Cohesin depleted cells rebuild functional nuclear compartments after endomitosis

Cohesin plays an essential role in chromatin loop extrusion, but its impact on a compartmentalized nuclear architecture, linked to nuclear functions, is less well understood. Using live-cell and super-resolved 3D microscopy, here we find that cohesin depletion in a human colon cancer derived cell line results in endomitosis and a single multilobulated nucleus with chromosome territories pervaded by interchromatin channels. Chromosome territories contain chromatin domain clusters with a zonal organization of repressed chromatin domains in the interior and transcriptionally competent domains located at the periphery. These clusters form microscopically defined, active and inactive compartments, which likely correspond to A/B compartments, which are detected with ensemble Hi-C. Splicing speckles are observed nearby within the lining channel system. We further observe that the multilobulated nuclei, despite continuous absence of cohesin, pass through S-phase with typical spatio-temporal patterns of replication domains. Evidence for structural changes of these domains compared to controls suggests that cohesin is required for their full integrity

Cohesin depleted cells pass through mitosis and reconstitute a functional nuclear architecture

The human genome forms thousands of “contact domains”, which are intervals of enhanced contact frequency. Some, called “loop domains” are thought to form by cohesin-mediated loop extrusion. Others, called “compartmental domains”, form due to the segregation of active and inactive chromatin into A and B compartments. Recently, Hi-C studies revealed that the depletion of cohesin leads to the disappearance of all loop domains within a few hours, but strengthens compartment structure. Here, we combine live cell microscopy, super-resolution microscopy, Hi-C, and studies of replication timing to examine the longer-term consequences of cohesin degradation in HCT-116 human colorectal carcinoma cells, tracking cells for up to 30 hours. Surprisingly, cohesin depleted cells proceed through an aberrant mitosis, yielding a single postmitotic cell with a multilobulated nucleus. Hi-C reveals the continued disappearance of loop domains, whereas A and B compartments are maintained. In line with Hi-C, microscopic observations demonstrate the reconstitution of chromosome territories and chromatin domains. An interchromatin channel system (IC) expands between chromatin domain clusters and carries splicing speckles. The IC is lined by active chromatin enriched for RNA Pol II and depleted in H3K27me3. Moreover, the cells exhibit typical early-, mid-, and late- DNA replication timing patterns. Our observations indicate that the functional nuclear compartmentalization can be maintained in cohesin depleted pre- and postmitotic cells. However, we find that replication foci – sites of active DNA synthesis – become physically larger consistent with a model where cohesin dependent loop extrusion tends to compact intervals of replicating chromatin, whereas their genomic boundaries are associated with compartmentalization, and do not change.3D FISH3D fluorescence in situ hybridization3D SIM3D structured illumination microscopyAIDauxin inducible degronANC / INCactive / inactive nuclear compartmentCTchromosome territoryCD(C)chromatin domain (cluster)CTCFCCCTC binding factorDAPI4’,6-diamidino-2-phenylindoleEdU5-Ethynyl-2’-deoxyuridineHi-Cchromosome conformation capturing combined with deep sequencingICinterchromatin compartmentMLNmultilobulated nucleusNCnucleosome clusterPBSphosphate buffered salinePBSTphosphate buffered saline with 0.02% TweenPRperichromatin regionRDreplication domainRLreplication labelingTADtopologically associating domai

Are the processes of DNA replication and DNA repair reading a common structural chromatin unit?

Decades of investigation on genomic DNA have brought us deeper insights into its organization within the nucleus and its metabolic mechanisms. This was fueled by the parallel development of experimental techniques and has stimulated model building to simulate genome conformation in agreement with the experimental data. Here, we will discuss our recent discoveries on the chromatin units of DNA replication and DNA damage response. We will highlight their remarkable structural similarities and how both revealed themselves as clusters of nanofocal structures each on the hundred thousand base pair size range corresponding well with chromatin loop sizes. We propose that the function of these two global genomic processes is determined by the loop level organization of chromatin structure with structure dictating function. 3D-SIM: 3D-structured illumination microscopy; 3C: chromosome conformation capture; DDR: DNA damage response; FISH: fluorescent in situ hybridization; Hi-C: high conformation capture; HiP-HoP: highly predictive heteromorphic polymer model; IOD: inter-origin distance; LAD: lamina associated domain; STED: stimulated emission depletion microscopy; STORM: stochastic optical reconstruction microscopy; SBS: strings and binders switch model; TAD: topologically associated domain

Tolerancia Crítica y Ciudadanía Activa: Una Introducción Práctica al Debate Educativo

Critical Tolerance and Active Citizenship: A Practical Introduction to Educational Debate, Idebate Press\u27s first release in Spanish, offers educators a hands-on guide for implementing debate practices in classrooms and non-formal educational settings. The work consists of a series of 15 sessions devoted to specific aspects of debating. Topics include: general characteristics of debate formats, public speaking, creating debate resolutions, argumentation, reasoning and fallacies, preparation of cases, taking notes, formulating questions, and judging debates. Each session includes tips for facilitating introductory debate training and exercises to put learning into practice. The work includes activity and resource sheets for students as well as an appendix with a sample debate.https://digitalcommons.linfield.edu/linfauth/1004/thumbnail.jp

Oceanic Plateau and island arcs of southwestern Ecuador: their place in the geodynamic evolution of northwestern South America.

