Chromatin opening elements allow tetracycline inducible gene expression in stem cells

In modern biomedicine, cell therapeutics are produced from reprogrammed stem cells (induced pluripotent stem cells, iPS). For this purpose, these cells can also undergo an additional genetic modification. However, unintentional mechanisms frequently occur, which can deactivate gene expression. Researchers from the Paul-Ehrlich-Institut (PEI) have developed inducible vectors containing ubiquitous chromatin opening elements (UCOE) which keep the relevant gene modified locus permanently open so that it remains active. The journal Biomaterials reports on these results in its online version of 24 November 2018. (Mehr in: Pressemitteilungen – idw – Informationsdienst Wissenschaft)

Chromatin-öffnende Elemente erlauben Tetrazyklin-induzierbare Genexpression in Stammzellen

In der modernen Biomedizin werden aus reprogrammierten Stammzellen (induzierte pluripotente Stammzellen, iPS) Zelltherapeutika hergestellt. Hierzu können diese Zellen im späteren Verlauf noch zusätzlich genetisch modifiziert werden. Allerdings treten häufig unbeabsichtigte Mechanismen auf, welche die Genexpression stilllegen. Forschende des Paul-Ehrlich-Instituts (PEI) haben induzierbare (aktivierbare) Vektoren entwickelt, die mit Hilfe eines Chromatin-öffnenden Elements (UCOE, ubiquitious chromatin opening element) den relevanten genmodifizierten Abschnitt dauerhaft offen und damit aktiv halten. Über die Forschungsergebnisse berichtet die Zeitschrift Biomaterials online vorab am 24.11.18. (Mehr in: Pressemitteilungen – idw – Informationsdienst Wissenschaft)

Chromatin structure mutes enhancer activity

In early embryos it is not possible to distinguish between arms and legs: their developmental precursors are virtually identical. Dynamic changes in the 3D structure of the chromatin in the leg precursor cells bring the Pen enhancer into spatial proximity to the Pitx1 gene, thus enabling the activation of this critical gene specifically in the future leg. As a result, forelimb and hindlimb buds subsequently differentiate from each other and produce arms and legs, respectively. This surprising molecular mechanism has been discovered by scientists around Stefan Mundlos and Guillaume Andrey from the Max Planck Institute for Molecular Genetics and the Charitè-Universitätsmedizin Berlin. (Mehr in: Pressemitteilungen – idw – Informationsdienst Wissenschaft)

Chromatin-Struktur legt Enhancer still

Im frühen Embryo lässt sich nicht erkennen, was ein Arm wird und was ein Bein: Die Knospen für die Extremitäten sehen zunächst identisch aus. Durch dynamische Veränderungen in der 3D-Struktur des Chromatins wird bei den unteren Extremitätenknospen der Enhancer Pen in räumliche Nähe zu dem Gen Pitx1 gebracht und kann es aktivieren. So erwachsen aus den oberen Knospen Arme und aus den unteren Beine. Den erstaunlichen molekularen Mechanismus haben Wissenschaftlerinnen und Wissenschaftler um Guillaume Andrey und Stefan Mundlos vom Max-Planck-Institut für molekulare Genetik und der Charité – Universitätsmedizin Berlin entdeckt und jetzt im renommierten Fachblatt „Nature Genetics“ veröffentlicht. (Mehr in: Pressemitteilungen – idw – Informationsdienst Wissenschaft)

Chromatinstruktur – Packen und entpacken

Im Zellkern ist das Erbgut dicht gewickelt. Trotzdem muss die Zelle immer wieder unterschiedliche Gene zugänglich machen. LMU-Forscher haben nun einen Mechanismus entschlüsselt, wie die molekularen Maschinen dabei DNA mit einem Zollstock aus Proteinen abmessen. (Mehr in: Pressemitteilungen – idw – Informationsdienst Wissenschaft)

3D structure of DNA forms defined room for dissociated lncRNAs to activate gene expression

