| 1. |
- Boija, Ann, et al.
(författare)
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CBP Regulates Recruitment and Release of Promoter-Proximal RNA Polymerase II
- 2017
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Ingår i: Molecular Cell. - : Elsevier BV. - 1097-2765 .- 1097-4164. ; 68:3, s. 491-503.e5
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Tidskriftsartikel (refereegranskat)abstract
- Transcription activation involves RNA polymerase II (Pol II) recruitment and release from the promoter into productive elongation, but how specific chromatin regulators control these steps is unclear. Here, we identify a novel activity of the histone acetyltransferase p300/CREB-binding protein (CBP) in regulating promoter-proximal paused Pol II. We find that Drosophila CBP inhibition results in "dribbling" of Pol II from the pause site to positions further downstream but impedes transcription through the +1 nucleosome genome-wide. Promoters strongly occupied by CBP and GAGA factor have high levels of paused Pol II, a unique chromatin signature, and are highly expressed regardless of cell type. Interestingly, CBP activity is rate limiting for Pol II recruitment to these highly paused promoters through an interaction with TFIIB but for transit into elongation by histone acetylation at other genes. Thus, CBP directly stimulates both Pol II recruitment and the ability to traverse the first nucleosome, thereby promoting transcription of most genes.
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| 2. |
- Greibe, Tine, 1974, et al.
(författare)
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Are "Pinholes" the Cause of Excess Current in Superconducting Tunnel Junctions? A Study of Andreev Current in Highly Resistive Junctions
- 2011
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Ingår i: Physical Review Letters. - 1079-7114 .- 0031-9007. ; 106:9
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Tidskriftsartikel (refereegranskat)abstract
- In highly resistive superconducting tunnel junctions, excess subgap current is usually observed and is often attributed to microscopic pinholes in the tunnel barrier. We have studied the subgap current in superconductor-insulator-superconductor (SIS) and superconductor-insulator-normal-metal ( SIN) junctions. In Al/AlOx/Al junctions, we observed a decrease of 2 orders of magnitude in the current upon the transition from the SIS to the SIN regime, where it then matched theory. In Al/AlOx/Cu junctions, we also observed generic features of coherent diffusive Andreev transport in a junction with a homogenous barrier. We use the quasiclassical Keldysh-Green function theory to quantify single- and two-particle tunneling and find good agreement with experiment over 2 orders of magnitude in transparency. We argue that our observations rule out pinholes as the origin of the excess current.
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| 3. |
- Holmqvist, Per-Henrik, et al.
(författare)
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Preferential Genome Targeting of the CBP Co-Activator by Rel and Smad Proteins in Early Drosophila melanogaster Embryos
- 2012
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Ingår i: PLOS Genetics. - San Francisco : Public Library of Science (PLoS). - 1553-7390 .- 1553-7404. ; 8:6
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Tidskriftsartikel (refereegranskat)abstract
- CBP and the related p300 protein are widely used transcriptional co-activators in metazoans that interact with multiple transcription factors. Whether CBP/p300 occupies the genome equally with all factors or preferentially binds together with some factors is not known. We therefore compared Drosophila melanogaster CBP (nejire) ChIP-seq peaks with regions bound by 40 different transcription factors in early embryos, and we found high co-occupancy with the Rel-family protein Dorsal. Dorsal is required for CBP occupancy in the embryo, but only at regions where few other factors are present. CBP peaks in mutant embryos lacking nuclear Dorsal are best correlated with TGF-beta/Dpp-signaling and Smad-protein binding. Differences in CBP occupancy in mutant embryos reflect gene expression changes genome-wide, but CBP also occupies some non-expressed genes. The presence of CBP at silent genes does not result in histone acetylation. We find that Polycomb-repressed H3K27me3 chromatin does not preclude CBP binding, but restricts histone acetylation at CBP-bound genomic sites. We conclude that CBP occupancy in Drosophila embryos preferentially overlaps factors controlling dorsoventral patterning and that CBP binds silent genes without causing histone hyperacetylation.
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| 4. |
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| 5. |
- Nylinder, Josefine, 1974, et al.
