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Influence of Repressive Histone and DNA Methylation upon D4Z4 Transcription in Non-Myogenic Cells
Influence of Repressive Histone and DNA Methylation upon D4Z4 Transcription in Non-Myogenic Cells
We looked at a disease-associated macrosatellite array D4Z4 and focused on epigenetic factors influencing its chromatin state outside of the disease-context. We used the HCT116 cell line that contains the non-canonical polyadenylation (poly-A) signal required to stabilize somatic transcripts of the human double homeobox gene DUX4, encoded from D4Z4. In HCT116, D4Z4 is packaged into constitutive heterochromatin, characterized by DNA methylation and histone H3 tri-methylation at lysine 9 (H3K9me3), resulting in low basal levels of D4Z4-derived transcripts. However, a double knockout (DKO) of DNA methyltransferase genes, DNMT1 and DNMT3B, but not either alone, results in significant loss of DNA and H3K9 methylation. This is coupled with upregulation of transcript levels from the array, including DUX4 isoforms (DUX4-fl) that are abnormally expressed in somatic muscle in the disease Facioscapulohumeral muscular dystrophy (FSHD) along with DUX4 protein, as indicated indirectly by upregulation of bondafide targets of DUX4 in DKO but not HCT116 cells. Results from treatment with a chemical inhibitor of histone methylation in HCT116 suggest that in the absence of DNA hypomethylation, H3K9me3 loss alone is sufficient to facilitate DUX4-fl transcription. Additionally, characterization of a cell line from a patient with Immunodeficiency, Centromeric instability and Facial anomalies syndrome 1 (ICF1) possessing a non-canonical poly-A signal and DNA hypomethylation at D4Z4 showed DUX4 target gene upregulation in the patient when compared to controls in spite of retention of H3K9me3. Taken together, these data suggest that both DNA methylation and H3K9me3 are determinants of D4Z4 silencing. Moreover, we show that in addition to testis, there is appreciable expression of spliced and polyadenylated D4Z4 derived transcripts that contain the complete DUX4 open reading frame (ORF) along with DUX4 target gene expression in the thymus, suggesting that DUX4 may provide normal function in this somatic tissue., Keywords: dux4 gene, facioscapulohumeral muscular-dystrophy, human cancer-cells, human genome, immunodeficiency syndrome, inactive x-chromosome, macrosatellite dxz4, methyltransferase gene, muscle-cells, repeated elements, Publication Note: The publisher’s version of record is available at https://doi.org/10.1371/journal.pone.0160022
Influence of Repressive Histone and DNA Methylation upon D4Z4 Transcription in Non-Myogenic Cells.
Influence of Repressive Histone and DNA Methylation upon D4Z4 Transcription in Non-Myogenic Cells.
We looked at a disease-associated macrosatellite array D4Z4 and focused on epigenetic factors influencing its chromatin state outside of the disease-context. We used the HCT116 cell line that contains the non-canonical polyadenylation (poly-A) signal required to stabilize somatic transcripts of the human double homeobox gene DUX4, encoded from D4Z4. In HCT116, D4Z4 is packaged into constitutive heterochromatin, characterized by DNA methylation and histone H3 tri-methylation at lysine 9 (H3K9me3), resulting in low basal levels of D4Z4-derived transcripts. However, a double knockout (DKO) of DNA methyltransferase genes, DNMT1 and DNMT3B, but not either alone, results in significant loss of DNA and H3K9 methylation. This is coupled with upregulation of transcript levels from the array, including DUX4 isoforms (DUX4-fl) that are abnormally expressed in somatic muscle in the disease Facioscapulohumeral muscular dystrophy (FSHD) along with DUX4 protein, as indicated indirectly by upregulation of bondafide targets of DUX4 in DKO but not HCT116 cells. Results from treatment with a chemical inhibitor of histone methylation in HCT116 suggest that in the absence of DNA hypomethylation, H3K9me3 loss alone is sufficient to facilitate DUX4-fl transcription. Additionally, characterization of a cell line from a patient with Immunodeficiency, Centromeric instability and Facial anomalies syndrome 1 (ICF1) possessing a non-canonical poly-A signal and DNA hypomethylation at D4Z4 showed DUX4 target gene upregulation in the patient when compared to controls in spite of retention of H3K9me3. Taken together, these data suggest that both DNA methylation and H3K9me3 are determinants of D4Z4 silencing. Moreover, we show that in addition to testis, there is appreciable expression of spliced and polyadenylated D4Z4 derived transcripts that contain the complete DUX4 open reading frame (ORF) along with DUX4 target gene expression in the thymus, suggesting that DUX4 may provide normal function in this somatic tissue., Publication Note: This NIH-funded author manuscript originally appeared in PubMed Central at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4965136.