Enhancers are regulatory regions of the DNA, giving rise to “long non-coding RNAs” (lncRNAs), which are known as crucial regulators of gene expression. Scientists from the Max Planck Institute for Molecular Genetics in Berlin now have shown that a lncRNA called A-ROD is only functional the moment it is released from chromatin into the nucleoplasm. In the current issue of Nature Communications the researchers demonstrate that the regulatory interaction requires dissociation of A-ROD from chromatin, with target specificity ensured within the pre-established chromosomal proximity. This can heavily influence our understanding of dynamic regulation of gene expression in biological processes. (Mehr in: Pressemitteilungen – idw – Informationsdienst Wissenschaft)

3D-Struktur der DNA bildet umschriebenen Raum zur Aktivierung der Genexpression durch freie lncRNAs

Forscher/innen des Max-Planck-Instituts für molekulare Genetik (MPIMG) in Berlin konnten zeigen, dass eine lncRNA namens A-ROD nur funktionstüchtig ist, wenn sie vom Chromatin in das Kernplasma abgegeben wird. In der aktuellen Ausgabe der Zeitschrift Nature Communications beschreiben die Wissenschaftler/innen, dass sich A-ROD vollständig vom Chromatin ablösen muss, um regulatorisch aktiv werden zu können. Durch die dreidimensionale Struktur der DNA wird gewährleistet, dass sich A-ROD bei der Ablösung bereits in direkter Nähe zu seinem Zielgen befindet. Dies kann unser Verständnis der dynamischen Regulation der Genexpression in biologischen Prozessen stark beeinflussen. (Mehr in: Pressemitteilungen – idw – Informationsdienst Wissenschaft)

Probing RNA epigenetics and chromatin structures to predict drug resistance in leukemia

A research team has begun to unravel the role of RNA epigenetics and chromatin structure in the regulation of 5-azacytidine, a DNA hypomethylating agent in certain leukemias. The results could lead to novel strategies and biomarkers that could reduce the risk of drug resistance. (Mehr in: Cancer News — ScienceDaily)

‚Icebreaker‘ protein opens genome for T-cell development, researchers find

Researchers describe the role of a transcription factor called TCF-1 in targeting the condensed chromatin and regulating the availability of genome sequences in T-cell development. The new connection between TCF-1 and chromatin will aid in developing new therapies using epigenetic drugs to alter T-cell fate in cancer, autoimmune disorders, and infectious diseases. (Mehr in: Cancer News — ScienceDaily)

Unveiling how nucleosome repositioning occurs to shed light on genetic diseases

For the first time, researchers have unveiled the three-dimensional structure of an overlapping dinucleosome, a newly discovered chromatin structural unit. This may explain how nucleosome repositioning occurs and provide valuable information for understanding chromatin dynamics during transcription and developing drugs to treat genetic diseases. (Mehr in: Cancer News — ScienceDaily)

Powerful optical imaging technology catches DNA naturally fluorescing

Biomedical engineers have developed imaging technology that is the first to see DNA ‚blink,‘ or fluoresce. The tool enables researchers to study individual biomolecules (DNA, chromatin, proteins) as well as important global patterns of gene expression, which could yield insights into cancer. (Mehr in: Cancer News — ScienceDaily)

Mechanical force triggers gene expression by stretching chromatin

How genes in our DNA are expressed into traits within a cell is a complicated mystery with many players, the main suspects being chemical. However, a new study has demonstrated that external mechanical force can directly regulate gene expression. The study also identified the pathway that conveys the force from the outside of the cell into the nucleus. (Mehr in: Cancer News — ScienceDaily)