(författare)
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Modelling uncertainty for nitrate leaching and nitrous oxide emissions based on a Swedish field experiment with organic crop rotation
- 2011
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Ingår i: Agriculture, Ecosystems & Environment. - : Elsevier BV. - 0167-8809 .- 1873-2305. ; 141:1-2, s. 167-183
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Tidskriftsartikel (refereegranskat)abstract
- High uncertainties are common in detailed quantification of the N budget of agricultural cropping systems. The process-based CoupModel, integrated with the parameter calibration method known as Generalized likelihood uncertainty estimation (GLUE), was used here to define parameter values and estimate an N budget based on experimental data from an organic farming experiment in south-west Sweden. Data on nitrate (NO3-) leaching and nitrous oxide (N2O) emissions were used as a basis for quantifying N budget pools. A complete N budget with uncertainties associated with the different components of the N cycle compartments for two different fields (B2 and B4) is presented. Simulated N2O emissions contributed 1-2% of total N output, which corresponded to 7% and 8.7% of total N leaching for B2 and B4, respectively. Measured N2O emissions contributed 3.5% and 10.3% of total N leaching from B2 and B4, respectively. Simulated N inputs (deposition, plant N fixation and fertilisation) and outputs (emissions, leaching and harvest) showed a relatively small range of uncertainty, while the differences in N storage in the soil exhibited a larger range of uncertainty. One-fifth of the GLUE-calibrated parameters had a significant impact on simulated NO3- leaching and/or N2O emissions data. Emissions of N2O were strongly associated with the nitrification process. The high degree of equifinality indicated that a simpler model could be calibrated to the same field data.
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| 6. |
- Bernenko, Dolores, et al.
(författare)
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Mapping the semi-nested community structure of 3D chromosome contact networks
- 2022
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Mammalian DNA folds into 3D structures that facilitate and regulate genetic processes such as transcription, DNA repair, and epigenetics. Several insights derive from chromosome capture methods, such as Hi-C, which allow researchers to construct contact maps depicting 3D interactions among all DNA segment pairs. These maps show a complex cross-scale organization spanning megabase-pair compartments to short-ranged DNA loops. To better understand the organizing principles, several groups analyzed Hi-C data assuming a Russian-doll-like nested hierarchy where DNA regions of similar sizes merge into larger and larger structures. Apart from being a simple and appealing description, this model explains, e.g., the omnipresent chequerboard pattern seen in Hi-C maps, known as A/B compartments, and foreshadows the co-localization of some functionally similar DNA regions. However, while successful, this model is incompatible with the two competing mechanisms that seem to shape a significant part of the chromosomes’ 3D organization: loop extrusion and phase separation. This paper aims to map out the chromosome’s actual folding hierarchy from empirical data. To this end, we take advantage of Hi-C experiments and treat the measured DNA-DNA interactions as a weighted network. From such a network, we extract 3D communities using the generalized Louvain algorithm. This algorithm has a resolution parameter that allows us to scan seamlessly through the community size spectrum, from A/B compartments to topologically associated domains (TADs). By constructing a hierarchical tree connecting these communities, we find that chromosomes are more complex than a perfect hierarchy. Analyzing how communities nest relative to a simple folding model, we found that chromosomes exhibit a significant portion of nested and non-nested community pairs alongside considerable randomness. In addition, by examining nesting and chromatin types, we discovered that nested parts are often associated with active chromatin. These results highlight that crossscale relationships will be essential components in models aiming to reach a deep understanding of the causal mechanisms of chromosome folding.
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| 7. |
- Bernenko, Dolores, et al.
