scholarly journals Architectural proteins CTCF and cohesin have distinct roles in modulating the higher order structure and expression of the CFTR locus

2014 ◽  
Vol 42 (15) ◽  
pp. 9612-9622 ◽  
Author(s):  
Nehal Gosalia ◽  
Daniel Neems ◽  
Jenny L. Kerschner ◽  
Steven T. Kosak ◽  
Ann Harris
Keyword(s):  
Nuclear Periphery ◽  
Three Dimensional ◽  
Higher Order ◽  
Ctcf Binding ◽  
Cftr Gene ◽  
Dimensional Structure ◽  
Order Structure ◽  
Cis Acting ◽  
Architectural Proteins ◽  
Enhancer Blocking

Abstract Higher order chromatin structures across the genome are maintained in part by the architectural proteins CCCTC binding factor (CTCF) and the cohesin complex, which co-localize at many sites across the genome. Here, we examine the role of these proteins in mediating chromatin structure at the cystic fibrosis transmembrane conductance regulator (CFTR) gene. CFTR encompasses nearly 200 kb flanked by CTCF-binding enhancer-blocking insulator elements and is regulated by cell-type-specific intronic enhancers, which loop to the promoter in the active locus. SiRNA-mediated depletion of CTCF or the cohesin component, RAD21, showed that these two factors have distinct roles in regulating the higher order organization of CFTR. CTCF mediates the interactions between CTCF/cohesin binding sites, some of which have enhancer-blocking insulator activity. Cohesin shares this tethering role, but in addition stabilizes interactions between the promoter and cis-acting intronic elements including enhancers, which are also dependent on the forkhead box A1/A2 (FOXA1/A2) transcription factors (TFs). Disruption of the three-dimensional structure of the CFTR gene by depletion of CTCF or RAD21 increases gene expression, which is accompanied by alterations in histone modifications and TF occupancy across the locus, and causes internalization of the gene from the nuclear periphery.

scholarly journals Nuclear peripheral chromatin-lamin B1 interaction is required for global integrity of chromatin architecture and dynamics in human cells

2020 ◽  
Author(s):  
Lei Chang ◽  
Mengfan Li ◽  
Shipeng Shao ◽  
Chen Li ◽  
Shanshan Ai ◽  
...  
Keyword(s):  
Nuclear Periphery ◽  
Three Dimensional ◽  
Higher Order ◽  
Order Structure ◽  
Chromatin Dynamics ◽  
Lamin B1 ◽  
Chromatin Architecture ◽  
Chromatin Interactions ◽  
Molecular Machinery

Abstract The eukaryotic genome is folded into higher-order conformation accompanied with constrained dynamics for coordinated genome functions. However, the molecular machinery underlying these hierarchically organized three-dimensional (3D) chromatin architecture and dynamics remains poorly understood. Here by combining imaging and sequencing, we studied the role of lamin B1 in chromatin architecture and dynamics. We found that lamin B1 depletion leads to detachment of lamina-associated domains (LADs) from the nuclear periphery accompanied with global chromatin redistribution and decompaction. Consequently, the inter-chromosomal as well as inter-compartment interactions are increased, but the structure of topologically associating domains (TADs) is not affected. Using live-cell genomic loci tracking, we further proved that depletion of lamin B1 leads to increased chromatin dynamics, owing to chromatin decompaction and redistribution toward nucleoplasm. Taken together, our data suggest that lamin B1 and chromatin interactions at the nuclear periphery promote LAD maintenance, chromatin compaction, genomic compartmentalization into chromosome territories and A/B compartments and confine chromatin dynamics, supporting their crucial roles in chromatin higher-order structure and chromatin dynamics.

scholarly journals Hit the brakes – a new perspective on the loop extrusion mechanism of cohesin and other SMC complexes

2021 ◽  
Vol 134 (1) ◽  
pp. jcs247577
Author(s):  
Avi Matityahu ◽  
Itay Onn
Keyword(s):  
Present Model ◽  
Protein Complexes ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Order Structure ◽  
Chromatid Cohesion ◽  
Topologically Associated Domains ◽  
Extrusion Mechanism ◽  
New Perspective ◽  
Structural Maintenance Of Chromosome