Neuer Therapie-Ansatz gegen Leukämien entzieht Krebsgenen die Kontrolle

Krebsforscher des Universitären Centrums für Tumorerkrankungen (UCT) Mainz entwickelt zielgerichtete epigenetische Therapie gegen aggressive Leukämieform
In Leukämiezellen sind häufig Gene reaktiviert, die normalerweise eine Selbsterneuerung von Blutstammzellen vermitteln. Dr. Michael Kühn, Onkologe am UCT der Universitätsmedizin Mainz, konnte zusammen mit Wissenschaftlern aus den USA zeigen, dass die gezielte medikamentöse Inaktivierung von zwei bestimmten Chromatin-Regulatoren eine Abschaltung des Stammzellprogramms und eine Rückentwicklung der NPM1mut- Leukämiezellen zu normalen Blutzellen bewirkt. Die Ergebnisse sind online von Cancer Discovery publiziert. (Mehr in: Pressemitteilungen – idw – Informationsdienst Wissenschaft)

Open chromatin profiling key to identifying leukemia cells of origin

Researchers have found a precise and reliable way — whole-genome profiling of open chromatin — to identify the kind of cell that leads to a given case of leukemia, a valuable key to cancer prognosis and outcome. (Mehr in: Cancer News — ScienceDaily)

Jumping Genes Regulation of the Brain

Molecular Colors

The regulation of DNA is fantastically complex with many different layers: changing 3D shapes of the chromatin and loops of DNA; regional differences in nuclear DNA; large numbers of different epigenetic tags on DNA nucleotides and protective protein histone molecules; complex DNA repair mechanisms and alternative…

(Mehr in: Jon Lieff, M.D.)

More detailed analysis of how cells react to stress

Stress in the body’s cells is both the cause and consequence of inflammatory diseases or cancer. The cells react to stress to protect themselves. Researchers have now developed a new technique that allows studying a fundamental response to stress in much more detail than previously possible: the ADP-ribosylation of chromatin. In the long term, this method could help finding ways of blocking disease-causing processes. (Mehr in: Cancer News — ScienceDaily)

More detailed analysis of how cells react to stress

Stress in the body’s cells is both the cause and consequence of inflammatory diseases or cancer. The cells react to stress to protect themselves. Researchers at the University of Zurich have now developed a new technique that allows studying a fundamental response to stress in much more detail than previously possible: the ADP-ribosylation of chromatin. In the long term, this method could help finding ways of blocking disease-causing processes. (Mehr in: Pressemitteilungen – idw – Informationsdienst Wissenschaft)

Exaktere Analyse der Zellreaktionen auf Stress

Stress in den Zellen des Körpers sind sowohl Ursache als auch Konsequenz von Entzündungskrankheiten oder Krebs. Die Zellen reagieren auf diesen Stress, um sich vor Schädigungen zu schützen. Forschende der Universität Zürich haben nun ein neues Verfahren entwickelt, mit der sich eine elementare Stressantwort viel detaillierter als bisher untersuchen lässt: die ADP-Ribosylierung des Chromatins. Langfristig soll diese Methode helfen, krankmachende Prozesse zu unterbinden. (Mehr in: Pressemitteilungen – idw – Informationsdienst Wissenschaft)

How DNA is organized in our cells

A critical role for two proteins in chromatin structure has been uncovered by researchers. Their breakthrough helps explain how DNA is organized in our cells. This discovery could lead to a better understanding of what causes certain types of cancer, such as lymphoma. (Mehr in: Cancer News — ScienceDaily)

Vast Complexity of Chromatin 3D Shapes

B0003529 Chondrocyte showing organelles - coloured

Humans have only 21,000 genes—the same as a worm—and they are identical in all of the different types of cells. It is not the inherited code of the genes that determines the different cellular functions. Rather, it is way that genes are utilized differently in each type of cell that determines which proteins will…

(Mehr in: Jon Lieff, M.D.)

A global assessment of cancer genomic alterations in epigenetic mechanisms

Muhammad A Shah, Emily L Denton, Cheryl H Arrowsmith, Mathieu Lupien and Matthieu Schapira

Abstract

Background

The notion that epigenetic mechanisms may be central to cancer initiation and progression is supported by recent next-generation sequencing efforts revealing that genes involved in chromatin-mediated signaling are recurrently mutated in cancer patients.

