Insulation of gene expression by CTCF and cohesin-based subTAD (ubiquitous intra-TAD) loop structures in mouse liver

Computational prediction of CTCF/cohesin-based intra-TAD loops that insulate chromatin contacts and gene expression in mouse liver

eLife ◽

10.7554/elife.34077 ◽

2018 ◽

Vol 7 ◽

Cited By ~ 21

Author(s):

Bryan J Matthews ◽

David J Waxman

Keyword(s):

Nuclear Organization ◽

Computational Prediction ◽

Computational Method ◽

Regulatory Control ◽

Ctcf Binding ◽

Distal Enhancer ◽

Enhancer Activity ◽

Repressive Histone Mark ◽

Functional Features ◽

High Level

CTCF and cohesin are key drivers of 3D-nuclear organization, anchoring the megabase-scale Topologically Associating Domains (TADs) that segment the genome. Here, we present and validate a computational method to predict cohesin-and-CTCF binding sites that form intra-TAD DNA loops. The intra-TAD loop anchors identified are structurally indistinguishable from TAD anchors regarding binding partners, sequence conservation, and resistance to cohesin knockdown; further, the intra-TAD loops retain key functional features of TADs, including chromatin contact insulation, blockage of repressive histone mark spread, and ubiquity across tissues. We propose that intra-TAD loops form by the same loop extrusion mechanism as the larger TAD loops, and that their shorter length enables finer regulatory control in restricting enhancer-promoter interactions, which enables selective, high-level expression of gene targets of super-enhancers and genes located within repressive nuclear compartments. These findings elucidate the role of intra-TAD cohesin-and-CTCF binding in nuclear organization associated with widespread insulation of distal enhancer activity.

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The role of CTCF in coordinating the expression of single gene loci

Biochemistry and Cell Biology ◽

10.1139/o11-040 ◽

2011 ◽

Vol 89 (5) ◽

pp. 489-494 ◽

Cited By ~ 4

Author(s):

Austin E Gillen ◽

Ann Harris

Keyword(s):

Gene Expression ◽

Mhc Class Ii ◽

Noncoding Rna ◽

Single Gene ◽

Functional Model ◽

Gene Clusters ◽

Ctcf Binding ◽

Chromatin Interactions ◽

Insulator Elements

The CCCTC-binding factor (CTCF), which binds insulator elements in vertebrates, also facilitates coordinated gene expression at several gene clusters, including the β-globin, Igf2/H19 (insulin like growth factor 2/H19 noncoding RNA), and major histocompatibility complex (MHC) class II loci. CTCF controls expression of these genes both by enabling insulator function and facilitating higher order chromatin interactions. While the role of CTCF in gene regulation is best studied at these multi-gene loci, there is also evidence that CTCF contributes to the regulated expression of single genes. Here, we discuss how CTCF participates in coordinating gene expression at the CFTR (cystic fibrosis transmembrane conductance regulator) and IFNG (interferon-gamma) loci. We consider the structural similarities between the loci with regard to CTCF-binding elements, the possible interaction between nuclear receptors and CTCF, and the role of CTCF in chromatin looping at these genes. These comparisons reveal a functional model that may be applicable to other single-gene loci that require CTCF for coordinated gene expression.

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Roles of Elongator Dependent tRNA Modification Pathways in Neurodegeneration and Cancer

Genes ◽

10.3390/genes10010019 ◽

2018 ◽

Vol 10 (1) ◽

pp. 19 ◽

Cited By ~ 18

Author(s):

Harmen Hawer ◽

Alexander Hammermeister ◽

Keerthiraju Ravichandran ◽

Sebastian Glatt ◽

Raffael Schaffrath ◽

...