ABSTRACTThe three-dimensional structure of chromatin is determined by the action of protein complexes of the structural maintenance of chromosome (SMC) family. Eukaryotic cells contain three SMC complexes, cohesin, condensin, and a complex of Smc5 and Smc6. Initially, cohesin was linked to sister chromatid cohesion, the process that ensures the fidelity of chromosome segregation in mitosis. In recent years, a second function in the organization of interphase chromatin into topologically associated domains has been determined, and loop extrusion has emerged as the leading mechanism of this process. Interestingly, fundamental mechanistic differences exist between mitotic tethering and loop extrusion. As distinct molecular switches that aim to suppress loop extrusion in different biological contexts have been identified, we hypothesize here that loop extrusion is the default biochemical activity of cohesin and that its suppression shifts cohesin into a tethering mode. With this model, we aim to provide an explanation for how loop extrusion and tethering can coexist in a single cohesin complex and also apply it to the other eukaryotic SMC complexes, describing both similarities and differences between them. Finally, we present model-derived molecular predictions that can be tested experimentally, thus offering a new perspective on the mechanisms by which SMC complexes shape the higher-order structure of chromatin.

scholarly journals Chromosomal G-dark Bands Determine the Spatial Organization of Centromeric Heterochromatin in the Nucleus

2001 ◽  
Vol 12 (11) ◽  
pp. 3563-3572 ◽  
Author(s):  
Célia Carvalho ◽  
Henrique M. Pereira ◽  
João Ferreira ◽  
Cristina Pina ◽  
Denise Mendonça ◽  
...  
Keyword(s):  
Satellite Dna ◽  
Spatial Organization ◽  
Nuclear Periphery ◽  
Three Dimensional ◽  
Lymphoid Cells ◽  
Spatial Proximity ◽  
Centromeric Heterochromatin ◽  
Chromosome 11 ◽  
Cis Acting ◽  
Tight Correlation

Gene expression can be silenced by proximity to heterochromatin blocks containing centromeric α-satellite DNA. This has been shown experimentally through cis-acting chromosome rearrangements resulting in linear genomic proximity, or throughtrans-acting changes resulting in intranuclear spatial proximity. Although it has long been been established that centromeres are nonrandomly distributed during interphase, little is known of what determines the three-dimensional organization of these silencing domains in the nucleus. Here, we propose a model that predicts the intranuclear positioning of centromeric heterochromatin for each individual chromosome. With the use of fluorescence in situ hybridization and confocal microscopy, we show that the distribution of centromeric α-satellite DNA in human lymphoid cells synchronized at G0/G1is unique for most individual chromosomes. Regression analysis reveals a tight correlation between nuclear distribution of centromeric α-satellite DNA and the presence of G-dark bands in the corresponding chromosome. Centromeres surrounded by G-dark bands are preferentially located at the nuclear periphery, whereas centromeres of chromosomes with a lower content of G-dark bands tend to be localized at the nucleolus. Consistent with the model, a t(11; 14) translocation that removes G-dark bands from chromosome 11 causes a repositioning of the centromere, which becomes less frequently localized at the nuclear periphery and more frequently associated with the nucleolus. The data suggest that “chromosomal environment” plays a key role in the intranuclear organization of centromeric heterochromatin. Our model further predicts that facultative heterochromatinization of distinct genomic regions may contribute to cell-type specific patterns of centromere localization.

scholarly journals Three-dimensional localization of CENP-A suggests a complex higher order structure of centromeric chromatin

2008 ◽  
Vol 183 (7) ◽  
pp. 1193-1202 ◽  
Author(s):  
Owen J. Marshall ◽  
Alan T. Marshall ◽  
K.H. Andy Choo
Keyword(s):  
Chromatin Immunoprecipitation ◽  
Protein A ◽  
Three Dimensional ◽  
Higher Order ◽  
Order Structure ◽  
Mitotic Chromosomes ◽  
Centromeric Chromatin ◽  
Centromere Protein A ◽  
Transmission Electron ◽  
Histone H3 Variant

The histone H3 variant centromere protein A (CENP-A) is central to centromere formation throughout eukaryotes. A long-standing question in centromere biology has been the organization of CENP-A at the centromere and its implications for the structure of centromeric chromatin. In this study, we describe the three-dimensional localization of CENP-A at the inner kinetochore plate through serial-section transmission electron microscopy of human mitotic chromosomes. At the kinetochores of normal centromeres and at a neocentromere, CENP-A occupies a compact domain at the inner kinetochore plate, stretching across two thirds of the length of the constriction but encompassing only one third of the constriction width and height. Within this domain, evidence of substructure is apparent. Combined with previous chromatin immunoprecipitation results (Saffery, R., H. Sumer, S. Hassan, L.H. Wong, J.M. Craig, K. Todokoro, M. Anderson, A. Stafford, and K.H.A. Choo. 2003. Mol. Cell. 12:509–516; Chueh, A.C., L.H. Wong, N. Wong, and K.H.A. Choo. 2005. Hum. Mol. Genet. 14:85–93), our data suggest that centromeric chromatin is arranged in a coiled 30-nm fiber that is itself coiled or folded to form a higher order structure.

