Nuclear repositioning of the VSG promoter during developmental silencing in Trypanosoma brucei

Interphase nuclear repositioning of chromosomes has been implicated in the epigenetic regulation of RNA polymerase (pol) II transcription. However, little is known about the nuclear position–dependent regulation of RNA pol I–transcribed loci. Trypanosoma brucei is an excellent model system to address this question because its two main surface protein genes, procyclin and variant surface glycoprotein (VSG), are transcribed by pol I and undergo distinct transcriptional activation or downregulation events during developmental differentiation. Although the monoallelically expressed VSG locus is exclusively localized to an extranucleolar body in the bloodstream form, in this study, we report that nonmutually exclusive procyclin genes are located at the nucleolar periphery. Interestingly, ribosomal DNA loci and pol I transcription activity are restricted to similar perinucleolar positions. Upon developmental transcriptional downregulation, however, the active VSG promoter selectively undergoes a rapid and dramatic repositioning to the nuclear envelope. Subsequently, the VSG promoter region was subjected to chromatin condensation. We propose a model whereby the VSG expression site pol I promoter is selectively targeted by temporal nuclear repositioning during developmental silencing

Phase field model of fluid-driven fracture in elastic media: Immersed-fracture formulation and validation with analytical solutions

Propagation of fluid-driven fractures plays an important role in natural and engineering processes, including transport of magma in the lithosphere, geologic sequestration of carbon dioxide, and oil and gas recovery from low-permeability formations, among many others. The simulation of fracture propagation poses a computational challenge as a result of the complex physics of fracture and the need to capture disparate length scales. Phase field models represent fractures as a diffuse interface and enjoy the advantage that fracture nucleation, propagation, branching, or twisting can be simulated without ad hoc computational strategies like remeshing or local enrichment of the solution space. Here we propose a new quasi-static phase field formulation for modeling fluid-driven fracturing in elastic media at small strains. The approach fully couples the fluid flow in the fracture (described via the Reynolds lubrication approximation) and the deformation of the surrounding medium. The flow is solved on a lower dimensionality mesh immersed in the elastic medium. This approach leads to accurate coupling of both physics. We assessed the performance of the model extensively by comparing results for the evolution of fracture length, aperture, and fracture fluid pressure against analytical solutions under different fracture propagation regimes. The excellent performance of the numerical model in all regimes builds confidence in the applicability of phase field approaches to simulate fluid-driven fracture.United States. Department of Energy (Grant DE-SC0009286)Spain. Ministerio de Economía y Competitividad (Grant RyC-2012-11704)Spain. Ministerio de Economía y Competitividad (Grant CTM2014-54312-P

Cohesin regulates VSG monoallelic expression in trypanosomes

Antigenic variation allows Trypanosoma brucei to evade the host immune response by switching the expression of 1 out of ∼15 telomeric variant surface glycoprotein (VSG) expression sites (ESs). VSG ES transcription is mediated by RNA polymerase I in a discrete nuclear site named the ES body (ESB). However, nothing is known about how the monoallelic VSG ES transcriptional state is maintained over generations. In this study, we show that during S and G2 phases and early mitosis, the active VSG ES locus remains associated with the single ESB and exhibits a delay in the separation of sister chromatids relative to control loci. This delay is dependent on the cohesin complex, as partial knockdown of cohesin subunits resulted in premature separation of sister chromatids of the active VSG ES. Cohesin depletion also prompted transcriptional switching from the active to previously inactive VSG ESs. Thus, in addition to maintaining sister chromatid cohesion during mitosis, the cohesin complex plays an essential role in the correct epigenetic inheritance of the active transcriptional VSG ES state

NoMeplot: analysis of DNA methylation and nucleosome occupancy at the single molecule

Supplementary information accompanies this paper at https://doi.org/10.1038/s41598-019-44597-2.We are very grateful to Peter A. Jones for sharing protocols and advice and we thank Serafin Moral for constructive and useful discussion.Recent technical advances highlight that to understand mammalian development and human disease we need to consider transcriptional and epigenetic cell-to-cell differences within cell populations. This is particularly important in key areas of biomedicine like stem cell differentiation and intratumor heterogeneity. The recently developed nucleosome occupancy and methylome (NOMe) assay facilitates the simultaneous study of DNA methylation and nucleosome positioning on the same DNA strand. NOMe-treated DNA can be sequenced by sanger (NOMe-PCR) or high throughput approaches (NOMe-seq). NOMe-PCR provides information for a single locus at the single molecule while NOMe-seq delivers genome-wide data that is usually interrogated to obtain population-averaged measures. Here, we have developed a bioinformatic tool that allow us to easily obtain locus-specific information at the single molecule using genome-wide NOMe-seq datasets obtained from bulk populations. We have used NOMePlot to study mouse embryonic stem cells and found that polycomb-repressed bivalent gene promoters coexist in two different epigenetic states, as defined by the nucleosome binding pattern detected around their transcriptional start site.This study was supported by the Spanish ministry of economy and competitiveness (SAF2013-40891-R; BFU2016-75233-P) and the andalusian regional government (PC-0246-2017). David Landeira is a Ramón y Cajal researcher of the Spanish ministry of economy and competitiveness (RYC-2012- 10019)