Keyword(s):

Gene Expression ◽

Human Disease ◽

Messenger Rna ◽

Environmental Changes ◽

Mrna Translation ◽

Regulatory Control ◽

Trna Modification ◽

Positive Role ◽

Trna Modifications

Transfer RNA (tRNA) is subject to a multitude of posttranscriptional modifications which can profoundly impact its functionality as the essential adaptor molecule in messenger RNA (mRNA) translation. Therefore, dynamic regulation of tRNA modification in response to environmental changes can tune the efficiency of gene expression in concert with the emerging epitranscriptomic mRNA regulators. Several of the tRNA modifications are required to prevent human diseases and are particularly important for proper development and generation of neurons. In addition to the positive role of different tRNA modifications in prevention of neurodegeneration, certain cancer types upregulate tRNA modification genes to sustain cancer cell gene expression and metastasis. Multiple associations of defects in genes encoding subunits of the tRNA modifier complex Elongator with human disease highlight the importance of proper anticodon wobble uridine modifications (xm5U34) for health. Elongator functionality requires communication with accessory proteins and dynamic phosphorylation, providing regulatory control of its function. Here, we summarized recent insights into molecular functions of the complex and the role of Elongator dependent tRNA modification in human disease.

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The first enhancer in an enhancer chain safeguards subsequent enhancer-promoter contacts from a distance

Genome Biology ◽

10.1186/s13059-019-1808-y ◽

2019 ◽

Vol 20 (1) ◽

Cited By ~ 5

Author(s):

Wei Song ◽

Roded Sharan ◽

Ivan Ovcharenko

Keyword(s):

Gene Expression ◽

Cell Lines ◽

Target Gene ◽

Regulatory Elements ◽

Distal Enhancer ◽

Human Genes ◽

Multiple Cell ◽

A Chain ◽

Target Promoter

Abstract Background Robustness and evolutionary stability of gene expression in the human genome are established by an array of redundant enhancers. Results Using Hi-C data in multiple cell lines, we report a comprehensive map of promoters and active enhancers connected by chromatin contacts, spanning 9000 enhancer chains in 4 human cell lines associated with 2600 human genes. We find that the first enhancer in a chain that directly contacts the target promoter is commonly located at a greater genomic distance from the promoter than the second enhancer in a chain, 96 kb vs. 45 kb, respectively. The first enhancer also features higher similarity to the promoter in terms of tissue specificity and higher enrichment of loop factors, suggestive of a stable primary contact with the promoter. In contrast, a chain of enhancers which connects to the target promoter through a neutral DNA segment instead of an enhancer is associated with a significant decrease in target gene expression, suggesting an important role of the first enhancer in initiating transcription using the target promoter and bridging the promoter with other regulatory elements in the locus. Conclusions The widespread chained structure of gene enhancers in humans reveals that the primary, critical enhancer is distal, commonly located further away than other enhancers. This first, distal enhancer establishes contacts with multiple regulatory elements and safeguards a complex regulatory program of its target gene.

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Global transcriptional response of Methylorubrum extorquens to formaldehyde stress expands the role of EfgA and is distinct from antibiotic translational inhibition

10.1101/2021.01.07.425672 ◽

2021 ◽

Author(s):

Jannell Bazurto ◽

Siavash Riazi ◽

Simon D'Alton ◽

Daniel E. Deatherage ◽

Eric L. Bruger ◽

...

Keyword(s):