(författare)
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Mapping the semi-nested community structure of 3D chromosome contact networks
- 2023
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Ingår i: PloS Computational Biology. - : Public Library of Science (PLoS). - 1553-734X .- 1553-7358. ; 19:7
-
Tidskriftsartikel (refereegranskat)abstract
- Mammalian DNA folds into 3D structures that facilitate and regulate genetic processes such as transcription, DNA repair, and epigenetics. Several insights derive from chromosome capture methods, such as Hi-C, which allow researchers to construct contact maps depicting 3D interactions among all DNA segment pairs. These maps show a complex cross-scale organization spanning megabase-pair compartments to short-ranged DNA loops. To better understand the organizing principles, several groups analyzed Hi-C data assuming a Russian-doll-like nested hierarchy where DNA regions of similar sizes merge into larger and larger structures. Apart from being a simple and appealing description, this model explains, e.g., the omnipresent chequerboard pattern seen in Hi-C maps, known as A/B compartments, and foreshadows the co-localization of some functionally similar DNA regions. However, while successful, this model is incompatible with the two competing mechanisms that seem to shape a significant part of the chromosomes' 3D organization: loop extrusion and phase separation. This paper aims to map out the chromosome's actual folding hierarchy from empirical data. To this end, we take advantage of Hi-C experiments and treat the measured DNA-DNA interactions as a weighted network. From such a network, we extract 3D communities using the generalized Louvain algorithm. This algorithm has a resolution parameter that allows us to scan seamlessly through the community size spectrum, from A/B compartments to topologically associated domains (TADs). By constructing a hierarchical tree connecting these communities, we find that chromosomes are more complex than a perfect hierarchy. Analyzing how communities nest relative to a simple folding model, we found that chromosomes exhibit a significant portion of nested and non-nested community pairs alongside considerable randomness. In addition, by examining nesting and chromatin types, we discovered that nested parts are often associated with active chromatin. These results highlight that cross-scale relationships will be essential components in models aiming to reach a deep understanding of the causal mechanisms of chromosome folding.
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| 8. |
- Brindefalk, B., et al.
(författare)
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Bacterial composition in Swedish raw drinking water reveals three major interacting ubiquitous metacommunities
- 2022
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Ingår i: Microbiologyopen. - : Wiley. - 2045-8827. ; 11:5
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Tidskriftsartikel (refereegranskat)abstract
- Background Surface raw water used as a source for drinking water production is a critical resource, sensitive to contamination. We conducted a study on Swedish raw water sources, aiming to identify mutually co-occurring metacommunities of bacteria, and environmental factors driving such patterns. Methods The water sources were different regarding nutrient composition, water quality, and climate characteristics, and displayed various degrees of anthropogenic impact. Water inlet samples were collected at six drinking water treatment plants over 3 years, totaling 230 samples. The bacterial communities of DNA sequenced samples (n = 175), obtained by 16S metabarcoding, were analyzed using a joint model for taxa abundance. Results Two major groups of well-defined metacommunities of microorganisms were identified, in addition to a third, less distinct, and taxonomically more diverse group. These three metacommunities showed various associations to the measured environmental data. Predictions for the well-defined metacommunities revealed differing sets of favored metabolic pathways and life strategies. In one community, taxa with methanogenic metabolism were common, while a second community was dominated by taxa with carbohydrate and lipid-focused metabolism. Conclusion The identification of ubiquitous persistent co-occurring bacterial metacommunities in freshwater habitats could potentially facilitate microbial source tracking analysis of contamination issues in freshwater sources.
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| 9. |
- Cameron, Sarina R., et al.
(författare)
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PTE, a novel module to target Polycomb Repressive Complex 1 to the human cyclin D2 (CCND2) oncogene
- 2018
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Ingår i: Journal of Biological Chemistry. - : American Society for Biochemistry and Molecular Biology. - 0021-9258 .- 1083-351X. ; 293:37, s. 14342-14358
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Tidskriftsartikel (refereegranskat)abstract
- Polycomb group proteins are essential epigenetic repressors. They form multiple protein complexes of which two kinds, PRC1 and PRC2, are indispensable for repression. Although much is known about their biochemical properties, how mammalian PRC1 and PRC2 are targeted to specific genes is poorly understood. Here, we establish the cyclin D2 (CCND2) oncogene as a simple model to address this question. We provide the evidence that the targeting of PRC1 to CCND2 involves a dedicated PRC1-targeting element (PTE). The PTE appears to act in concert with an adjacent cytosine-phosphate-guanine (CpG) island to arrange for the robust binding of PRC1 and PRC2 to repressed CCND2. Our findings pave the way to identify sequence-specific DNA-binding proteins implicated in the targeting of mammalian PRC1 complexes and provide novel link between polycomb repression and cancer.