Low-frequency seismology and the three-dimensional structure of the Earth

1989 ◽  
Vol 328 (1599) ◽  
pp. 329-349 ◽  
Keyword(s):  
Large Scale ◽  
Inner Core ◽  
Three Dimensional ◽  
Low Frequency ◽  
Quantitative Agreement ◽  
Phase Behaviour ◽  
Dimensional Structure ◽  
Order Structure ◽  
Three Dimensional Structure ◽  
The Earth

We attempt to catalogue those features of the three-dimensional structure of the Earth that are well-constrained by low-frequency data (i.e. periods longer than about 125 seconds). The dominant signals in such data are the surface-wave equivalent modes whose phase characteristics are mainly affected by a large scale structure of harmonic degree two in the upper mantle. Available aspherical models predict this phase behaviour quite well, but do not give an accurate prediction of the observed waveforms and we must appeal to higher-order structure an d /o r coupling effects to give the observed complexity of the data. Strong splitting of modes which sample the cores of the Earth is also observed and, though we do not yet have a model of aspherical structure which gives quantitative agreement with these data, anisotropy or large-scale aspherical structure in the inner core appears to be required to model the observed signal.

scholarly journals Capturing the complexity of topologically associating domains through multi-feature optimization

2021 ◽  
Author(s):  
Natalie Sauerwald ◽  
Carl Kingsford
Keyword(s):  
Binding Sites ◽  
Three Dimensional ◽  
Replication Timing ◽  
Biological Properties ◽  
Ctcf Binding ◽  
Dimensional Structure ◽  
Biological Targets ◽  
Topologically Associating Domains ◽  
Contact Frequency ◽  
Algorithmic Framework

AbstractThe three-dimensional structure of human chromosomes is tied to gene regulation and replication timing, but there is still a lack of consensus on the computational and biological definitions for chromosomal substructures such as topologically associating domains (TADs). TADs are described and identified by various computational properties leading to different TAD sets with varying compatibility with biological properties such as boundary occupancy of structural proteins. We unify many of these computational and biological targets into one algorithmic framework that jointly maximizes several computational TAD definitions and optimizes TAD selection for a quantifiable biological property. Using this framework, we explore the variability of TAD sets optimized for six different desirable properties of TAD sets: high occupancy of CTCF, RAD21, and H3K36me3 at boundaries, reproducibility between replicates, high intra- vs inter-TAD difference in contact frequencies, and many CTCF binding sites at boundaries. The compatibility of these biological targets varies by cell type, and our results suggest that these properties are better reflected as subpopulations or families of TADs rather than a singular TAD set fitting all TAD definitions and properties. We explore the properties that produce similar TAD sets (reproducibility and inter- vs intra-TAD difference, for example) and those that lead to very different TADs (such as CTCF binding sites and inter- vs intra-TAD contact frequency difference).

scholarly journals Analytical Techniques for Structural Characterization of Proteins in Solid Pharmaceutical Forms: An Overview

Pharmaceutics ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 534
Author(s):  
Aljoša Bolje ◽  
Stanislav Gobec
Keyword(s):  
Drug Product ◽  
Conformational Stability ◽  
Physical Stability ◽  
Three Dimensional ◽  
Chemical Properties ◽  
Analytical Techniques ◽  
Dimensional Structure ◽  
Order Structure ◽  
Careful Study ◽  
Formulation Development

