo (MC) approaches dedicated to the inference of thermodynamic properties. In this chapter, we explain how to perform such Monte-Carlo simulations, following two objectives. On one hand, we present the self-avoiding supercoiled Worm-Li…
DNA supercoiling, the under or overwinding of DNA, is a key physical mechanism both participating to compaction of bacterial genomes and making genomic sequences adopt various structural forms. DNA supercoiling may lead to the formation of braided superstructures (plectonemes), or it may locally destabilize canonical B-DNA to generate denaturation bubbles, left-handed Z-DNA and other functional alternative forms. Prediction of the relative fraction of these structures has been limited because of a lack of predictive polymer models that can capture the multiscale properties of long DNA molecule…
Among several quantitative invariants found in evolutionary genomics, one of the most striking is the scaling of the overall abundance of proteins, or protein domains, sharing a specific functional annotation across genomes of given size. The size of these functional categories change, on average, as power-laws in the total number of protein-coding genes. Here, we show that such regularities are not restricted to the overall behavior of high-level functional categories, but also exist systematically at the level of single evolutionary families of protein domains. Specifically, the number of pr…
In-vivo DNA organization at large length scales ($\sim 100nm$) is highly debated and polymer models have proved useful to understand the principle of DNA-organization. Here, we show that $<2$% cross-links at specific points in a ring polymer can lead to a distinct spatial organization of the polymer. The specific pairs of cross-linked monomers were extracted from contact maps of bacterial DNA. We are able to predict the structure of 2 DNAs using Monte Carlo simulations of the bead-spring polymer with cross-links at these special positions. Simulations with cross-links at random positions along…
The volume occupied by unconstrained bacterial DNA in physiological solutions exceeds 1000 times the volume of the cell. Still, it is confined to a well defined region of the cell called the nucleoid, which occupies only a fraction of the cell volume. There is still no general agreement on the mechanism leading to the compaction of the DNA and the formation of the nucleoid. However, advances in in vivo sub-wavelength resolution microscopy techniques have recently allowed the observation of the nucleoid at an unprecedented level of detail. In particular, these observations show that the compact…
The chromosomal DNA of bacteria is folded into a compact body called the nucleoid, which is composed essentially of DNA (80%), RNA (10%), and a number of different proteins (10%). These nucleoid proteins act as regulators of gene expression and influence the organization of the nucleoid by bridging, bending, or wrapping the DNA. These so-called architectural properties of nucleoid proteins are still poorly understood. For example, the reason why certain proteins compact the DNA coil in certain environments but make instead the DNA more rigid in other environments is the matter of ongoing debat…
Since the sequencing of large genomes, many statistical features of their sequences have been found. One intriguing feature is that certain subsequences are much more abundant than others. In fact, abundances of subsequences of a given length are distributed with a scale-free power-law tail, resembling properties of human texts, such as the Zipf's law. Despite recent efforts, the understanding of this phenomenon is still lacking. Here we find that selfish DNA elements, such as those belonging to the Alu family of repeats, dominate the power-law tail. Interestingly, for the Alu elements the pow…
The mobility of protein is fundamental in the machinery of life. Here, we have investigated the effect of DNA binding in conjunction with DNA internal motion of the bacterial Hfq master regulator devoid of its amyloid C-terminus domain. Hfq is one of the most abundant nucleoid associated proteins that shape the bacterial chromosome and is involved in several aspects of nucleic acid metabolism. Fluorescence microscopy has been used to track a C-terminus domain lacking mutant form of Hfq on double stranded DNA, which is stretched by confinement to a rectangular nanofluidic channel. The mobility …