Polycomb regulation is coupled to cell cycle transition in pluripotent stem cells

When self-renewing pluripotent cells receive a differentiation signal, ongoing cell duplication needs to be coordinated with entry into a differentiation program. Accordingly, transcriptional activation of lineage specifier genes and cell differentiation is confined to the G1 phase of the cell cycle by unknown mechanisms. We found that Polycomb repressive complex 2 (PRC2) subunits are differentially recruited to lineage specifier gene promoters across cell cycle in mouse embryonic stem cells (mESCs). Jarid2 and the catalytic subunit Ezh2 are markedly accumulated at target promoters during S and G2 phases, while the transcriptionally activating subunits EPOP and EloB are enriched during G1 phase. Fluctuations in the recruitment of PRC2 subunits promote changes in RNA synthesis and RNA polymerase II binding that are compromised in Jarid2 −/− mESCs. Overall, we show that differential recruitment of PRC2 subunits across cell cycle enables the establishment of a chromatin state that facilitates the induction of cell differentiation in G1 phase.This study was supported by the Spanish Ministry of Economy and Competitiveness (SAF2013-40891-R and BFU2016-75233-P) and the Andalusian Regional Government (PC-0246-2017). D.L. is a Ramón y Cajal researcher of the Spanish Ministry of Economy and Competitiveness (RYC-2012-10019)

Valuable Secondary Habitats or Hazardous Ecological Traps? Environmental Risk Assessment of Minor and Trace Elements in Fly Ash Deposits across the Czech Republic

Deposits of coal combustion wastes, especially fly ash, are sources of environmental and health risks in industrial regions. Recently, fly ash deposits have been reported as habitat surrogates for some threatened arthropods in Central Europe. However, the potential environmental risks of fly ash have not yet been assessed in the region. We analysed concentrations of 19 minor and trace elements in 19 lignite combustion waste deposits in the Czech Republic. We assessed their environmental risks by comparison with the national and EU legislation limits, and with several commonly used indices. Over 50% of the samples exceeded the Czech national limits for As, Cu, V, or Zn, whilst only V exceeded the EU limits. For some studied elements, the high-risk indices were detected in several localities. Nevertheless, the measured water characteristics, the long-term presence of fly ash, previous leaching by acid rains, and the low amount of organic matter altogether can infer low biological availability of these elements. We presume the revealed high concentrations of some heavy metals at some studied sites can be harmful for some colonising species. Nevertheless, more ecotoxicological research on particular species is needed for final decision on their conservation potential for terrestrial and freshwater biota.info:eu-repo/semantics/publishedVersio

The molecular clock protein Bmal1 regulates cell differentiation in mouse embryonic stem cells

Mammals optimize their physiology to the light–dark cycle by synchronization of the master circadian clock in the brain with peripheral clocks in the rest of the tissues of the body. Circadian oscillations rely on a negative feedback loop exerted by the molecular clock that is composed by transcriptional activators Bmal1 and Clock, and their negative regulators Period and Cryptochrome. Components of the molecular clock are expressed during early development, but onset of robust circadian oscillations is only detected later during embryogenesis. Here, we have used na¨ıve pluripotent mouse embryonic stem cells (mESCs) to study the role of Bmal1 during early development. We found that, compared to wild-type cells, Bmal12/2 mESCs express higher levels of Nanog protein and altered expression of pluripotencyassociated signalling pathways. Importantly, Bmal12/2 mESCs display deficient multi-lineage cell differentiation capacity during the formation of teratomas and gastrula-like organoids. Overall, we reveal that Bmal1 regulates pluripotent cell differentiation and propose that the molecular clock is an hitherto unrecognized regulator of mammalian development.Ramon y Cajal grant of the Spanish ministry of economy and competitiveness RYC2012-10019Spanish ministry of economy and competitiveness BFU2016-75233-PAndalusian regional government PC-0246-2017Fundacion Progreso y Salud (FPS)Instituto de Salud Carlos III European Union (EU) CPII17/00032 PI17/01574University of Granad

Molecular mechanisms underlying the control of antigenic variation in African trypanosomes

