Chromatin is reprogrammed after fertilization to produce a totipotent zygote with the potential to generate a new organism. The maternal genome inherited from the oocyte and the paternal genome ...provided by sperm coexist as separate haploid nuclei in the zygote. How these two epigenetically distinct genomes are spatially organized is poorly understood. Existing chromosome conformation capture-based methods are not applicable to oocytes and zygotes owing to a paucity of material. To study three-dimensional chromatin organization in rare cell types, we developed a single-nucleus Hi-C (high-resolution chromosome conformation capture) protocol that provides greater than tenfold more contacts per cell than the previous method. Here we show that chromatin architecture is uniquely reorganized during the oocyte-to-zygote transition in mice and is distinct in paternal and maternal nuclei within single-cell zygotes. Features of genomic organization including compartments, topologically associating domains (TADs) and loops are present in individual oocytes when averaged over the genome, but the presence of each feature at a locus varies between cells. At the sub-megabase level, we observed stochastic clusters of contacts that can occur across TAD boundaries but average into TADs. Notably, we found that TADs and loops, but not compartments, are present in zygotic maternal chromatin, suggesting that these are generated by different mechanisms. Our results demonstrate that the global chromatin organization of zygote nuclei is fundamentally different from that of other interphase cells. An understanding of this zygotic chromatin 'ground state' could potentially provide insights into reprogramming cells to a state of totipotency.
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IJS, KISLJ, NUK, SBMB, UL, UM, UPUK
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
TAD in mouse embryonic stem cells using super-resolution live-cell imaging and quantified looping dynamics by Bayesian inference. Unexpectedly, the
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
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.
Several general principles of global 3D genome organization have recently been established, including non-random positioning of chromosomes and genes in the cell nucleus, distinct chromatin ...compartments, and topologically associating domains (TADs). However, the extent and nature of cell-to-cell and cell-intrinsic variability in genome architecture are still poorly characterized. Here, we systematically probe heterogeneity in genome organization. High-throughput optical mapping of several hundred intra-chromosomal interactions in individual human fibroblasts demonstrates low association frequencies, which are determined by genomic distance, higher-order chromatin architecture, and chromatin environment. The structure of TADs is variable between individual cells, and inter-TAD associations are common. Furthermore, single-cell analysis reveals independent behavior of individual alleles in single nuclei. Our observations reveal extensive variability and heterogeneity in genome organization at the level of individual alleles and demonstrate the coexistence of a broad spectrum of genome configurations in a cell population.
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•Genome organization is highly variable between individual cells•Physical interaction between any two genome regions is a relatively rare event•The structure of TADs is malleable and variable between individual cells•The interactions of two alleles in the same nucleus are independent of each other
High-throughput imaging of several hundred chromatin interactions in individual cells reveals extensive heterogenity in spatial genome organization at the single-cell level.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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.
Synopsis
Zygotic genomes fold into loops and domains that depend on cohesin and are regulated in size by Wapl. Single‐nucleus Hi‐C and polymer models of chromosomes provide evidence for cohesin‐dependent loop extrusion organizing genomes over multiple scales.
Zygotic genomes are organized into cohesin‐dependent chromatin loops and Topologically Associating Domains (TADs).
Loop extrusion leads to different loop strengths in maternal and paternal genomes.
Cohesin restricts inter‐chromosomal interactions by altering chromosome surface area.
Loop extrusion organizes chromatin at multiple genomic scales.
Single‐nucleus Hi‐C and microscopy show that the zygotic genome folds into loops and domains in a cohesin‐dependent manner with differences in maternal and paternal chromosome organization.
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FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Structural maintenance of chromosomes (SMC) complexes play critical roles in chromosome dynamics in virtually all organisms, but how they function remains poorly understood. In the bacterium Bacillus ...subtilis, SMC-condensin complexes are topologically loaded at centromeric sites adjacent to the replication origin. Here we provide evidence that these ring-shaped assemblies tether the left and right chromosome arms together while traveling from the origin to the terminus (>2 megabases) at rates >50 kilobases per minute. Condensin movement scales linearly with time, providing evidence for an active transport mechanism. These data support a model in which SMC complexes function by processively enlarging DNA loops. Loop formation followed by processive enlargement provides a mechanism by which condensin complexes compact and resolve sister chromatids in mitosis and by which cohesin generates topologically associating domains during interphase.