Results

Here, we analyze mutational and transcriptional profiles from TCGA and the ICGC across a collection 441 chromatin factors and histones. Chromatin factors essential for rapid replication are frequently overexpressed, and those that maintain genome stability frequently mutated. We identify novel mutation hotspots such as K36M in histone H3.1, and uncover a general trend in which transcriptional profiles and somatic mutations in tumor samples favor increased transcriptionally repressive histone methylation, and defective chromatin remodeling.

Conclusions

This unbiased approach confirms previously published data, uncovers novel cancer-associated aberrations targeting epigenetic mechanisms, and justifies continued monitoring of chromatin-related alterations as a class, as more cancer types and distinct cancer stages are represented in cancer genomics data repositories.

Continue reading „A global assessment of cancer genomic alterations in epigenetic mechanisms“

Three-dimensional super-resolution microscopy of the inactive X chromosome territory reveals a collapse of its active nuclear compartment harboring distinct Xist RNA foci

3D-SIM-based DAPI intensity classification in the Barr body versus the entire nucleus of C2C12 cells. (A) Mid z-section of a DAPI-stained nucleus. The area below the dashed line illustrates the resolution level obtained by wide-field deconvolution microscopy, for comparison. Inset magnifications show the non-uniformly compacted structure of the Barr body resolvable with 3D-SIM (1) and an arbitrary autosomal region with CDCs (2). Scale bars: 5 μm, insets 1 μm. (B) X chromosome-specific painting (green) of Xi (left) and Xa territories (right) of the same nucleus in different z-sections. Note the high convergence between the painted Xi and the DAPI visualized Barr body (arrowheads). Scale bars: 2 μm, insets 1 μm. (C) 3D DAPI intensity classification exemplified for the nucleus shown in (A). Seven DAPI intensity classes displayed in false-color code ranging from class 1 (blue) representing pixels close to background intensity, largely representing the IC, up to class 7 (white) representing pixels with highest density, mainly associated with chromocenters. Framed areas of the Barr body (inset 1) and a representative autosomal region (inset 2) are shown on the right at resolution levels of 3D-SIM, deconvolution and conventional wide-field microscopy. The Xi territory pervaded by lower DAPI intensities becomes evident only at 3D-SIM resolution, whereas both wide-field and deconvolution microscopy imply a concentric increase of density in the Barr body. In the autosomal region, chromatin assigned to classes 2 to 3 lines compacted CDCs, represented by classes 4 to 6. (D) Left: average DAPI intensity classification profiles with standard deviations evaluated for entire nuclear volumes or the Barr body region only (dark grey bars). Right: over/underrepresentation of the average DAPI intensity class fraction sizes in the Barr body versus entire nuclear volumes (n = 12). Distribution differences on classes between Xi and entire nucleus P <0.001. 3D-SIM, three-dimensional structured illumination microscopy; CDC, chromatin domain cluster; DAPI, 4',6-diamidino-2-phenylindole; FISH, fluorescence in situ hybridization; IC, interchromatin compartment; Xa, active X chromosome; Xi, inactive X chromosome. Smeets et al. Epigenetics & Chromatin 2014 7:8   doi:10.1186/1756-8935-7-8
3D-SIM-based DAPI intensity classification in the Barr body versus the entire nucleus of C2C12 cells. (A) Mid z-section of a DAPI-stained nucleus. The area below the dashed line illustrates the resolution level obtained by wide-field deconvolution microscopy, for comparison. Inset magnifications show the non-uniformly compacted structure of the Barr body resolvable with 3D-SIM (1) and an arbitrary autosomal region with CDCs (2). Scale bars: 5 μm, insets 1 μm. (B) X chromosome-specific painting (green) of Xi (left) and Xa territories (right) of the same nucleus in different z-sections. Note the high convergence between the painted Xi and the DAPI visualized Barr body (arrowheads). Scale bars: 2 μm, insets 1 μm. (C) 3D DAPI intensity classification exemplified for the nucleus shown in (A). Seven DAPI intensity classes displayed in false-color code ranging from class 1 (blue) representing pixels close to background intensity, largely representing the IC, up to class 7 (white) representing pixels with highest density, mainly associated with chromocenters. Framed areas of the Barr body (inset 1) and a representative autosomal region (inset 2) are shown on the right at resolution levels of 3D-SIM, deconvolution and conventional wide-field microscopy. The Xi territory pervaded by lower DAPI intensities becomes evident only at 3D-SIM resolution, whereas both wide-field and deconvolution microscopy imply a concentric increase of density in the Barr body. In the autosomal region, chromatin assigned to classes 2 to 3 lines compacted CDCs, represented by classes 4 to 6. (D) Left: average DAPI intensity classification profiles with standard deviations evaluated for entire nuclear volumes or the Barr body region only (dark grey bars). Right: over/underrepresentation of the average DAPI intensity class fraction sizes in the Barr body versus entire nuclear volumes (n = 12). Distribution differences on classes between Xi and entire nucleus P Smeets et al. Epigenetics & Chromatin 2014 7:8 doi:10.1186/1756-8935-7-8