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| 10. |
- Dwibedi, Chinmay Kumar, et al.
(författare)
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Biological amplification of low frequency mutations unravels laboratory culture history of the bio-threat agent Francisella tularensis
- 2020
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Ingår i: Forensic Science International. - : Elsevier. - 1872-4973 .- 1878-0326. ; 45
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Tidskriftsartikel (refereegranskat)abstract
- Challenges of investigating a suspected bio attack include establishing if microorganisms have been cultured to produce attack material and to identify their source. Addressing both issues, we have investigated genetic variations that emerge during laboratory culturing of the bacterial pathogen Francisella tularensis. Key aims were to identify genetic variations that are characteristic of laboratory culturing and explore the possibility of using biological amplification to identify genetic variation present at exceedingly low frequencies in a source sample. We used parallel serial passage experiments and high-throughput sequencing of F. tularensis to explore the genetic variation. We found that during early laboratory culture passages of F. tularensis, gene duplications emerged in the pathogen genome followed by single-nucleotide polymorphisms in genes for bacterial capsule synthesis. Based on a biological enrichment scheme and the use of high-throughput sequencing, we identified genetic variation that likely pre-existed in a source sample. The results support that capsule synthesis gene mutations are common during laboratory culture, and that a biological amplification strategy is useful for linking a F. tularensis sample to a specific laboratory variant among many highly similar variants.
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| 11. |
- Figueiredo, Margarida L A, 1986-, et al.
(författare)
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HP1a Recruitment to Promoters Is Independent of H3K9 Methylation in Drosophila melanogaster
- 2012
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Ingår i: PLOS Genetics. - : Public Library of Science (PLoS). - 1553-7390 .- 1553-7404. ; 8:11, s. e1003061-
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Tidskriftsartikel (refereegranskat)abstract
- Heterochromatin protein 1 (HP1) proteins, recognized readers of the heterochromatin mark methylation of histone H3 lysine 9 (H3K9me), are important regulators of heterochromatin-mediated gene silencing and chromosome structure. In Drosophila melanogaster three histone lysine methyl transferases (HKMTs) are associated with the methylation of H3K9: Su(var)3-9, Setdb1, and G9a. To probe the dependence of HP1a binding on H3K9me, its dependence on these three HKMTs, and the division of labor between the HKMTs, we have examined correlations between HP1a binding and H3K9me patterns in wild type and null mutants of these HKMTs. We show here that Su(var)3-9 controls H3K9me-dependent binding of HP1a in pericentromeric regions, while Setdb1 controls it in cytological region 2L:31 and (together with POF) in chromosome 4. HP1a binds to the promoters and within bodies of active genes in these three regions. More importantly, however, HP1a binding at promoters of active genes is independent of H3K9me and POF. Rather, it is associated with heterochromatin protein 2 (HP2) and open chromatin. Our results support a hypothesis in which HP1a nucleates with high affinity independently of H3K9me in promoters of active genes and then spreads via H3K9 methylation and transient looping contacts with those H3K9me target sites.
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| 12. |
- Hägglund, Moa, et al.
(författare)
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Accounting for bacterial overlap between raw water communities and contaminating sources improves the accuracy of signature-based microbial source tracking
- 2018
-
Ingår i: Frontiers in Microbiology. - : Frontiers Media S.A.. - 1664-302X. ; 9
-
Tidskriftsartikel (refereegranskat)abstract
- Microbial source tracking (MST) analysis is essential to identifying and mitigating the fecal pollution of water resources. The signature-based MST method uses a library of sequences to identify contaminants based on operational taxonomic units (OTUs) that are unique to a certain source. However, no clear guidelines for how to incorporate OTU overlap or natural variation in the raw water bacterial community into MST analyses exist. We investigated how the inclusion of bacterial overlap between sources in the library affects source prediction accuracy. To achieve this, large-scale sampling-including feces from seven species, raw sewage, and raw water samples from water treatment plants - was followed by 16S rRNA amplicon sequencing. The MST library was defined using three settings: (i) no raw water communities represented; (ii) raw water communities selected through clustering analysis; and (iii) local water communities collected across consecutive years. The results suggest that incorporating either the local background or representative bacterial composition improves MST analyses, as the results were positively correlated to measured levels of fecal indicator bacteria and the accuracy at which OTUs were assigned to the correct contamination source increased fourfold. Using the proportion of OTUs with high source origin probability, underpinning a contaminating signal, is a solid foundation in a framework for further deciphering and comparing contaminating signals derived in signature-based MST approaches. In conclusion, incorporating background bacterial composition of water in MST can improve mitigation efforts for minimizing the spread of pathogenic and antibiotic resistant bacteria into essential freshwater resources.