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BFBNIB, NMLJ, NUK, ODKLJ, PNG, SAZU, UL, UM, UPUK
Mammalian genomes are organized and regulated through long-range chromatin interactions. Structural loops formed by CCCTC-binding factor (CTCF) and cohesin fold the genome into domains, while ...enhancers interact with promoters across vast genomic distances to regulate gene expression. Although genomics and fixed-cell imaging approaches help illuminate many aspects of chromatin interactions, temporal information is usually lost. Here, we discuss how 3D super-resolution live-cell imaging (SRLCI) can resolve open questions on the dynamic formation and dissolution of chromatin interactions. We discuss SRLCI experimental design, implementation strategies, and data interpretation and highlight associated pitfalls. We conclude that, while technically demanding, SRLCI approaches will likely emerge as a critical tool to dynamically probe 3D genome structure and function and to study enhancer–promoter interactions and chromatin looping.
•Super-resolution live-cell imaging (SRLCI) can track chromatin interactions.•SRLCI can help distinguish alternative models of enhancer-promoter interaction.•Discussion of challenges in SRLCI experimental design and data interpretation.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Expansins refer to a family of closely related non-enzymatic proteins found in the plant cell wall that are involved in the cell wall loosening. In addition, expansins appear to be involved in ...different physiological and environmental responses in plants such as leaf and stem initiation and growth, stomata opening and closing, reproduction, ripening and stress tolerance. Sugarcane (Saccharum spp.) is one of the main crops grown worldwide. Lignocellulosic biomass from sugarcane is one of the most promising raw materials for the ethanol industry. However, the efficient use of lignocellulosic biomass requires the optimization of several steps, including the access of some enzymes to the hemicellulosic matrix. The addition of expansins in an enzymatic cocktail or their genetic manipulation could drastically improve the saccharification process of feedstock biomass by weakening the hydrogen bonds between polysaccharides present in plant cell walls. In this study, the expansin gene family in sugarcane was identified and characterized by in silico analysis. Ninety two putative expansins in sugarcane (SacEXPs) were categorized in three subfamilies after phylogenetic analysis. The expression profile of some expansin genes in leaves of sugarcane in different developmental stages was also investigated. This study intended to provide suitable expansin targets for genetic manipulation of sugarcane aiming at biomass and yield improvement.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Fertilization is defined as the union of two gametes. During fertilization, sperm and egg fuse to form a diploid zygote to initiate prenatal development. In mammals, fertilization involves multiple ...ordered steps, including the acrosome reaction, zona pellucida penetration, sperm-egg attachment, and membrane fusion. Given the success of in vitro fertilization, one would think that the mechanisms of fertilization are understood; however, the precise details for many of the steps in fertilization remain a mystery. Recent studies using genetic knockout mouse models and structural biology are providing valuable insight into the molecular basis of sperm-egg attachment and fusion. Here, we review the cell biology of fertilization, specifically summarizing data from recent structural and functional studies that provide insights into the interactions involved in human gamete attachment and fusion.
The new generation of radio interferometers is characterized by high sensitivity, wide fields of view and large fractional bandwidth. To synthesize the deepest images enabled by the high dynamic ...range of these instruments requires us to take into account the direction-dependent Jones matrices, while estimating the spectral properties of the sky in the imaging and deconvolution algorithms. In this paper we discuss and implement a wideband wide-field spectral deconvolution framework (ddfacet) based on image plane faceting, that takes into account generic direction-dependent effects. Specifically, we present a wide-field co-planar faceting scheme, and discuss the various effects that need to be taken into account to solve for the deconvolution problem (image plane normalization, position-dependent Point Spread Function, etc). We discuss two wideband spectral deconvolution algorithms based on hybrid matching pursuit and sub-space optimisation respectively. A few interesting technical features incorporated in our imager are discussed, including baseline dependent averaging, which has the effect of improving computing efficiency. The version of ddfacet presented here can account for any externally defined Jones matrices and/or beam patterns.
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FMFMET, NUK, UL, UM, UPUK
Chromosome organization mediated by structural maintenance of chromosomes (SMC) complexes is vital in many organisms. SMC complexes act as motors that extrude DNA loops, but it remains unclear what ...happens when multiple complexes encounter one another on the same DNA in living cells and how these interactions may help to organize an active genome. We therefore created a crash-course track system to study SMC complex encounters in vivo by engineering defined SMC loading sites in the Bacillus subtilis chromosome. Chromosome conformation capture (Hi-C) analyses of over 20 engineered strains show an amazing variety of chromosome folding patterns. Through three-dimensional polymer simulations and theory, we determine that these patterns require SMC complexes to bypass each other in vivo, as recently seen in an in vitro study. We posit that the bypassing activity enables SMC complexes to avoid traffic jams while spatially organizing the genome.
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GEOZS, IJS, IMTLJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBMB, UL, UM, UPUK, ZAGLJ