Insect Herbivores Increase The Spatial Aggregation Of A Clonal Plant
Insect Herbivores Increase The Spatial Aggregation Of A Clonal Plant
Premise of research.In a plant population, the pattern of spatial dispersion from uniform to strongly aggregated can influence the intensity of ecological interactions, including those between plants and their consumers, such as herbivorous insects. Whether insect herbivores also contribute to the generation of dispersion patterns within plant populations, however, is not well understood. We investigated how insect herbivory and plant density interact to influence the degree of aggregation of stems in experimental plots of the clonal perennial herb Solanum carolinense, a noxious weed in the southeastern United States and invasive in other parts of the world.Methodology.We planted S. carolinense in 40 experimental plots at five densities and maintained each plot at one of two herbivory levels. Stems in every plot were initially planted in a uniform dispersion pattern. For 3 yr, we quantified the dispersion of clonal recruits in each plot at six spatial scales.Pivotal results.At larger spatial scales, we found significant interactions between plant density and insect herbivory. In plots with herbivores, spatial aggregation increased as plant density decreased, but this relationship was weak or nonexistent in plots where herbivores were excluded (herbivore-excluded plots exhibited random dispersion of stems across all densities). Solanum carolinense in plots exposed to herbivores were on average twice as aggregated as those that had herbivores excluded.Conclusions.These results suggest that insect herbivores can influence clonal growth or ramet survival in ways that increase population-level aggregation of S. carolinense. Our findings also imply that S. carolinense populations in its invasive range may have increased spatial spread and decreased intraspecific competition when herbivory is low. We discuss the potential for specific types of feedbacks between insect damage and plant dispersion, highlighting important targets for future research., Keywords: patterns, solidago-altissima, Solanum carolinense, solanum-carolinense, seed dispersal, biological-control, carex-arenaria, density effects, dispersion, herbivory, old-field, patch size, physiological integration, population dynamics, Ripley's K, trientalis-europaea l, Publication Note: The publisher's version of record is available at https://doi.org/10.1086/696685
Integrating Top-down And Bottom-up Effects Of Local Density Across Scales And A Complex Life Cycle
Integrating Top-down And Bottom-up Effects Of Local Density Across Scales And A Complex Life Cycle
Effects of group size (local conspecific density) on individual performance can be substantial, yet it is unclear how these translate to larger-scale and longer-term outcomes. Effects of group size can be mediated by both top-down and bottom-up interactions, can change in type or direction across the life cycle, and can depend on the spatial scale at which group size is assessed. Only by determining how these different processes combine can we make predictions about how selection operates on group size or link hierarchical patterns of density dependence with population dynamics. We manipulated the density of a leaf beetle,Leptinotarsa juncta, at three nested spatial scales (patch, plant within a patch, and leaf within plant) to investigate how conspecific density affects predator-mediated survival and resource-mediated growth during different life stages and across multiple spatial scales. We then used data from field predation experiments to assess howL. junctadensities at hierarchical scales affect different aspects of predation. Finally, we incorporated predator- and resource-mediated effects of density in a model to explore how changes in group size due to density-dependent predation might affect mass at pupation for survivors. The effects ofL. junctadensity on predation risk differed among scales. Per capita predation risk of both eggs and late instars was lowest at high patch-scale densities, but increased with plant-scale density. The final mass of late instars declined with increasing plant-scale larval density, potentially because of truncated development of high-density larvae. Predation incidence (i.e., group attack rate) increased with larval density at all spatial scales. A high coefficient of variation (i.e., greater aggregation) ofL. junctadensity was associated with lower predation incidence at some scales. Our model suggested that predator- and resource-mediated effects of density interact: lower survival at high larval density is mitigated by high final mass of larvae in the resulting smaller groups. Our results emphasize the importance of spatial scale and demonstrate that effects of top-down and bottom-up interactions are not necessarily independent. To understand how group size influences fitness, predator- and resource-mediated effects of density should be measured in their demographic and spatial context, and not in isolation., mortality, adult size, bottom-up interactions, density dependence, dependence, foraging behavior, group-size, hierarchical predator foraging, insect herbivores, larval aggregation, Leptinotarsajuncta, local density, optimal group size, predation risk, prey, spatial scale, spatial scale, top-down interactions, The publisher's version of record is availible at https://doi.org/10.1002/ecy.3118
Integrative Encode Resource For Cancer Genomics
Integrative Encode Resource For Cancer Genomics