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| 13. |
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| 14. |
- Karlsson, Edvin, et al.
(författare)
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Airborne microbial biodiversity and seasonality in Northern and Southern Sweden
- 2020
-
Ingår i: PeerJ. - : PeerJ. - 2167-8359. ; 8
-
Tidskriftsartikel (refereegranskat)abstract
- Microorganisms are essential constituents of ecosystems. To improve our understanding of how various factors shape microbial diversity and composition in nature it is important to study how microorganisms vary in space and time. Factors shaping microbial communities in ground level air have been surveyed in a limited number of studies, indicating that geographic location, season and local climate influence the microbial communities. However, few have surveyed more than one location, at high latitude or continuously over more than a year. We surveyed the airborne microbial communities over two full consecutive years in Kiruna, in the Arctic boreal zone, and Ljungbyhed, in the Southern nemoral zone of Sweden, by using a unique collection of archived air filters. We mapped both geographic and seasonal differences in bacterial and fungal communities and evaluated environmental factors that may contribute to these differences and found that location, season and weather influence the airborne communities. Location had stronger influence on the bacterial community composition compared to season, while location and season had equal influence on the fungal community composition. However, the airborne bacterial and fungal diversity showed overall the same trend over the seasons, regardless of location, with a peak during the warmer parts of the year, except for the fungal seasonal trend in Ljungbyhed, which fluctuated more within season. Interestingly, the diversity and evenness of the airborne communities were generally lower in Ljungbyhed. In addition, both bacterial and fungal communities varied significantly within and between locations, where orders like Rhizobiales, Rhodospirillales and Agaricales dominated in Kiruna, whereas Bacillales, Clostridiales and Sordariales dominated in Ljungbyhed. These differences are a likely reflection of the landscape surrounding the sampling sites where the landscape in Ljungbyhed is more homogenous and predominantly characterized by artificial and agricultural surroundings. Our results further indicate that local landscape, as well as seasonal variation, shapes microbial communities in air.
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| 15. |
- Karlsson, Edvin, 1985-
(författare)
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Using airborne eDNA to study ecosystem dynamics
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- In this era of global biodiversity crisis, the need to monitor ecological communities over space and time is more pressing than ever to effectively direct biodiversity conservation and management efforts. To understand the natural dynamics of an ecosystem, and the impact of anthropogenic activities related to environmental and climate change, long time series are needed to accurately link such processes to ecosystem change. This thesis uses a unique resource of archived air filters collected in Sweden originally intended for radioactive particle measurements to reconstruct historical eDNA abundance in the most extensive time-series of airborne eDNA abundance to date - spanning across four decades. By using metabarcoding and metagenomic analysis, it is evident that airborne eDNA from a very large diversity of species is present in air, representing all major branches of the tree of life, including bacteria, fungi, plants, metazoans, and viruses, from a wide range of terrestrial and aquatic sources. These have seasonal as well as long term trends that in part can be explained by temporal variation in climate and regional differences. The data generated in this thesis comprise an extensive resource for analysis of trends related to climate and environmental change and will also allow deeper studies on phenology, phenological change, functional genomics, and potentially antibiotic resistance. The results presented here show the potential of using airborne eDNA to monitor species in the local ecosystem over time and the methods provides an efficient tool for assessment of broad scale biodiversity, in a non-invasive and cost-effective way.
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| 16. |
- Kumar, Rajendra, et al.
