scholarly journals High-resolution mapping of regulatory element interactions and genome architecture using ARC-C

2018 ◽  
Author(s):  
Ni Huang ◽  
Wei Qiang Seow ◽  
Julie Ahringer
Keyword(s):  
High Resolution ◽  
Regulatory Element ◽  
Regulatory Elements ◽  
Genome Architecture ◽  
Chromosome Conformation ◽  
Regulatory Interactions ◽  
C Elegans ◽  
Chromatin Regulators ◽  
Accessible Region ◽  
Resolution Mapping

AbstractInteractions between cis-regulatory elements such as promoters and enhancers are important for transcription but global identification of these interactions remains a major challenge. Leveraging the chromatin accessiblity of regulatory elements, we developed ARC-C (accessible region chromosome conformation capture), which profiles chromatin regulatory interactions genome-wide at high resolution. Applying ARC-C to C. elegans, we identify ~15,000 significant interactions at 500bp resolution. Regions bound by transcription factors and chromatin regulators such as cohesin and condensin II are enriched for interactions, and we use ARC-C to show that the BLMP-1 transcription factor mediates interactions between its targets. Investigating domain level architecture, we find that C. elegans chromatin domains defined by either active or repressive modifications form topologically associating domains (TADs) and that these domains interact to form A/B (active/inactive) compartment structure. ARC-C is a powerful new tool to interrogate genome architecture and regulatory interactions at high resolution.

scholarly journals Accessible Region Conformation Capture (ARC-C) gives high resolution insights into genome architecture and regulation

2021 ◽  
pp. gr.275669.121
Author(s):  
Ni Huang ◽  
Wei Qiang Seow ◽  
Alex Appert ◽  
Yan Dong ◽  
Przemyslaw Stempor ◽  
...  
Keyword(s):  
High Resolution ◽  
Regulatory Elements ◽  
Genome Architecture ◽  
Regulatory Interactions ◽  
C Elegans ◽  
Chromatin Regulators ◽  
Chromatin Interactions ◽  
Genome Wide ◽  
Accessible Region ◽  
Domain Level

Nuclear organization and chromatin interactions are important for genome function, yet determining chromatin connections at high-resolution remains a major challenge. To address this, we developed Accessible Region Conformation Capture (ARC-C), which profiles interactions between regulatory elements genome-wide without a capture step. Applied to C. elegans, we identify ~15,000 significant interactions between regulatory elements at 500bp resolution. Of 105 TFs or chromatin regulators tested, we find that the binding sites of 60 are enriched for interacting with each other, making them candidates for mediating interactions. These include cohesin and condensin II. Applying ARC-C to a mutant of transcription factor BLMP-1 detected changes in interactions between its targets. ARC-C simultaneously profiles domain level architecture, and we observe that C. elegans chromatin domains defined by either active or repressive modifications form topologically associating domains (TADs) which interact with A/B (active/inactive) compartment-like structure. Furthermore, we discovered that inactive compartment interactions are dependent on H3K9 methylation. ARC-C is a powerful new tool to interrogate genome architecture and regulatory interactions at high resolution.

scholarly journals High-resolution analysis of cis -acting regulatory networks at the α-globin locus

2013 ◽  
Vol 368 (1620) ◽  
pp. 20120361 ◽  
Author(s):  
Jim R. Hughes ◽  
Karen M. Lower ◽  
Ian Dunham ◽  
Stephen Taylor ◽  
Marco De Gobbi ◽  
...  
Keyword(s):  
High Resolution ◽  
Gene Cluster ◽  
Regulatory Networks ◽  
Single Gene ◽  
Regulatory Elements ◽  
Circular Chromosome ◽  
Chromosome Conformation ◽  
Cis Acting ◽  
High Resolution Analysis ◽  
Resolution Analysis

We have combined the circular chromosome conformation capture protocol with high-throughput, genome-wide sequence analysis to characterize the cis -acting regulatory network at a single locus. In contrast to methods which identify large interacting regions (10–1000 kb), the 4C approach provides a comprehensive, high-resolution analysis of a specific locus with the aim of defining, in detail, the cis -regulatory elements controlling a single gene or gene cluster. Using the human α-globin locus as a model, we detected all known local and long-range interactions with this gene cluster. In addition, we identified two interactions with genes located 300 kb (NME4) and 625 kb (FAM173a) from the α-globin cluster.