Coastal Ecuador is made up of an oceanic igneous basement overlain by Upper Cretaceous to Lower Paleocene (approximately 98-60 Ma) volcaniclastic and volcanic rocks of island-arc affinities. The igneous basement, known as the Pinon Formation, locally dated at 123 Ma, consists of olivine-free basalts and dolerites. Relative to N-MORB, both types of rocks exhibit high concentrations in Nb (0.3-10.75 ppm), Ta (0.03-0.67 ppm), Th (0.11-1.44 ppm), light and medium rare earth elements, and low Zr (22-105 ppm) and Hf (0.59-2.8 ppm) contents, thus showing oceanic plateau basalts affinities. Most of these oceanic plateau basalts tholeiites display rather homogeneous epsilonNd(T = 123 Ma) ratios (approximately +7), with the exception of two rocks with higher (+10) and lower (+4.5) epsilonNd(T = 123 Ma), respectively. All these basalts plot, with one exception, within the ocean island basalts field. Their (87Sr/86Sr)i ratios are highly variable (0.7032-0.7048), probably due to hydrothermal oceanic alteration or assimilation of altered oceanic crust. The rocks of the Pinon Formation are geochemically similar to the oceanic plateau tholeiites from Nauru and Ontong Java Plateaus and to the Upper Cretaceous (92-88 Ma) Carribbean Oceanic Plateau lavas. The basalts and dolerites of the Upper Cretaceous-Lower Paleocene island arcs show calc-alkaline affinities. The epsilonNd ratios to Cretaceous intra-oceanic are rocks from North, Central and South America and from the Greater Antilles. Since the Pinon oceanic plateau tholeiites are locally overlain by early-Late Cretaceous sediments (approximately 98-83 Ma) and yielded locally an Early Cretaceous age, they do not belong to the Late Cretaceous Caribbean Oceanic Plateau. The basement of coastal Ecuador is interpreted as an accreted fragment of an overthickened and buoyant oceanic plateau... (D'après résumé d'auteur

Géodynamique andine : résumés étendus = Andean geodynamics : extended abstracts = Geodinamica andina : resumenes expandidos

Des plateaux océaniques se sont accrétés à différentes périodes à la marge équatorienne. La suture d'âge crétacé supérieur contient des roches cumulatives mafiques et ultamafiques datées à 123 Ma (San Jaun). Plus à l'ouest, la suture paléocène contient des picrites et basaltes riches en Mg qui diffèrent des précédentes. Elles ont en particulier des _Nd plus élevés et sont beaucoup plus radiogéniques en Pb que celles de San Juan. Elles présentent de fortes similitudes avec les roches crétacé supérieur du Plateau Caraïbe et pourraient représenter des fragments de ce dernier, accrétés en Equateur vers 55 Ma. (Résumé d'auteur

The Early Cretaceous San Juan Plutonic Suite, Ecuador: A magma chamber in an oceanic plateau ?

Sections through an oceanic plateau are preserved in tectonic slices in the Western Cordillera of Ecuador (South America). The San Juan section is a sequence of mafic-ultramafic cumulates. To establish that these plutonic rocks formed in an oceanic plateau setting, we have developed criteria that discriminate intrusions of oceanic plateaus from those of other tectonic settings. The mineralogy and crystallization sequence of the cumulates are similar to those of intra-plate magmas. Clinopyroxene predominates throughout, and orthopyroxene is only a minor component. Rocks of intermediate composition are absent, and hornblende is restricted to the uppermost massive gabbros within the sequence. The ultramafic cumulates are very depleted in light rare-earth elements (LREE), whereas the gabbros have flat or slightly enriched LREE patterns. The composition of the basaltic liquid in equilibrium with the peridotite, calculated using olivine compositions and REE contents of clinopyroxene, contains between 16% and 8% MgO and has a flat REE pattern. This melt is geochemically similar to other accreted oceanic plateau basalts, isotropic gabbros, and differentiated sills in western Ecuador. The Ecuadorian intrusive and extrusive rocks have a narrow range of epsilonNd(i) (+8 to +5) and have a rather large range of Pb isotopic ratios. Pb isotope systematics of the San Juan plutonic rocks and mineral separates lie along a mixing line between the depleted mantle (DMM) and the enriched-plume end members. This suggests that the Ecuadorian plutonic rocks generated from the mixing of two mantle sources, a depleted mid-oceanic ridge basalt (MORB) source and an enriched one. The latter is characterized by high (Pb-207/Pb-204)(i) ratios and could reflect a contamination by recycled either lower continental crust or oceanic pelagic sediments and (or) altered oceanic crust (enriched mantle type I, EMI). These data suggest that the San Juan sequence represents the plutonic components of an Early Cretaceous oceanic plateau, which accreted in the Late Cretaceous to the Ecuadorian margin

The Chromatin Architectural Protein CTCF Is Critical for Cell Survival upon Irradiation-Induced DNA Damage

CTCF is a nuclear protein initially discovered for its role in enhancer-promoter insulation. It has been shown to play a role in genome architecture and in fact its DNA binding sites are enriched at the borders of chromatin domains. Recently, we showed that depletion of CTCF impairs the DNA damage response to ionizing radiation. To investigate the relationship between chromatin domains and DNA damage repair, we present here clonogenic survival assays in different cell lines upon CTCF knockdown and ionizing irradiation. The application of a wide range of ionizing irradiation doses (0-10 Gy) allowed us to investigate the survival response through a biophysical model that accounts for the double strand breaks probability distribution onto chromatin domains. We demonstrate that the radiosensitivity of different cell lines is increased upon lowering the amount of the architectural protein. Our model shows that the deficiency in DNA repair ability is related to the changes in the size of chromatin domains that occur when different amounts of CTCF are present in the nucleus.Published Versio