ENCODE comprises thousands of functional genomics datasets, and the encyclopedia covers hundreds of cell types, providing a universal annotation for genome interpretation. However, for particular applications, it may be advantageous to use a customized annotation. Here, we develop such a custom annotation by leveraging advanced assays, such as eCLIP, Hi-C, and whole-genome STARR-seq on a number of data-rich ENCODE cell types. A key aspect of this annotation is comprehensive and experimentally derived networks of both transcription factors and RNA-binding proteins (TFs and RBPs). Cancer, a disease of system-wide dysregulation, is an ideal application for such a network-based annotation. Specifically, for cancer-associated cell types, we put regulators into hierarchies and measure their network change (rewiring) during oncogenesis. We also extensively survey TF-RBP crosstalk, highlighting how SUB1, a previously uncharacterized RBP, drives aberrant tumor expression and amplifies the effect of MYC, a well-known oncogenic TF. Furthermore, we show how our annotation allows us to place oncogenic transformations in the context of a broad cell space; here, many normal-to-tumor transitions move towards a stem-like state, while oncogene knockdowns show an opposing trend. Finally, we organize the resource into a coherent workflow to prioritize key elements and variants, in addition to regulators. We showcase the application of this prioritization to somatic burdening, cancer differential expression and GWAS. Targeted validations of the prioritized regulators, elements and variants using siRNA knockdowns, CRISPR-based editing, and luciferase assays demonstrate the value of the ENCODE resource. ENCODE is a resource comprising thousands of functional genomic datasets. Here, the authors present custom annotation within ENCODE for cancer, highlighting a workflow that can help prioritise key elements in oncogenesis., landscape, cells, dysregulation, genes, binding, network analysis, repair, somatic mutations, transcription factors, tumor heterogeneity, The publisher's version of record is availible at https://doi.org/10.1038/s41467-020-14743-w
Integrin-mediated Traction Force Enhances Paxillin Molecular Associations And Adhesion Dynamics That Increase The Invasiveness Of Tumor Cells Into A Three-dimensional Extracellular Matrix
Integrin-mediated Traction Force Enhances Paxillin Molecular Associations And Adhesion Dynamics That Increase The Invasiveness Of Tumor Cells Into A Three-dimensional Extracellular Matrix
Metastasis requires tumor cells to navigate through a stiff stroma and squeeze through confined microenvironments. Whether tumors exploit unique biophysical properties to metastasize remains unclear. Data show that invading mammary tumor cells, when cultured in a stiffened three-dimensional extracellular matrix that recapitulates the primary tumor stroma, adopt a basal-like phenotype. Metastatic tumor cells and basal-like tumor cells exert higher integrin-mediated traction forces at the bulk and molecular levels, consistent with a motor-clutch model in which motors and clutches are both increased. Basal-like nonmalignant mammary epithelial cells also display an altered integrin adhesion molecular organization at the nanoscale and recruit a suite of paxillin-associated proteins implicated in invasion and metastasis. Phosphorylation of paxillin by Src family kinases, which regulates adhesion turnover, is similarly enhanced in the metastatic and basal-like tumor cells, fostered by a stiff matrix, and critical for tumor cell invasion in our assays. Bioinformatics reveals an unappreciated relationship between Src kinases, paxillin, and survival of breast cancer patients. Thus adoption of the basal-like adhesion phenotype may favor the recruitment of molecules that facilitate tumor metastasis to integrin-based adhesions. Analysis of the physical properties of tumor cells and integrin adhesion composition in biopsies may be predictive of patient outcome., Keywords: angle interference microscopy, breast-cancer metastasis, epithelial-mesenchymal transition, focal adhesions, growth-factor-beta, hybrid epithelial/mesenchymal phenotype, linked kinase ilk, malignant mammary epithelium, nuclear-deformation, transgenic mouse model, Publication Note: The publisher's version of record is available at https://doi.org/10.1091/mbc.E16-09-0654
Integrin-mediated traction force enhances paxillin molecular associations and adhesion dynamics that increase the invasiveness of tumor cells into a three-dimensional extracellular matrix.
Integrin-mediated traction force enhances paxillin molecular associations and adhesion dynamics that increase the invasiveness of tumor cells into a three-dimensional extracellular matrix.
Metastasis requires tumor cells to navigate through a stiff stroma and squeeze through confined microenvironments. Whether tumors exploit unique biophysical properties to metastasize remains unclear. Data show that invading mammary tumor cells, when cultured in a stiffened three-dimensional extracellular matrix that recapitulates the primary tumor stroma, adopt a basal-like phenotype. Metastatic tumor cells and basal-like tumor cells exert higher integrin-mediated traction forces at the bulk and molecular levels, consistent with a motor-clutch model in which motors and clutches are both increased. Basal-like nonmalignant mammary epithelial cells also display an altered integrin adhesion molecular organization at the nanoscale and recruit a suite of paxillin-associated proteins implicated in invasion and metastasis. Phosphorylation of paxillin by Src family kinases, which regulates adhesion turnover, is similarly enhanced in the metastatic and basal-like tumor cells, fostered by a stiff matrix, and critical for tumor cell invasion in our assays. Bioinformatics reveals an unappreciated relationship between Src kinases, paxillin, and survival of breast cancer patients. Thus adoption of the basal-like adhesion phenotype may favor the recruitment of molecules that facilitate tumor metastasis to integrin-based adhesions. Analysis of the physical properties of tumor cells and integrin adhesion composition in biopsies may be predictive of patient outcome., Grant Number: R01 CA192914, U01 CA202241, U54 CA210184, R01 GM112998, U54 CA210190, T32 CA108462, R01 CA174929, R01 CA172986, R01 HL082792, Publication Note: This NIH-funded author manuscript originally appeared in PubMed Central at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5449147.