Gene Expression ◽

Carbon Source ◽

Transcriptional Response ◽

Genotoxic Stress ◽

Biological Molecules ◽

Regulatory Control ◽

Translation Inhibition ◽

Translational Inhibition ◽

Potential Carbon

The potency and indiscriminate nature of formaldehyde reactivity upon biological molecules make it a universal stressor. However, some organisms such as Methylorubrum extorquens possess means to rapidly and effectively mitigate formaldehyde-induced damage. EfgA is a recently identified formaldehyde sensor predicted to halt translation in response to elevated formaldehyde as a means to protect cells. Herein, we investigate growth and changes in gene expression to understand how M. extorquens responds to formaldehyde with and without the EfgA-formaldehyde-mediated translational response, and how this mechanism compares to antibiotic-mediated translation inhibition. These distinct mechanisms of translation inhibition have notable differences: they each involve different specific players and in addition, formaldehyde also acts as a general, multi-target stressor and a potential carbon source. We present findings demonstrating that in addition to its characterized impact on translation, functional EfgA allows for a rapid and robust transcriptional response to formaldehyde and that removal of EfgA leads to heightened proteotoxic and genotoxic stress in the presence of increased formaldehyde levels. We also found that many downstream consequences of translation inhibition were shared by EfgA-formaldehyde- and kanamycin-mediated translation inhibition. Our work uncovered additional layers of regulatory control enacted by functional EfgA upon experiencing formaldehyde stress, and further demonstrated the importance this protein plays at both transcriptional and translational levels in this model methylotroph.

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Contribution of CTCF binding to transcriptional activity at the HOXA locus in NPM1-mutant AML cells

10.1101/2020.02.17.952390 ◽

2020 ◽

Author(s):

Reza Ghasemi ◽

Heidi Struthers ◽

Elisabeth R. Wilson ◽

David H. Spencer

Keyword(s):

Gene Expression ◽

Transcriptional Activity ◽

Three Dimensional ◽

Ctcf Binding ◽

Chromatin Domain ◽

Intrinsic Factors ◽

Chromatin Loops ◽

Chromatin Interactions ◽

The Common

AbstractTranscriptional regulation of the HOXA genes is thought to involve CTCF-mediated chromatin loops and the opposing actions of the COMPASS and Polycomb epigenetic complexes. We investigated the role of these mechanisms at the HOXA cluster in AML cells with the common NPM1c mutation, which express both HOXA and HOXB genes. CTCF binding at the HOXA locus is conserved across primary AML samples, regardless of HOXA gene expression, and defines a continuous chromatin domain marked by COMPASS-associated histone H3 trimethylation in NPM1-mutant primary AML samples. Profiling of the three-dimensional chromatin architecture of NPM1-mutant OCI-AML3 cells identified chromatin loops between the active HOXA9-HOXA11 genes and loci in the SNX10 gene and an intergenic region located 1.4Mbp upstream of the HOXA locus. Deletion of CTCF binding sites in OCI-AML3 cells reduced these interactions, but resulted in new, CTCF-independent loops with regions in the SKAP2 gene that were marked by enhancer-associated histone modifications in primary AML samples. HOXA gene expression was maintained in the CTCF deletion mutants, indicating that transcriptional activity at the HOXA locus in NPM1-mutant AML cells does not require long-range CTCF-mediated chromatin interactions, and instead may be driven by intrinsic factors within the HOXA gene cluster.

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Global Transcriptional Response of Methylorubrum extorquens to Formaldehyde Stress Expands the Role of EfgA and Is Distinct from Antibiotic Translational Inhibition

Microorganisms ◽

10.3390/microorganisms9020347 ◽

2021 ◽

Vol 9 (2) ◽

pp. 347

Author(s):

Jannell V. Bazurto ◽

Siavash Riazi ◽

Simon D’Alton ◽

Daniel E. Deatherage ◽

Eric L. Bruger ◽

...

Keyword(s):