scholarly journals Analysis of sub-kilobase chromatin topology reveals nano-scale regulatory interactions with variable dependence on cohesin and CTCF

2021 ◽  
Author(s):  
Abrar Aljahani ◽  
Peng Hua ◽  
Magdalena A. Karpinska ◽  
Kimberly Quililan ◽  
James O. J. Davies ◽  
...  
Keyword(s):  
Large Scale ◽  
Regulation Of Gene Expression ◽  
Regulatory Elements ◽  
Interaction Data ◽  
Chromosome Conformation ◽  
Regulatory Interactions ◽  
Nano Scale ◽  
Topologically Associating Domains ◽  
Scale Structures ◽  
Local Insulation

Enhancers and promoters predominantly interact within large-scale topologically associating domains (TADs), which are formed by loop extrusion mediated by cohesin and CTCF. However, it is unclear whether complex chromatin structures exist at sub-kilobase-scale and to what extent fine-scale regulatory interactions depend on loop extrusion. To address these questions, we present an MNase-based chromosome conformation capture (3C) approach, which has enabled us to generate the most detailed local interaction data to date and precisely investigate the effects of cohesin and CTCF depletion on chromatin architecture. Our data reveal that cis-regulatory elements have distinct internal nano-scale structures, within which local insulation is dependent on CTCF, but which are independent of cohesin. In contrast, we find that depletion of cohesin causes a subtle reduction in longer-range enhancer-promoter interactions and that CTCF depletion can cause rewiring of regulatory contacts. Together, our data show that loop extrusion is not essential for enhancer-promoter interactions, but contributes to their robustness and specificity and to precise regulation of gene expression.

scholarly journals Three classes of epigenomic regulators converge to hyperactivate the essential maternal gene deadhead within a heterochromatin mini-domain.

2021 ◽  
Author(s):  
Daniela Torres-Campana ◽  
Béatrice Horard ◽  
Sandrine Denaud ◽  
Gérard Benoit ◽  
Benjamin Loppin ◽  
...  
Keyword(s):  
Regulatory Element ◽  
Critical Role ◽  
Extreme Case ◽  
Regulatory Elements ◽  
Chromatin Regulators ◽  
Data Analysis Procedure ◽  
Chromatin Profiling ◽  
Dna Regulatory Elements ◽  
Female Germline ◽  
Maternal Gene

The formation of a diploid zygote is a highly complex cellular process that is entirely controlled by maternal gene products stored in the egg cytoplasm. This highly specialized transcriptional program is tightly controlled at the chromatin level in the female germline. As an extreme case in point, the massive and specific ovarian expression of the essential thioredoxin Deadhead (DHD) is critically regulated in Drosophila by the histone demethylase Lid and its partner, the histone deacetylase complex scaffold Sin3A, via yet unknown mechanisms. Here, we identified the Brahma chromatin remodeler sub-unit Snr1 and the insulator component Mod(mdg4) as essential for dhd expression and investigated how these epigenomic effectors act with Lid and Sin3A to hyperactivate dhd . Using Cut&Run chromatin profiling with a dedicated data analysis procedure, we found that dhd is intriguingly embedded in an H3K27me3/H3K9me3-enriched mini-domain flanked by DNA regulatory elements, including a dhd promoter-proximal element essential for its expression. Surprisingly, Lid, Sin3A, Snr1 and Mod(mdg4) impact H3K27me3 and this regulatory element in distinct manners. However, we show that these effectors activate dhd independently of H3K27me3/H3K9me3, and that these marks are not required to repress dhd . Together, our study demonstrates an atypical and critical role for chromatin regulators Lid, Sin3A, Snr1 and Mod(mdg4) to trigger tissue-specific hyperactivation within a unique heterochromatin mini-domain.

scholarly journals Determinants of Transcription Initiation Directionality in Metazoans

2017 ◽  
Author(s):  
Mahmoud M. Ibrahim ◽  
Aslihan Karabacak ◽  
Alexander Glahs ◽  
Ena Kolundzic ◽  
Antje Hirsekorn ◽  
...  
Keyword(s):  
Transcription Initiation ◽  
Core Promoter ◽  
Regulatory Element ◽  
Promoter Sequence ◽  
Regulatory Elements ◽  
Integrative Model ◽  
Promoter Regions ◽  
Promoter Sequences ◽  
C Elegans ◽  
Divergent Transcription