Interactive Effects Of Precipitation And Nitrogen Enrichment On Multi-trophic Dynamics In Plant-arthropod Communities
Interactive Effects Of Precipitation And Nitrogen Enrichment On Multi-trophic Dynamics In Plant-arthropod Communities
Patterns of precipitation and nitrogen (N) deposition are changing in ecosystems worldwide. Simultaneous increases in precipitation and N deposition can relieve co-limiting soil resource conditions for plants and result in synergistic plant responses, which may affect animals and plant responses to higher trophic levels. However, the potential for synergistic effects of precipitation and N deposition on animals and plant responses to herbivores and predators (via trophic cascades) is unclear. We investigated the influence of precipitation and N enrichment on ecological dynamics across three trophic levels, hypothesizing that herbivores and plants would exhibit synergistic responses to the combined influence of precipitation, N amendments and predators. To test this, we conducted a field experiment with arthropods on two model plant species, Nicotiana tabacum and Nicotiana rustica. First, we characterized the plant-arthropod assemblages, finding that N.tabacum hosted greater abundances of caterpillars, while N. rustica hosted more sap-sucking herbivores. Next, we evaluated the effects of rainwater, soil N, and predatory spider manipulations for both plantarthropod assemblages. On N. tabacum, water and N availability had an interactive effect on caterpillars, where caterpillars were most abundant with rainwater additions and least abundant when both rainwater and N were added. For N. rustica, foliar chemistry had a synergistic response to all three experimental factors. Compared to spider-absent conditions, leaf N concentration increased and C/N decreased when spiders were present, but this response only occurred under high water and N availability. Spiders indirectly altered plant chemistry via a facilitative effect of spiders on sap-sucking herbivores, potentially due to intra-guild predation, and a positive effect of sap-suckers on foliar N concentration. Our study suggests that predictions of the ecological impacts of altered precipitation and N deposition may need to account for the effects of resource co-limitation on dynamics across trophic levels., Keywords: terrestrial ecosystems, fresh-water, bottom-up, top-down, food-web, herbivore interactions, nicotiana-attenuata, nutrient enrichment, oxyopes-salticus araneae, striped lynx spider, Publication Note: The publisher’s version of record is available at https://doi.org/10.1371/journal.pone.0201219
Interactive effects of precipitation and nitrogen enrichment on multi-trophic dynamics in plant-arthropod communities.
Interactive effects of precipitation and nitrogen enrichment on multi-trophic dynamics in plant-arthropod communities.
Patterns of precipitation and nitrogen (N) deposition are changing in ecosystems worldwide. Simultaneous increases in precipitation and N deposition can relieve co-limiting soil resource conditions for plants and result in synergistic plant responses, which may affect animals and plant responses to higher trophic levels. However, the potential for synergistic effects of precipitation and N deposition on animals and plant responses to herbivores and predators (via trophic cascades) is unclear. We investigated the influence of precipitation and N enrichment on ecological dynamics across three trophic levels, hypothesizing that herbivores and plants would exhibit synergistic responses to the combined influence of precipitation, N amendments and predators. To test this, we conducted a field experiment with arthropods on two model plant species, Nicotiana tabacum and Nicotiana rustica. First, we characterized the plant-arthropod assemblages, finding that N. tabacum hosted greater abundances of caterpillars, while N. rustica hosted more sap-sucking herbivores. Next, we evaluated the effects of rainwater, soil N, and predatory spider manipulations for both plant-arthropod assemblages. On N. tabacum, water and N availability had an interactive effect on caterpillars, where caterpillars were most abundant with rainwater additions and least abundant when both rainwater and N were added. For N. rustica, foliar chemistry had a synergistic response to all three experimental factors. Compared to spider-absent conditions, leaf N concentration increased and C/N decreased when spiders were present, but this response only occurred under high water and N availability. Spiders indirectly altered plant chemistry via a facilitative effect of spiders on sap-sucking herbivores, potentially due to intra-guild predation, and a positive effect of sap-suckers on foliar N concentration. Our study suggests that predictions of the ecological impacts of altered precipitation and N deposition may need to account for the effects of resource co-limitation on dynamics across trophic levels., Publication Note: This NIH-funded author manuscript originally appeared in PubMed Central at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6072000.