Gene Expression ◽

Carbon Source ◽

Transcriptional Response ◽

Genotoxic Stress ◽

Biological Molecules ◽

Regulatory Control ◽

Translation Inhibition ◽

Translational Inhibition ◽

Potential Carbon

The potency and indiscriminate nature of formaldehyde reactivity upon biological molecules make it a universal stressor. However, some organisms such as Methylorubrum extorquens possess means to rapidly and effectively mitigate formaldehyde-induced damage. EfgA is a recently identified formaldehyde sensor predicted to halt translation in response to elevated formaldehyde as a means to protect cells. Herein, we investigate growth and changes in gene expression to understand how M. extorquens responds to formaldehyde with and without the EfgA-formaldehyde-mediated translational response, and how this mechanism compares to antibiotic-mediated translation inhibition. These distinct mechanisms of translation inhibition have notable differences: they each involve different specific players and in addition, formaldehyde also acts as a general, multi-target stressor and a potential carbon source. We present findings demonstrating that in addition to its characterized impact on translation, functional EfgA allows for a rapid and robust transcriptional response to formaldehyde and that removal of EfgA leads to heightened proteotoxic and genotoxic stress in the presence of increased formaldehyde levels. We also found that many downstream consequences of translation inhibition were shared by EfgA-formaldehyde- and kanamycin-mediated translation inhibition. Our work uncovered additional layers of regulatory control enacted by functional EfgA upon experiencing formaldehyde stress, and further demonstrated the importance this protein plays at both transcriptional and translational levels in this model methylotroph.

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CTCF-mediated Genomic Effects of BART Region on Epstein-Barr Virus Chromatin 3D Structure in Gastric Carcinoma Cells

10.1101/2020.12.03.409722 ◽

2020 ◽

Author(s):

Kyoung-Dong Kim ◽

Subin Cho ◽

Taelyn Kim ◽

Sora Huh ◽

Lina Kim ◽

...

Keyword(s):

Gene Expression ◽

Gastric Carcinoma ◽

Genome Structure ◽

Viral Gene Expression ◽

Ctcf Binding ◽

Viral Gene ◽

3D Genome ◽

Barr Virus ◽

Epstein Barr

AbstractEBV latent infection in gastric carcinoma (GC) cells is characterized by distinct viral gene expression programs. CCCTC-binding factor (CTCF) is a chromatin structural factor that has been involved in coordinated chromatin interactions between multiple loci of Epstein-Barr virus (EBV) genes. Here, we investigate the role of CTCF in regulating EBV gene expression and chromosome conformation in model of EBV-associated gastric carcinoma (EBVaGC). Chromatin immunoprecipitation followed by sequencing (ChIP-seq) against CTCF revealed 16 CTCF binding sites (BS) in EBV genome of EBVaGC, SNU719 cells. Among the CTCF BSs, one site named as BARTp (BamHI A right transcript promoter) CTCF BS is located at upstream of 11.8-kb BART region (EBV genome: 139724-151554) and was not yet defined its biological functions in EBV life cycle. EBV BART encodes a complex miRNA cluster of highly spliced transcripts that is implicated in EBV cancer pathogenesis. This present study investigated the functional role of the CTCF binding site at BARTp (BARTp CTCF BS) in regulating EBV gene transcription and EBV three-dimensional (3D) genome structure as DNA loop maker. Circular chromatin confirmation capture (4C)-seq and chromatin confirmation capture (3C)-semi-quantitative(sq)PCR assays using SNU719 cells revealed that BARTp CTCF BS interacts with CTCF BSs of LMP1/2, Cp/OriP, and Qp in EBV genome. We generated mutations in BARTp CTCF BS (S13) in bacmids with (BART+) or without (BART−) the 11.8-kb BART transcript unit (B(+/−)). ChIP-qPCR assay demonstrated that CTCF binding was ablated from BARTp in EBV B(+/−) S13− genomes (mutant S13), elevated at several other sites such as LMP1, OriP, and Cp in EBV B(-) (BART−) S13− genome, and decreased at the same sites in EBV B(+) S13− genome. Infection assay showed that BARTp CTCF BS mutation reduced infectivity, while BART transcript deletion has no detectable effects. Gene expression tests showed that EBNA1 was highly downregulated in B(+/−) S13− EBVs related to B(+/−) S13+ EBVs (wild-type S13). LMP1 and BZLF1 were more downregulated in B(-) S13− EBV than B(+) S13− EBV. Taken together, these findings suggest that the CTCF binding and BART region contribute to EBV 3D genome structure via a cluster of DNA loops formed by BARTp CTCF BS (S13) and are important for coordinated viral gene expression and EBV infectivity.