(författare)
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Genome contact map explorer : a platform for the comparison, interactive visualization and analysis of genome contact maps
- 2017
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Ingår i: Nucleic Acids Research. - : Oxford University Press (OUP). - 0305-1048 .- 1362-4962. ; 45:17
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Tidskriftsartikel (refereegranskat)abstract
- Hi-C experiments generate data in form of large genome contact maps (Hi-C maps). These show that chromosomes are arranged in a hierarchy of three-dimensional compartments. But to understand how these compartments form and by how much they affect genetic processes such as gene regulation, biologists and bioinformaticians need efficient tools to visualize and analyze Hi-C data. However, this is technically challenging because these maps are big. In this paper, we remedied this problem, partly by implementing an efficient file format and developed the genome contact map explorer platform. Apart from tools to process Hi-C data, such as normalization methods and a programmable interface, we made a graphical interface that let users browse, scroll and zoom Hi-C maps to visually search for patterns in the Hi-C data. In the software, it is also possible to browse several maps simultaneously and plot related genomic data. The software is openly accessible to the scientific community.
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| 17. |
- Kumar, Rajendra, et al.
(författare)
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Genomic 3D compartments emerge from unfolding mitotic chromosomes
- 2019
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Ingår i: Chromosoma. - : Springer. - 0009-5915 .- 1432-0886. ; 128:1, s. 15-20
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Tidskriftsartikel (refereegranskat)abstract
- The 3D organisation of the genome in interphase cells is not a randomly folded polymer. Rather, experiments show that chromosomes arrange into a network of 3D compartments that correlate with biological processes, such as transcription, chromatin modifications and protein binding. However, these compartments do not exist during cell division when the DNA is condensed, and it is unclear how and when they emerge. In this paper, we focus on the early stages after cell division as the chromosomes start to decondense. We use a simple polymer model to understand the types of 3D structures that emerge from local unfolding of a compact initial state. From simulations, we recover 3D compartments, such as TADs and A/B compartments that are consistently detected in chromosome capture experiments across cell types and organisms. This suggests that the large-scale 3D organisation is a result of an inflation process.
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| 18. |
- Lee, Sang Hoon, et al.
(författare)
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Mapping the spectrum of 3D communities in human chromosome conformation capture data
- 2019
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Ingår i: Scientific Reports. - : Nature Publishing Group. - 2045-2322. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- Several experiments show that the three dimensional (3D) organization of chromosomes affects genetic processes such as transcription and gene regulation. To better understand this connection, researchers developed the Hi-C method that is able to detect the pairwise physical contacts of all chromosomal loci. The Hi-C data show that chromosomes are composed of 3D compartments that range over a variety of scales. However, it is challenging to systematically detect these cross-scale structures. Most studies have therefore designed methods for specific scales to study foremost topologically associated domains (TADs) and A/B compartments. To go beyond this limitation, we tailor a network community detection method that finds communities in compact fractal globule polymer systems. Our method allows us to continuously scan through all scales with a single resolution parameter. We found: (i) polymer segments belonging to the same 3D community do not have to be in consecutive order along the polymer chain. In other words, several TADs may belong to the same 3D community. (ii) CTCF proteins-a loop-stabilizing protein that is ascribed a big role in TAD formation-are well correlated with community borders only at one level of organization. (iii) TADs and A/B compartments are traditionally treated as two weakly related 3D structures and detected with different algorithms. With our method, we detect both by simply adjusting the resolution parameter. We therefore argue that they represent two specific levels of a continuous spectrum 3D communities, rather than seeing them as different structural entities.