Iseg
Iseg
Background: Identification of functional elements of a genome often requires dividing a sequence of measurements along a genome into segments where adjacent segments have different properties, such as different mean values. Despite dozens of algorithms developed to address this problem in genomics research, methods with improved accuracy and speed are still needed to effectively tackle both existing and emerging genomic and epigenomic segmentation problems. Results: We designed an efficient algorithm, called iSeg, for segmentation of genomic and epigenomic profiles. iSeg first utilizes dynamic programming to identify candidate segments and test for significance. It then uses a novel data structure based on two coupled balanced binary trees to detect overlapping significant segments and update them simultaneously during searching and refinement stages. Refinement and merging of significant segments are performed at the end to generate the final set of segments. By using an objective function based on the p-values of the segments, the algorithm can serve as a general computational framework to be combined with different assumptions on the distributions of the data. As a general segmentation method, it can segment different types of genomic and epigenomic data, such as DNA copy number variation, nucleosome occupancy, nuclease sensitivity, and differential nuclease sensitivity data. Using simple t-tests to compute p-values across multiple datasets of different types, we evaluate iSeg using both simulated and experimental datasets and show that it performs satisfactorily when compared with some other popular methods, which often employ more sophisticated statistical models. Implemented in C++, iSeg is also very computationally efficient, well suited for large numbers of input profiles and data with very long sequences. Conclusions: We have developed an efficient general-purpose segmentation tool and showed that it had comparable or more accurate results than many of the most popular segment-calling algorithms used in contemporary genomic data analysis. iSeg is capable of analyzing datasets that have both positive and negative values. Tunable parameters allow users to readily adjust the statistical stringency to best match the biological nature of individual datasets, including widely or sparsely mapped genomic datasets or those with non-normal distributions., Keywords: gene-expression, identification, design, high-resolution, array cgh data, chromatin reveals, circular binary segmentation, dna copy number, hidden markov model, maize genome, Publication Note: The publisher’s version of record is available at https://doi.org/10.1186/s12859-018-2140-3
Kaposi's Sarcoma-Associated Herpesvirus Inhibitor of cGAS (KicGAS), Encoded by ORF52, Is an Abundant Tegument Protein and Is Required for Production of Infectious Progeny Viruses.
Kaposi's Sarcoma-Associated Herpesvirus Inhibitor of cGAS (KicGAS), Encoded by ORF52, Is an Abundant Tegument Protein and Is Required for Production of Infectious Progeny Viruses.
Although Kaposi's sarcoma-associated herpesvirus (KSHV) ORF52 (also known as KSHV inhibitor of cGAS [KicGAS]) has been detected in purified virions, the roles of this protein during KSHV replication have not been characterized. Using specific monoclonal antibodies, we revealed that ORF52 displays true late gene expression kinetics and confirmed its cytoplasmic localization in both transfected and KSHV-infected cells. We demonstrated that ORF52 comigrates with other known virion proteins following sucrose gradient centrifugation. We also determined that ORF52 resides inside the viral envelope and remains partially associated with capsid when extracellular virions are treated with various detergents and/or salts. There results indicate that ORF52 is a tegument protein abundantly present in extracellular virions. To characterize the roles of ORF52 in the KSHV life cycle, we engineered a recombinant KSHV ORF52-null mutant virus and found that loss of ORF52 results in reduced virion production and a further defect in infectivity. Upon analysis of the virion composition of ORF52-null viral particles, we observed a decrease in the incorporation of ORF45, as well as other tegument proteins, suggesting that ORF52 is important for the packaging of other virion proteins. In summary, our results indicate that, in addition to its immune evasion function, KSHV ORF52 is required for the optimal production of infectious virions, likely due to its roles in virion assembly as a tegument protein. The tegument proteins of herpesviruses, including Kaposi's sarcoma-associated herpesvirus (KSHV), play key roles in the viral life cycle. Each of the three subfamilies of herpesviruses (alpha, beta, and gamma) encode unique tegument proteins with specialized functions. We recently found that one such gammaherpesvirus-specific protein, ORF52, has an important role in immune evasion during KSHV primary infection, through inhibition of the host cytosolic DNA sensing pathway. In this report, we further characterize ORF52 as a tegument protein with vital roles during KSHV lytic replication. We found that ORF52 is important for the production of infectious viral particles, likely through its role in virus assembly, a critical process for KSHV replication and pathogenesis. More comprehensive investigation of the functions of tegument proteins and their roles in viral replication may reveal novel targets for therapeutic interventions against KSHV-associated diseases., Grant Number: F31 CA183250, R01 DE016680, Publication Note: This NIH-funded author manuscript originally appeared in PubMed Central at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4934757.
Label-Free Relative Quantitation of Isobaric and Isomeric Human Histone H2A and H2B Variants by Fourier Transform Ion Cyclotron Resonance Top-Down MS/MS.
Label-Free Relative Quantitation of Isobaric and Isomeric Human Histone H2A and H2B Variants by Fourier Transform Ion Cyclotron Resonance Top-Down MS/MS.