This work addresses the question of the interplay of DNA demixing and supercoiling in bacterial cells. Demixing of DNA from other globular macromolecules results from the overall repulsion between all components of the system and leads to the formation of the nucleoid, which is the region of the cell that contains the genomic DNA in a rather compact form. Supercoiling describes the coiling of the axis of the DNA double helix to accommodate the torsional stress injected in the molecule by topoisomerases. Supercoiling is able to induce some compaction of the bacterial DNA, although to a lesser e…
The unconstrained genomic DNA of bacteria forms a coil, which volume exceeds 1000 times the volume of the cell. Since prokaryotes lack a membrane-bound nucleus, in sharp contrast with eukaryotes, the DNA may consequently be expected to occupy the whole available volume when constrained to fit in the cell. Still, it has been known for more than half a century that the DNA is localized in a well defined region of the cell, called the nucleoid, which occupies only 15% to 25% of the total volume. Although this problem has focused the attention of many scientists for the past decades, there is stil…
The bacterial genome is organized in a structure called the nucleoid by a variety of associated proteins. These proteins can form complexes on DNA that play a central role in various biological processes, including chromosome segregation. A prominent example is the large ParB-DNA complex, which forms an essential component of the segregation machinery in many bacteria. ChIP-Seq experiments show that ParB proteins localize around centromere-like parS sites on the DNA to which ParB binds specifically, and spreads from there over large sections of the chromosome. Recent theoretical and experiment…
The mechanism responsible for the compaction of the genomic DNA of bacteria inside a structure called the nucleoid is a longstanding but still lively debated question. Most puzzling is the fact that the nucleoid occupies only a small fraction of the cell, although it is not separated from the rest of the cytoplasm by any membrane and would occupy a volume about thousand times larger outside from the cell. Here, by performing numerical simulations with coarse-grained models, we elaborate on the conjecture that the formation of the nucleoid may result from a segregative phase separation mechanis…
In vitro binding energies capture Klf4 occupancy across the human genome
Max Planck Institute of Molecular Cell Biology and Genetics（马克斯·普朗克分子细胞生物学与遗传学研究所）；Max Planck Institute for the Physics of Complex Systems（马克斯·普朗克复杂系统物理研究所）；Center for Systems Biology Dresden（德累斯顿系统生物学中心）；Cluster of Excellence Physics of Life, Technical University Dresden（德累斯顿工业大学生命物理卓越集群）
We showed in our previous studies that just $3\%$ cross-links, at special points along the contour of the bacterial DNA help the DNA-polymer to get organized at micron length scales \cite{jpcm,epl}. In this work, we investigate how does the release of topological constraints help in the organization of the DNA-polymer. Furthermore, we show that the chain compaction induced by the crowded environment in the bacterial cytoplasm contributes to the organization of the DNA-polymer. We model the DNA chain as a flexible bead-spring ring polymer, where each bead represents $1000$ base pairs. The speci…
DNA的纳米孔测序是一种单分子技术，其可以以最少的样品制备和仪器实现长读取、低成本和高速。在这里，我们建立在纳米孔分辨率和DNA控制方面的最新进展来解释DNA通过孔MspA易位过程中观察到的离子电流水平的处理。由于大约四个核苷酸影响每个水平的离子电流，我们测量了对应于所有256个四核苷酸组合（四聚体）的离子电流。该四聚体图谱高度预测先前未测量的源自噬菌体phi X 174基因组的序列的离子电流水平。此外，我们显示了长度高达4，500个碱基的phi X 174的纳米孔测序读段，其可以明确地与phi X 174参考基因组对齐，并证明了关于杂交基因组组装和多态性检测的概念验证实用性。所有方法和数据都充分提供。
Zygotic genome activation (ZGA), the onset of transcription after initial quiescence, is a major developmental step in many species, which occurs after ten cell divisions in Zebrafish embryos. How transcription factor-chromatin interactions evolve during early development to support ZGA is largely unknown. We established single molecule tracking in live developing Zebrafish embryos using reflected light-sheet microscopy to visualize the general transcription factor TATA-binding protein (TBP), and developed a novel data acquisition and analysis scheme to extract kinetic information during fast …
In genome mapping experiments, long DNA molecules are stretched by confining them to very narrow channels, so that the locations of sequence-specific fluorescent labels along the channel axis provide large-scale genomic information. It is difficult, however, to make the channels narrow enough so that the DNA molecule is fully stretched. In practice its conformations may form hairpins that change the spacings between internal segments of the DNA molecule, and thus the label locations along the channel axis. Here we describe a theory for the distribution of label spacings that explains the heavy…