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| 19. |
- Lewerentz, Jacob, 1992-
(författare)
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Genomic adaptation and gene-dosage regulation of Drosophila melanogaster cells, and long-read software developments
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Cells are the vehicles that allows genetic code to proliferate in the world, taking on various forms – as illustrated by the tree of life. The cell features are determined by the manufacturing of proteins, a process that has its blueprints encoded as genes in the genome. It is crucial for all cells to have the right amount of protein, regardless of context (part of a multicellular organism or self-sustained). The protein landscape (amount and type) vary depending on the environment. Cells of the multicellular organism should maintain the protein balance to provide its’ intended function in the organism tissue. The cells of multicellular organisms are faced with an imbalance due to sex-related chromosomal imbalances and other genome effects that change the number of gene copies. Restoration from the imbalance is done by dosage compensation systems. Cells that are isolated from the organism and grown inside the lab are common in research, known as cell lines. Cancer cells are similar to cell lines and have lost their original function in the organism in favor of a self-sustained lifestyle. The new environment (context) for these isolated cells impose a challenge; the cells must reorganize their genomes (holding the blueprints for proteins) to obtain autonomy.In this thesis, the genome evolution of isolated cells, cell lines, has been studied using Drosophila melanogaster (the fruit fly). Compared to normal cells of the host organism, cell line genomes are highly mutated and rearranged. With the emergence of novel sequencing technologies that can read long fragments of the genome, this complexity of cell line genomes can be captured. On the topic of novel sequencing technologies, new software implementations are presented and the future of software for long reads and complex genomes is discussed. The main focus of this thesis is to describe how an established and commonly used cell line has reorganized its’ genome to sustain a culture environment. Important information about the genome structure is provided to the research community. The thesis also describes the genome reorganization in new cell lines, covering the early adaptations to cell autonomy. Together, these investigations are of high relevance to human cancer research. This thesis has also studied the fundamentals for regulation of protein balance in organismal cells. Specifically, a recognition sequence to the X chromosome of the protein Painting of Fourth. This protein is related to dosage compensation and primarily enhance transcription from the 4 thchromosome in Drosophila melanogaster, but has been observed tooccasionally bind to the X chromosome.
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| 20. |
- Lewerentz, Jacob, 1992-, et al.
(författare)
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The path to immortalization of cells starts by managing stress through gene duplications
- 2023
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Ingår i: Experimental Cell Research. - : Elsevier. - 0014-4827 .- 1090-2422. ; 422:1
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Tidskriftsartikel (refereegranskat)abstract
- The genomes of immortalized cell lines (and cancer cells) are characterized by multiple types of aberrations, ranging from single nucleotide polymorphisms (SNPs) to structural rearrangements that have accumulated over time. Consequently, it is difficult to estimate the relative impact of different aberrations, the order of events, and which gene functions were under selective pressure at the early stage towards cellular immortalization. Here, we have established novel cell cultures derived from Drosophila melanogaster embryos that were sampled at multiple time points over a one-year period. Using short-read DNA sequencing, we show that copy-number gain in preferentially stress-related genes were acquired in a dominant fraction of cells in 300-days old cultures. Furthermore, transposable elements were active in cells of all cultures. Only a few (<1%) SNPs could be followed over time, and these showed no trend to increase or decrease. We conclude that the early cellular responses of a novel culture comprise sequence duplication and transposable element activity. During immortalization, positive selection first occurs on genes that are related to stress response before shifting to genes that are related to growth.
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| 21. |
- Lewerentz, Jacob, et al.
(författare)
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Transposon activity, local duplications and propagation of structural variants across haplotypes drive the evolution of the Drosophila S2 cell line
- 2022
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Ingår i: BMC Genomics. - : BioMed Central. - 1471-2164. ; 23:1
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Tidskriftsartikel (refereegranskat)abstract
- Background: Immortalized cell lines are widely used model systems whose genomes are often highly rearranged and polyploid. However, their genome structure is seldom deciphered and is thus not accounted for during analyses. We therefore used linked short- and long-read sequencing to perform haplotype-level reconstruction of the genome of a Drosophila melanogaster cell line (S2-DRSC) with a complex genome structure.Results: Using a custom implementation (that is designed to use ultra-long reads in complex genomes with nested rearrangements) to call structural variants (SVs), we found that the most common SV was repetitive sequence insertion or deletion (> 80% of SVs), with Gypsy retrotransposon insertions dominating. The second most common SV was local sequence duplication. SNPs and other SVs were rarer, but several large chromosomal translocations and mitochondrial genome insertions were observed. Haplotypes were highly similar at the nucleotide level but structurally very different. Insertion SVs existed at various haplotype frequencies and were unlinked on chromosomes, demonstrating that haplotypes have different structures and suggesting the existence of a mechanism that allows SVs to propagate across haplotypes. Finally, using public short-read data, we found that transposable element insertions and local duplications are common in other D. melanogaster cell lines.Conclusions: The S2-DRSC cell line evolved through retrotransposon activity and vast local sequence duplications, that we hypothesize were the products of DNA re-replication events. Additionally, mutations can propagate across haplotypes (possibly explained by mitotic recombination), which enables fine-tuning of mutational impact and prevents accumulation of deleterious events, an inherent problem of clonal reproduction. We conclude that traditional linear homozygous genome representation conceals the complexity when dealing with rearranged and heterozygous clonal cells.