Histone variants are known to play a central role in genome regulation and maintenance. However, many variants are inaccessible by antibody-based methods or bottom-up tandem mass spectrometry due to their highly similar sequences. For many, the only tractable approach is with intact protein top-down tandem mass spectrometry. Here, ultra-high-resolution FT-ICR MS and MS/MS yield quantitative relative abundances of all detected HeLa H2A and H2B isobaric and isomeric variants with a label-free approach. We extend the analysis to identify and relatively quantitate 16 proteoforms from 12 sequence variants of histone H2A and 10 proteoforms of histone H2B from three other cell lines: human embryonic stem cells (WA09), U937, and a prostate cancer cell line LaZ. The top-down MS/MS approach provides a path forward for more extensive elucidation of the biological role of many previously unstudied histone variants and post-translational modifications., Keywords: FT-ICR, FTMS, Histone, Post-translational modification, Proteoform, Sequence variants, Grant Number: P01 GM085354, Publication Note: This NIH-funded author manuscript originally appeared in PubMed Central at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6261780.
Lineage-specific dynamic and pre-established enhancer-promoter contacts cooperate in terminal differentiation.
Lineage-specific dynamic and pre-established enhancer-promoter contacts cooperate in terminal differentiation.
Chromosome conformation is an important feature of metazoan gene regulation; however, enhancer-promoter contact remodeling during cellular differentiation remains poorly understood. To address this, genome-wide promoter capture Hi-C (CHi-C) was performed during epidermal differentiation. Two classes of enhancer-promoter contacts associated with differentiation-induced genes were identified. The first class ('gained') increased in contact strength during differentiation in concert with enhancer acquisition of the H3K27ac activation mark. The second class ('stable') were pre-established in undifferentiated cells, with enhancers constitutively marked by H3K27ac. The stable class was associated with the canonical conformation regulator cohesin, whereas the gained class was not, implying distinct mechanisms of contact formation and regulation. Analysis of stable enhancers identified a new, essential role for a constitutively expressed, lineage-restricted ETS-family transcription factor, EHF, in epidermal differentiation. Furthermore, neither class of contacts was observed in pluripotent cells, suggesting that lineage-specific chromatin structure is established in tissue progenitor cells and is further remodeled in terminal differentiation., Grant Number: T32 GM007276, T32 HG000044, R01 AR045192, U01 HG007919, R01 HL122887, Publication Note: This NIH-funded author manuscript originally appeared in PubMed Central at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5715812.
Long-range Enhancer Interactions Are Prevalent In Mouse Embryonic Stem Cells And Are Reorganized Upon Pluripotent State Transition
Long-range Enhancer Interactions Are Prevalent In Mouse Embryonic Stem Cells And Are Reorganized Upon Pluripotent State Transition
Transcriptional enhancers, including super-enhancers (SEs), form physical interactions with promoters to regulate cell-type-specific gene expression. SEs are characterized by high transcription factor occupancy and large domains of active chromatin, and they are commonly assigned to target promoters using computational predictions. How promoter-SE interactions change upon cell state transitions, and whether transcription factors maintain SE interactions, have not been reported. Here, we used promoter-capture Hi-C to identify promoters that interact with SEs in mouse embryonic stem cells (ESCs). We found that SEs form complex, spatial networks in which individual SEs contact multiple promoters, and a rewiring of promoter-SE interactions occurs between pluripotent states. We also show that long-range promoter-SE interactions are more prevalent in ESCs than in epiblast stem cells (EpiSCs) or Nanog-deficient ESCs. We conclude that SEs form cell-type-specific interaction networks that are partly dependent on core transcription factors, thereby providing insights into the gene regulatory organization of pluripotent cells., Keywords: genes, super-enhancers, differentiation, genome, self-renewal, capture hi-c, chromatin interactions, looping interactions, nanog locus, primed pluripotency, Publication Note: The publisher's version of record is available at https://doi.org/10.1016/j.celrep.2018.02.040
Loss Of Setdb1 Decompacts The Inactive X Chromosome In Part Through Reactivation Of An Enhancer In The Il1rapl1 Gene
Loss Of Setdb1 Decompacts The Inactive X Chromosome In Part Through Reactivation Of An Enhancer In The Il1rapl1 Gene
Background: The product of dosage compensation in female mammals is the inactive X chromosome (Xi). Xi facultative heterochromatin is organized into two different types, one of which is defined by histone H3 trimethylated at lysine 9 (H3K9me3). The rationale for this study was to assess SET domain bifurcated 1 (SETDB1) as a candidate for maintaining this repressive modification at the human Xi. Results: Here, we show that loss of SETDB1 does not result in large-scale H3K9me3 changes at the Xi, but unexpectedly we observed striking decompaction of the Xi territory. Close examination revealed a 0.5 Mb region of the Xi that transitioned from H3K9me3 heterochromatin to euchromatin within the 3' end of the IL1RAPL1 gene that is part of a common chromosome fragile site that is frequently deleted or rearranged in patients afflicted with intellectual disability and other neurological ailments. Centrally located within this interval is a powerful enhancer adjacent to an ERVL-MaLR element. In the absence of SETDB1, the enhancer is reactivated on the Xi coupled with bidirectional transcription from the ERVL-MaLR element. Xa deletion of the enhancer/ERVL-MaLR resulted in loss of full-length IL1RAPL1 transcript in cis, coupled with trans decompaction of the Xi chromosome territory, whereas Xi deletion increased detection of full-length IL1RAPL1 transcript in trans, but did not impact Xi compaction. Conclusions: These data support a critical role for SETDB1 in maintaining the ERVL-MaLR element and adjacent enhancer in the 3' end of the IL1RAPL1 gene in a silent state to facilitate Xi compaction., Keywords: human genome, transcription factor, cell-lines, X chromosome inactivation, ctcf binding, Enhancer, Euchromatin, facultative heterochromatin, h3 lysine-27 methylation, Heterochromatin, histone methyltransferase activity, il1rapl1, Inactive X chromosome, linked intellectual disability, mental-retardation, setdb1, xist rna, Publication Note: The publisher’s version of record is available at https://doi.org/10.1186/s13072-018-0218-9
Loss of SETDB1 decompacts the inactive X chromosome in part through reactivation of an enhancer in the IL1RAPL1 gene.