African trypanosomes escape the host adaptive immune response by switching their dense protective coat of Variant Surface Glycoprotein (VSG). Each cell expresses only one VSG gene at a time from a telomeric expression site (ES). The [`]pre-genomic' era saw the identification of the range of pathways involving VSG recombination in the context of mono-telomeric VSG transcription. A prominent feature of the early post-genomic era is the description of the molecular machineries involved in these processes. We describe the factors and sequences recently linked to mutually exclusive transcription and VSG recombination, and how these act in the control of the key virulence mechanism of antigenic variatio

Antigenic variation in <i>Trypanosoma brucei</i>: joining the DOTs

African trypanosomes, such as &lt;i&gt;Trypanosoma brucei&lt;/i&gt;, are protistan parasites that cause sleeping sickness. Though first described more than a century ago, trypanosomes remain a blight on the health of the human population and on the economy of sub-Saharan Africa. &lt;i&gt;T. brucei&lt;/i&gt; replicates in the bloodstream of infected mammals and traverses the blood-brain barrier to enter the central nervous system in the late, frequently fatal, stages of the disease. Because of its extracellular lifestyle, &lt;i&gt;T. brucei&lt;/i&gt; is continuously exposed to antibody challenge. To circumvent this, the parasite uses antigenic variation of a surface protein named the variant surface glycoprotein (VSG). Around 107 VSG molecules are expressed on the parasite's cell surface, creating a dense coat that prevents adaptive immunity from detecting or accessing invariant antigens. However, antibodies against the expressed VSG are generated, and periodic switches to an immunologically distinct VSG coat are necessary for parasite survival. Such switches are pre-emptive of the immune response and contribute to the pattern of trypanosome growth seen in an infected host (Figure 1): parasite numbers increase, but then drop as VSG-specific antibodies are raised by the host. Cells that have switched to another VSG coat survive this killing and seed the outgrowth of a subsequent peak of parasites, which is again decimated by anti-VSG immune killing. As a survival strategy, antigenic variation succeeds by prolonging the time that the parasite

Changes in PRC1 activity during interphase modulate lineage transition in pluripotent cells

We thank the core facilities at GENYO for excellent technical support. We also thank the genomics unit at the CRG for assistance with RNA-seq and ChIP-seq experiments. The Landeira lab is supported by the Spanish ministry of science and innovation (PID2019-108108-100, EUR2021- 122005), the Andalusian regional government (PIER-0211-2019, PY20_00681) and the University of Granada (A-BIO-6-UGR20) grants. Research in the Klose lab is supported by the Wellcome Trust (209400/ Z/17/Z) and the European Research Council (681440). A.F. was sup- ported by a Sir Henry Wellcome Post-doctoral fellowship (110286/Z/15/ Z). Work in the Rada-Iglesias lab is funded by the Ministerio de Ciencia e Innovación, the Agencia Española de Investigación and the European Regional Development Fund (PGC2018-095301-B-I00 and RED2018- 102553-T); by the European Research Council (862022); and by the European Commission (H2020-MSCA-ITN-2019-860002).The online version contains supplementary material available at https://doi.org/10.1038/s41467-023-35859-9The potential of pluripotent cells to respond to developmental cues and trigger cell differentiation is enhanced during the G1 phase of the cell cycle, but the molecular mechanisms involved are poorly understood. Variations in polycomb activity during interphase progression have been hypothesized to regulate the cell-cycle-phase-dependent transcriptional activation of differentiation genes during lineage transition in pluripotent cells. Here, we show that recruitment of Polycomb Repressive Complex 1 (PRC1) and associated molecular functions, ubiquitination of H2AK119 and three-dimensional chromatin interactions, are enhanced during S and G2 phases compared to the G1 phase. In agreement with the accumulation of PRC1 at target promoters upon G1 phase exit, cells in S and G2 phases show firmer transcriptional repression of developmental regulator genes that is drastically perturbed upon genetic ablation of the PRC1 catalytic subunit RING1B. Importantly, depletion of RING1B during retinoic acid stimu- lation interferes with the preference of mouse embryonic stem cells (mESCs) to induce the transcriptional activation of differentiation genes in G1 phase. We propose that incremental enrolment of polycomb repressive activity during interphase progression reduces the tendency of cells to respond to develop- mental cues during S and G2 phases, facilitating activation of cell differentiation in the G1 phase of the pluripotent cell cycle.Ministry of Science and Innovation, Spain (MICINN) Spanish Government PID2019-108108-100, EUR2021-122005Andalusian regional government PIER-0211-2019, PY20_00681University of Granada A-BIO-6-UGR20Wellcome Trust 209400/Z/17/ZEuropean Research Council (ERC) European Commission 862022Wellcome Trust PGC2018-095301-B-I00Ministry of Science and Innovation, Spain (MICINN) Instituto de Salud Carlos III Spanish GovernmentEuropean Commission RED2018-102553-T, H2020-MSCA-ITN-2019-860002European Commission European Commission Joint Research Centre 681440Agencia Española de Investigación110286/Z/15/