Daniel Smeets, Yolanda Markaki, Volker J Schmid, Felix Kraus, Anna Tattermusch, Andrea Cerase, Michael Sterr, Susanne Fiedler, Justin Demmerle, Jens Popken, Heinrich Leonhardt, Neil Brockdorff, Thomas Cremer1, Lothar Schermelleh and Marion Cremer

Abstract

Background

A Xist RNA decorated Barr body is the structural hallmark of the compacted inactive X territory in female mammals. Using super-resolution three-dimensional structured illumination microscopy (3D-SIM) and quantitative image analysis, we compared its ultrastructure with active chromosome territories (CTs) in human and mouse somatic cells, and explored the spatio-temporal process of Barr body formation at onset of inactivation in early differentiating mouse embryonic stem cells (ESCs).

Results

We demonstrate that all CTs are composed of structurally linked chromatin domain clusters (CDCs). In active CTs the periphery of CDCs harbors low-density chromatin enriched with transcriptionally competent markers, called the perichromatin region (PR). The PR borders on a contiguous channel system, the interchromatin compartment (IC), which starts at nuclear pores and pervades CTs. We propose that the PR and macromolecular complexes in IC channels together form the transcriptionally permissive active nuclear compartment (ANC). The Barr body differs from active CTs by a partially collapsed ANC with CDCs coming significantly closer together, although a rudimentary IC channel system connected to nuclear pores is maintained. Distinct Xist RNA foci, closely adjacent to the nuclear matrix scaffold attachment factor-A (SAF-A) localize throughout Xi along the rudimentary ANC. In early differentiating ESCs initial Xist RNA spreading precedes Barr body formation, which occurs concurrent with the subsequent exclusion of RNA polymerase II (RNAP II). Induction of a transgenic autosomal Xist RNA in a male ESC triggers the formation of an ‘autosomal Barr body’ with less compacted chromatin and incomplete RNAP II exclusion.

Conclusions

3D-SIM provides experimental evidence for profound differences between the functional architecture of transcriptionally active CTs and the Barr body. Basic structural features of CT organization such as CDCs and IC channels are however still recognized, arguing against a uniform compaction of the Barr body at the nucleosome level. The localization of distinct Xist RNA foci at boundaries of the rudimentary ANC may be considered as snap-shots of a dynamic interaction with silenced genes. Enrichment of SAF-A within Xi territories and its close spatial association with Xist RNA suggests their cooperative function for structural organization of Xi.

Continue reading „Three-dimensional super-resolution microscopy of the inactive X chromosome territory reveals a collapse of its active nuclear compartment harboring distinct Xist RNA foci“