Loss of SETDB1 decompacts the inactive X chromosome in part through reactivation of an enhancer in the IL1RAPL1 gene.
The product of dosage compensation in female mammals is the inactive X chromosome (Xi). Xi facultative heterochromatin is organized into two different types, one of which is defined by histone H3 trimethylated at lysine 9 (H3K9me3). The rationale for this study was to assess SET domain bifurcated 1 (SETDB1) as a candidate for maintaining this repressive modification at the human Xi. Here, we show that loss of SETDB1 does not result in large-scale H3K9me3 changes at the Xi, but unexpectedly we observed striking decompaction of the Xi territory. Close examination revealed a 0.5 Mb region of the Xi that transitioned from H3K9me3 heterochromatin to euchromatin within the 3' end of the IL1RAPL1 gene that is part of a common chromosome fragile site that is frequently deleted or rearranged in patients afflicted with intellectual disability and other neurological ailments. Centrally located within this interval is a powerful enhancer adjacent to an ERVL-MaLR element. In the absence of SETDB1, the enhancer is reactivated on the Xi coupled with bidirectional transcription from the ERVL-MaLR element. Xa deletion of the enhancer/ERVL-MaLR resulted in loss of full-length IL1RAPL1 transcript in cis, coupled with trans decompaction of the Xi chromosome territory, whereas Xi deletion increased detection of full-length IL1RAPL1 transcript in trans, but did not impact Xi compaction. These data support a critical role for SETDB1 in maintaining the ERVL-MaLR element and adjacent enhancer in the 3' end of the IL1RAPL1 gene in a silent state to facilitate Xi compaction., Keywords: Enhancer, Euchromatin, Heterochromatin, IL1RAPL1, Inactive X chromosome, SETDB1, X chromosome inactivation, Grant Number: GM117003, R01 GM117003, Publication Note: This NIH-funded author manuscript originally appeared in PubMed Central at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6088404.
M(6)a Pathway Facilitates Sex Determination In Drosophila
M(6)a Pathway Facilitates Sex Determination In Drosophila
The conserved modification N-6-methyladenosine (m(6)A) modulates mRNA processing and activity. Here, we establish the Drosophila system to study the m(6)A pathway. We first apply miCLIP to map m(6)A across embryogenesis, characterize its m(6)A 'writer' complex, validate its YTH 'readers' CG6422 and YT521-B, and generate mutants in five m(6)A factors. While m(6)A factors with additional roles in splicing are lethal, m(6)A-specific mutants are viable but present certain developmental and behavioural defects. Notably, m(6)A facilitates the master female determinant Sxl, since multiple m(6)A components enhance female lethality in Sxl sensitized backgrounds. The m(6)A pathway regulates Sxl processing directly, since miCLIP data reveal Sxl as a major intronic m(6)A target, and female-specific Sxl splicing is compromised in multiple m(6)A pathway mutants. YT521-B is a dominant m(6)A effector for Sxl regulation, and YT521-B overexpression can induce female-specific Sxl splicing. Overall, our transcriptomic and genetic toolkit reveals in vivo biologic function for the Drosophila m(6)A pathway., Keywords: in-vivo, gene-expression, 3' utrs, clip data, fl(2)d gene, messenger-rna methylation, n-6-adenosine methylation, nuclear-rna, single-nucleotide-resolution, yth domain, Publication Note: The publisher's version of record is available at https://doi.org/10.1038/ncomms15737
Margatoxin-bound quantum dots as a novel inhibitor of the voltage-gated ion channel Kv1.3.