Using state-of-the-art techniques combining imaging methods and high-throughput genomic mapping tools leaded to the significant progress in detailing chromosome architecture of various organisms. However, a gap still remains between the rapidly growing structural data on the chromosome folding and the large-scale genome organization. Could a part of information on the chromosome folding be obtained directly from underlying genomic DNA sequences abundantly stored in the databanks? To answer this question, we developed an original discrete double Fourier transform (DDFT). DDFT serves for the det…
Using a bead-spring model of bacterial DNA polymers of {\em C. crescentus} and {\em E. coli} we show that just $33$ and $38$ effective cross-links at special positions along the chain contour of the DNA can lead to the large-scale organization of the DNA polymer, where confinement effects of the cell walls play a key role in the organization. The positions of the $33$ cross-links along the chain contour are chosen from the contact map data of {\em C. crescentus}. We represent $1000$ base pairs as a coarse-grained monomer in our bead-spring flexible ring polymer model of the DNA. Thus a $4017$ …
Hfq is a bacterial protein that is involved in several aspects of nucleic acids metabolism. It has been described as one of the nucleoid associated proteins shaping the bacterial chromosome, although it is better known to influence translation and turnover of cellular RNAs. Here, we explore the role of Escherichia coli Hfq C-terminal domain in the compaction of double stranded DNA. Various experimental methodologies, including fluorescence microscopy imaging of single DNA molecules confined inside nanofluidic channels, atomic force microscopy, isothermal titration microcalorimetry, and electro…
Prokaryotes do not make use of a nucleus membrane to segregate their genetic material from the cytoplasm, so that their nucleoid is potentially free to explore the whole volume of the cell. Nonetheless, high resolution images of bacteria with very compact nucleoids show that such spherical nucleoids are invariably positioned at the center of mononucleoid cells. The present work aims to determine whether such preferential localization results from generic (entropic) interactions between the nucleoid and the cell membrane or instead requires some specific mechanism, like the tethering of DNA at …
Nucleoid clustering drives stepwise expansion and segregation of replicating bacterial chromosomes
SUPA, School of Physics and Astronomy, University of Edinburgh（爱丁堡大学物理与天文学学院）；Dipartimento di Fisica e Astronomia and Sezione INFN, Universitá di Padova（帕多瓦大学物理与天文学系及INFN分部）
Vital biological processes such as genome repair require fast and efficient binding of selected proteins to specific target sites on DNA. Here we propose an active target search mechanism based on "chromophoresis", the dynamics of DNA-binding proteins up or down gradients in the density of epigenetic marks, or colours (biochemical tags on the genome). We focus on a set of proteins that deposit marks from which they are repelled---a case which is only encountered away from thermodynamic equilibrium. For suitable ranges of kinetic parameter values, chromophoretic proteins can perform unidirectio…
Certain retroviruses, including HIV, insert their DNA in a non-random fraction of the host genome via poorly understood selection mechanisms. Here, we develop a biophysical model for retroviral integrations as stochastic and quasi-equilibrium topological reconnections between polymers. We discover that physical effects, such as DNA accessibility and elasticity, play important and universal roles in this process. Our simulations predict that integration is favoured within nucleosomal and flexible DNA, in line with experiments, and that these biases arise due to competing energy barriers associa…
The solution space of genome-scale models of cellular metabolism provides a map between physically viable flux configurations and cellular metabolic phenotypes described, at the most basic level, by the corresponding growth rates. By sampling the solution space of E. coli's metabolic network, we show that empirical growth rate distributions recently obtained in experiments at single-cell resolution can be explained in terms of a trade-off between the higher fitness of fast-growing phenotypes and the higher entropy of slow-growing ones. Based on this, we propose a minimal model for the evolutio…
Impact of Oxygen on DNA Damage Distribution in 3D Genome and its Correlation to Oxygen Enhancement Ratio after High-LET Irradiation
Department of Engineering Physics, Tsinghua University（清华大学工程物理系）；Key Laboratory of Particle & Radiation Imaging, Tsinghua University, Ministry of Education（教育部粒子与辐射成像重点实验室）；Institute of Fluid Physics, China Academy of Engineering Physics（中国工程物理研究院流体物理研究所）