AbstractDivergent transcription from promoters and enhancers is pervasive in many species, but it remains unclear if it is a general and passive feature of all eukaryotic cis regulatory elements. To address this, we define promoters and enhancers in C. elegans, D. melanogaster and H. sapiens using ATAC-Seq and investigate the determinants of their transcription initiation directionalities by analyzing genome-wide nascent, cap-selected, polymerase run-on assays. All three species initiate divergent transcription from separate core promoter sequences. Sequence asymmetry downstream of forward and reverse initiation sites, known to be important for termination and stability in H. sapiens, is unique in each species. Chromatin states of divergent promoters are not entirely conserved, but in all three species, the levels of histone modifications on the +1 nucleosome are independent from those on the -1 nucleosome, arguing for independent initiation events. This is supported by an integrative model of H3K4me3 levels and core promoter sequence that is highly predictive of promoter directionality and of two types of promoters: those with balanced initiation directionality and those with skewed directionality. Lastly, D. melanogaster enhancers display variation in chromatin architecture depending on enhancer location, and D. melanogaster promoter regions with dual enhancer/promoter potential are enriched for divergent transcription. Our results point to a high degree of variation in regulatory element transcription initiation directionality within and between metazoans, and to non-passive regulatory mechanisms of transcription initiation directionality in those species.

scholarly journals Chromosome looping at the human α-globin locus is mediated via the major upstream regulatory element (HS −40)

Blood ◽  
2009 ◽  
Vol 114 (19) ◽  
pp. 4253-4260 ◽  
Author(s):  
Douglas Vernimmen ◽  
Fatima Marques-Kranc ◽  
Jacqueline A. Sharpe ◽  
Jacqueline A. Sloane-Stanley ◽  
William G. Wood ◽  
...  
Keyword(s):  
Long Range ◽  
Regulatory Element ◽  
Globin Gene ◽  
Regulatory Elements ◽  
Chromosome Conformation ◽  
Long Range Interactions ◽  
Upstream Regulatory Element ◽  
Physical Interactions ◽  
Globin Promoter

Abstract Previous studies in the mouse have shown that high levels of α-globin gene expression in late erythropoiesis depend on long-range, physical interactions between remote upstream regulatory elements and the globin promoters. Using quantitative chromosome conformation capture (q3C), we have now analyzed all interactions between 4 such elements lying 10 to 50 kb upstream of the human α cluster and their interactions with the α-globin promoter. All of these elements interact with the α-globin gene in an erythroid-specific manner. These results were confirmed in a mouse model of human α globin expression in which the human cluster replaces the mouse cluster in situ (humanized mouse). We have also shown that expression and all of the long-range interactions depend largely on just one of these elements; removal of the previously characterized major regulatory element (called HS −40) results in loss of all the interactions and α-globin expression. Reinsertion of this element at an ectopic location restores both expression and the intralocus interactions. In contrast to other more complex systems involving multiple upstream elements and promoters, analysis of the human α-globin cluster during erythropoiesis provides a simple and tractable model to understand the mechanisms underlying long-range gene regulation.

scholarly journals High-resolution targeted 3C interrogation of cis-regulatory element organization at genome-wide scale

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Damien J. Downes ◽  
Robert A. Beagrie ◽  
Matthew E. Gosden ◽  
Jelena Telenius ◽  
Stephanie J. Carpenter ◽  
...  
Keyword(s):  
Gene Expression ◽  
High Resolution ◽  
Regulatory Element ◽  
Gene Promoters ◽  
Entire Genome ◽  
Chromosome Conformation ◽  
Genome Wide ◽  
Wide Scale

AbstractChromosome conformation capture (3C) provides an adaptable tool for studying diverse biological questions. Current 3C methods generally provide either low-resolution interaction profiles across the entire genome, or high-resolution interaction profiles at limited numbers of loci. Due to technical limitations, generation of reproducible high-resolution interaction profiles has not been achieved at genome-wide scale. Here, to overcome this barrier, we systematically test each step of 3C and report two improvements over current methods. We show that up to 30% of reporter events generated using the popular in situ 3C method arise from ligations between two individual nuclei, but this noise can be almost entirely eliminated by isolating intact nuclei after ligation. Using Nuclear-Titrated Capture-C, we generate reproducible high-resolution genome-wide 3C interaction profiles by targeting 8055 gene promoters in erythroid cells. By pairing high-resolution 3C interaction calls with nascent gene expression we interrogate the role of promoter hubs and super-enhancers in gene regulation.