Margatoxin-bound quantum dots as a novel inhibitor of the voltage-gated ion channel Kv1.3.
Venom-derived ion channel inhibitors have strong channel selectivity, potency, and stability; however, tracking delivery to their target can be challenging. Herein, we utilized luminescent quantum dots (QDs) conjugated to margatoxin (MgTx) as a traceable vehicle to target a voltage-dependent potassium channel, Kv1.3, which has a select distribution and well-characterized role in immunity, glucose metabolism, and sensory ability. We screened both unconjugated (MgTx) and conjugated MgTx (QD-MgTx) for their ability to inhibit Shaker channels Kv1.1 to Kv1.7 using patch-clamp electrophysiology in HEK293 cells. Our data indicate that MgTx inhibits 79% of the outward current in Kv1.3-transfected cells and that the QD-MgTx conjugate is able to achieve a similar level of block, albeit a slightly reduced efficacy (66%) and at a slower time course (50% block by 10.9 ± 1.1 min, MgTx; vs. 15.3 ± 1.2 min, QD-MgTx). Like the unbound peptide, the QD-MgTx conjugate inhibits both Kv1.3 and Kv1.2 at a 1 nM concentration, whereas it does not inhibit other screened Shaker channels. We tested the ability of QD-MgTx to inhibit native Kv1.3 expressed in the mouse olfactory bulb (OB). In brain slices of the OB, the conjugate acted similarly to MgTx to inhibit Kv1.3, causing an increased action potential firing frequency attributed to decreased intraburst duration rather than interspike interval. Our data demonstrate a retention of known biophysical properties associated with block of the vestibule of Kv1.3 by QD-MgTx conjugate compared to that of MgTx, inferring QDs could provide a useful tool to deliver ion channel inhibitors to targeted tissues in vivo., Keywords: K channel, MgTx, Nanoparticle, Patch clamp, Pore blocker, Potassium channel, Grant Number: F31 DC014638, R01 DC013080, T32 DC000044, Publication Note: This NIH-funded author manuscript originally appeared in PubMed Central at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5250575.
Matching The Forecast Horizon With The Relevant Spatial And Temporal Processes And Data Sources
Matching The Forecast Horizon With The Relevant Spatial And Temporal Processes And Data Sources
Most phenomenological, statistical models used to generate ecological forecasts take either a time-series approach, based on long-term data from one location, or a space-for-time approach, based on data describing spatial patterns across environmental gradients. However, the magnitude and even the sign of environment-response relationships detected using these two approaches often differs, leading to contrasting predictions about responses to future environmental change. Here we consider how the forecast horizon determines whether more accurate predictions come from the time-series approach, the space-for-time approach or a combination of the two. As proof of concept, we use simulated case studies to show that forecasts for short and long forecast horizons need to focus on different ecological processes, which are reflected in different kinds of data. First, we simulated population or community dynamics under stationary temperature using two simple, mechanistic models. Second, we fit statistical models to the simulated data using a time-series approach, a space-for-time approach or a weighted average. We then forecast the response to a temperature increase using the statistical models, and compared these forecasts to temperature effects simulated by the mechanistic models. We found that the time-series approach made accurate short-term predictions because it captured initial conditions and effects of fast processes such as birth and death. The space-for-time approach made more accurate long-term predictions because it better captured the influence of slower processes such as evolutionary and ecological selection. The weighted average made accurate predictions at all time scales, including intermediate time-scales where the other two approaches performed poorly. A weighted average of time-series and space-for-time approaches shows promise, but making this weighted model operational will require new research to predict the rate at which slow processes begin to influence dynamics., scale, abundance, climate-change, dispersal, eco-evolutionary dynamics, ecological forecasting, global change, no-analog communities, prediction, selection, space, species distribution models, The publisher's version of record is availible at https://doi.org/10.1111/ecog.05271
Mayans
Mayans
In spite of the wealth of available cultural and archeological information as well as general interest in the Mayans, little is known about their genetics. In this study, for the first time, we attempt to alleviate this lacuna of knowledge by comprehensively investigating the Y chromosome composition of contemporary Mayan populations throughout their domain. To accomplish this, five geographically targeted and ethnically distinct Mayan populations are investigated using Y-SNP and Y-STR markers. overall, the Mayan populations as a group are highly homogeneous, basically made up of only two autochthonous haplogroups, Q1a2a1a1*-M3 and Q1a2a1*-L54. Although the Y-STR data illustrates diversity, this diversity, for the most part, is uniformly distributed among geographically distant Mayan populations. Similar haplotypes among populations, abundance of singletons and absence of population partitioning within networks among Mayan populations suggest recent population expansion and substantial gene flow within the Mayan dominion, possibly due to the development of agriculture, the establishment of interacting City-State systems and commerce., Publication Note: This NIH-funded author manuscript originally appeared in PubMed Central at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4989206.

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