Therapeutic proteins as biopharmaceuticals have emerged as a very important class of drugs for the treatment of many diseases. However, they are less stable compared to conventional pharmaceuticals. Their long-term stability in solid forms, which is critical for product performance, depends heavily on the retention of the native protein structure during the lyophilization (freeze-drying) process and, thereafter, in the solid state. Indeed, the biological function of proteins is directly related to the tertiary and secondary structure. Besides physical stability and biological activity, conformational stability (three-dimensional structure) is another important aspect when dealing with protein pharmaceuticals. Moreover, denaturation as loss of higher order structure is often a precursor to aggregation or chemical instability. Careful study of the physical and chemical properties of proteins in the dried state is therefore critical during biopharmaceutical drug development to deliver a final drug product with built-in quality that is safe, high-quality, efficient, and affordable for patients. This review provides an overview of common analytical techniques suitable for characterizing pharmaceutical protein powders, providing structural, and conformational information, as well as insights into dynamics. Such information can be very useful in formulation development, where selecting the best formulation for the drug can be quite a challenge.

scholarly journals Novel regulatory mechanisms for the CFTR gene

2009 ◽  
Vol 37 (4) ◽  
pp. 843-848 ◽  
Author(s):  
Christopher J. Ott ◽  
Neil P. Blackledge ◽  
Shih-Hsing Leir ◽  
Ann Harris
Keyword(s):  
Cystic Fibrosis ◽  
Expression Profiles ◽  
Regulatory Elements ◽  
Cftr Gene ◽  
Dimensional Structure ◽  
Regulatory Sequences ◽  
Physical Interaction ◽  
Basic Leucine Zipper ◽  
Tissue Specific ◽  
Cis Acting

The CFTR (cystic fibrosis transmembrane conductance regulator) gene, which when mutated causes cystic fibrosis, encompasses nearly 200 kb of genomic DNA at chromosome 7q31.2. It is flanked by two genes ASZ1 [ankyrin repeat, SAM (sterile α-motif) and basic leucine zipper] and CTTNBP2 (cortactin-binding protein 2), which have very different expression profiles. CFTR is expressed primarily in specialized epithelial cells, whereas ASZ1 is transcribed exclusively in the testis and ovary, and CTTNBP2 is highly expressed in the brain, kidney and pancreas, with lower levels of expression in other tissues. Despite its highly regulated pattern of expression, the promoter of the CFTR gene apparently lacks the necessary elements to achieve this. We previously suggested that cis-acting regulatory elements elsewhere in the locus, both flanking the gene and within introns, were required to co-ordinate regulated, tissue-specific expression of CFTR. We identified a number of crucial elements, including enhancer-blocking insulators flanking the locus, intronic tissue-specific enhancers and also characterized some of the interacting proteins. We recently employed a high-resolution method of mapping DHS (DNase I-hypersensitive sites) using tiled microarrays. DHS are often associated with regulatory elements and use of this technique generated cell-specific profiles of potential regulatory sequences in primary cells and cell lines. We characterized a set of cis-acting elements within the CFTR locus and demonstrated direct physical interaction between them and the CFTR promoter, by chromosome conformation capture (3C). These results provide the first insight into the three-dimensional structure of the active CFTR gene.

Intact “biological motion” and “structure from motion” perception in a patient with impaired motion mechanisms: A case study

1990 ◽  
Vol 5 (04) ◽  
pp. 353-369 ◽  
Author(s):  
Lucia M. Vaina ◽  
Marjorie Lemay ◽  
Don C. Bienfang ◽  
Albert Y. Choi ◽  
Ken Nakayama
Keyword(s):  
Structure From Motion ◽  
Random Noise ◽  
Higher Order ◽  
Image Motion ◽  
Coherent Motion ◽  
Dimensional Structure ◽  
Motion Processing ◽  
Order Structure ◽  
Impaired Performance ◽  
Early Motion

AbstactA series of psychophysical tests examining early and later aspects of image-motion processing were conducted in a patient with bilateral lesions involving the posterior visual pathways, affecting the lateral parietal-temporal-occipital cortex and the underlying white matter (as shown by magnetic resonance imaging studies and confirmed by neuro-ophthalmological and neuropsychological examinations). Visual acuity, form discrimination, color, and contrast-sensitivity discrimination were normal whereas spatial localization, line bisection, depth, and binocular stereopsis were severely impaired. Performance on early motion tasks was very poor. These include seeing coherent motion in random noise (Newsome & Paré, 1988), speed discrimination, and seeing two-dimensional form from relative speed of motion. However, on higher-order motion tasks the patient was able to identify actions from the evolving pattern of dots placed at the joints of a human actor (Johansson, 1973) as well as discriminating three-dimensional structure of a cylinder from motion in a dynamic random-dot field. The pattern of these results is at odds with the hypothesis that precise metrical comparison of early motion measurements is necessary for higher-order “structure from motion” tasks.

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