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Role of small nuclear RNAs in eukaryotic gene expression

Essays in Biochemistry ◽

10.1042/bse0540079 ◽

2013 ◽

Vol 54 ◽

pp. 79-90 ◽

Cited By ~ 34

Author(s):

Saba Valadkhan ◽

Lalith S. Gunawardane

Keyword(s):

Gene Expression ◽

Protein Interactions ◽

Rna Stability ◽

Splice Sites ◽

Branch Site ◽

Small Nuclear Rnas ◽

Eukaryotic Gene Expression ◽

Eukaryotic Gene ◽

Rna Biogenesis

Eukaryotic cells contain small, highly abundant, nuclear-localized non-coding RNAs [snRNAs (small nuclear RNAs)] which play important roles in splicing of introns from primary genomic transcripts. Through a combination of RNA–RNA and RNA–protein interactions, two of the snRNPs, U1 and U2, recognize the splice sites and the branch site of introns. A complex remodelling of RNA–RNA and protein-based interactions follows, resulting in the assembly of catalytically competent spliceosomes, in which the snRNAs and their bound proteins play central roles. This process involves formation of extensive base-pairing interactions between U2 and U6, U6 and the 5′ splice site, and U5 and the exonic sequences immediately adjacent to the 5′ and 3′ splice sites. Thus RNA–RNA interactions involving U2, U5 and U6 help position the reacting groups of the first and second steps of splicing. In addition, U6 is also thought to participate in formation of the spliceosomal active site. Furthermore, emerging evidence suggests additional roles for snRNAs in regulation of various aspects of RNA biogenesis, from transcription to polyadenylation and RNA stability. These snRNP-mediated regulatory roles probably serve to ensure the co-ordination of the different processes involved in biogenesis of RNAs and point to the central importance of snRNAs in eukaryotic gene expression.

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Post-Transcriptional Regulation of Cardiac Sarcomere Protein Titin Through 3’ Untranslated Regions

Vanderbilt Undergraduate Research Journal ◽

10.15695/vurj.v9i0.3772 ◽

2013 ◽

Vol 9 ◽

Cited By ~ 1

Author(s):

Tara A Shrout

Keyword(s):

Gene Expression ◽

Transcriptional Control ◽

Striated Muscle ◽

Binding Capacity ◽

Structural Features ◽

Neonatal Rat ◽

Regulatory Control ◽

Untranslated Regions ◽

Luciferase Reporter ◽

Regulatory Effects

Titin is the largest known protein in the human body, and forms the backbone of all striated muscle sarcomeres. The elastic nature of titin is an important component of muscle compliance and functionality. A significant amount of energy is expended to synthesize titin, thus we postulate that titin gene expression is under strict regulatory control in order to conserve cellular resources. In general, gene expression is mediated in part by post-transcriptional control elements located within the 5’ and 3’ untranslated regions (UTRs) of mature mRNA. The 3’UTR in particular contains structural features that affect binding capacity to other RNA components, such as MicroRNA, which control mRNA localization, translation, and degradation. The degree and significance of the regulatory effects mediated by two determined variants of titin’s 3’ UTR were evaluated in Neonatal Rat Ventricular Myocyte and Human Embryonic Kidney cell lines. Recombinant plasmids to transfect these cells lines were engineered by insertion of the variant titin 3’UTR 431- and 1047-base pairs sequences into luciferase reporter vectors. Expression due to an unaltered reporter vector served as the control. Quantitative changes in luciferase activity due to the recombinants proportionally represented the effect titin’s respective 3’UTR conferred on downstream post-transcriptional expression relative to the control. The effect due to titin’s shorter 3’UTR sequence was inconclusive; however, results illustrated that titin’s longer 3’UTR sequence caused a 35 percent decrease in protein expression. Secondary structural analysis of the two sequences revealed differential folding patterns that affect the stability and degree of MicroRNA-binding within titin’s variant 3’UTR sequences.

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