scholarly journals Three classes of epigenomic regulators converge to hyperactivate the essential maternal gene deadhead within a heterochromatin mini-domain

PLoS Genetics ◽  
2022 ◽  
Vol 18 (1) ◽  
pp. e1009615
Author(s):  
Daniela Torres-Campana ◽  
Béatrice Horard ◽  
Sandrine Denaud ◽  
Gérard Benoit ◽  
Benjamin Loppin ◽  
...  
Keyword(s):  
Regulatory Element ◽  
Critical Role ◽  
Extreme Case ◽  
Regulatory Elements ◽  
Chromatin Regulators ◽  
Data Analysis Procedure ◽  
Chromatin Profiling ◽  
Dna Regulatory Elements ◽  
Female Germline ◽  
Maternal Gene

The formation of a diploid zygote is a highly complex cellular process that is entirely controlled by maternal gene products stored in the egg cytoplasm. This highly specialized transcriptional program is tightly controlled at the chromatin level in the female germline. As an extreme case in point, the massive and specific ovarian expression of the essential thioredoxin Deadhead (DHD) is critically regulated in Drosophila by the histone demethylase Lid and its partner, the histone deacetylase complex Sin3A/Rpd3, via yet unknown mechanisms. Here, we identified Snr1 and Mod(mdg4) as essential for dhd expression and investigated how these epigenomic effectors act with Lid and Sin3A to hyperactivate dhd. Using Cut&Run chromatin profiling with a dedicated data analysis procedure, we found that dhd is intriguingly embedded in an H3K27me3/H3K9me3-enriched mini-domain flanked by DNA regulatory elements, including a dhd promoter-proximal element essential for its expression. Surprisingly, Lid, Sin3a, Snr1 and Mod(mdg4) impact H3K27me3 and this regulatory element in distinct manners. However, we show that these effectors activate dhd independently of H3K27me3/H3K9me3, and that dhd remains silent in the absence of these marks. Together, our study demonstrates an atypical and critical role for chromatin regulators Lid, Sin3A, Snr1 and Mod(mdg4) to trigger tissue-specific hyperactivation within a unique heterochromatin mini-domain.

High resolution mapping of nucleic acid containing complexes in vitro and in situ

1996 ◽  
Vol 54 ◽  
pp. 48-49
Author(s):  
D. P. Bazett-Jones ◽  
M. J. Hendzel
Keyword(s):  
Nucleic Acid ◽  
High Resolution ◽  
Transcription Initiation ◽  
Molecular Mechanisms ◽  
Heavy Atom ◽  
Regulation Of Transcription ◽  
High Exposure ◽  
Resolution Mapping ◽  
Tata Sequence

Structural analysis of combinations of nucleosomes and transcription factors on promoter and enhancer elements is necessary in order to understand the molecular mechanisms responsible for the regulation of transcription initiation. Such complexes are often not amenable to study by high resolution crystallographic techniques. We have been applying electron spectroscopic imaging (ESI) to specific problems in molecular biology related to transcription regulation. There are several advantages that this technique offers in studies of nucleoprotein complexes. First, an intermediate level of spatial resolution can be achieved because heavy atom contrast agents are not necessary. Second, mass and stoichiometric relationships of protein and nucleic acid can be estimated by phosphorus detection, an element in much higher proportions in nucleic acid than protein. Third, wrapping or bending of the DNA by the protein constituents can be observed by phosphorus mapping of the complexes. Even when ESI is used with high exposure of electrons to the specimen, important macromolecular information may be provided. For example, an image of the TATA binding protein (TBP) bound to DNA is shown in the Figure (top panel). It can be seen that the protein distorts the DNA away from itself and much of its mass sits off the DNA helix axis. Moreover, phosphorus and mass estimates demonstrate whether one or two TBP molecules interact with this particular promoter TATA sequence.

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