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| 22. |
- Lindgren, Petter, et al.
(författare)
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A likelihood ratio-based approach for improved source attribution in microbiological forensic investigations
- 2019
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Ingår i: Forensic Science International. - : Elsevier. - 0379-0738 .- 1872-6283. ; 302
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Tidskriftsartikel (refereegranskat)abstract
- A common objective in microbial forensic investigations is to identify the origin of a recovered pathogenic bacterium by DNA sequencing. However, there is currently no consensus about how degrees of belief in such origin hypotheses should be quantified, interpreted, and communicated to wider audiences. To fill this gap, we have developed a concept based on calculating probabilistic evidential values for microbial forensic hypotheses. The likelihood-ratio method underpinning this concept is widely used in other forensic fields, such as human DNA matching, where results are readily interpretable and have been successfully communicated in juridical hearings. The concept was applied to two case scenarios of interest in microbial forensics: (1) identifying source cultures among series of very similar cultures generated by parallel serial passage of the Tier 1 pathogen Francisella tularensis, and (2) finding the production facilities of strains isolated in a real disease outbreak caused by the human pathogen Listeria monocytogenes. Evidence values for the studied hypotheses were computed based on signatures derived from whole genome sequencing data, including deep-sequenced low-frequency variants and structural variants such as duplications and deletions acquired during serial passages. In the F. tularensis case study, we were able to correctly assign fictive evidence samples to the correct culture batches of origin on the basis of structural variant data. By setting up relevant hypotheses and using data on cultivated batch sources to define the reference populations under each hypothesis, evidential values could be calculated. The results show that extremely similar strains can be separated on the basis of amplified mutational patterns identified by high-throughput sequencing. In the L. monocytogenes scenario, analyses of whole genome sequence data conclusively assigned the clinical samples to specific sources of origin, and conclusions were formulated to facilitate communication of the findings. Taken together, these findings demonstrate the potential of using bacterial whole genome sequencing data, including data on both low frequency SNP signatures and structural variants, to calculate evidence values that facilitate interpretation and communication of the results. The concept could be applied in diverse scenarios, including both epidemiological and forensic source tracking of bacterial infectious disease outbreaks.
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| 23. |
- Lundberg, Lina E, et al.
(författare)
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Buffering and proteolysis are induced by segmental monosomy in Drosophila melanogaster
- 2012
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Ingår i: Nucleic Acids Research. - Oxford : Oxford University Press. - 0305-1048 .- 1362-4962. ; 40:13, s. 5926-5937
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Tidskriftsartikel (refereegranskat)abstract
- Variation in the number of individual chromosomes (chromosomal aneuploidy) or chromosome segments (segmental aneuploidy) is associated with developmental abnormalities and reduced fitness in all species examined; it is the leading cause of miscarriages and mental retardation and a hallmark of cancer. However, despite their documented importance in disease, the effects of aneuploidies on the transcriptome remain largely unknown. We have examined the expression effects of seven heterozygous chromosomal deficiencies, both singly and in all pairwise combinations, in Drosophila melanogaster. The results show that genes in one copy are buffered, i.e. expressed more strongly than the expected 50% of wild-type level, the buffering is general and not influenced by other monosomic regions. Furthermore, long genes are significantly more highly buffered than short genes and gene length appears to be the primary determinant of the buffering degree. For short genes the degree of buffering depends on expression level and expression pattern. Furthermore, the results show that in deficiency heterozygotes the expression of genes involved in proteolysis is enhanced and negatively correlates with the degree of buffering. Thus, enhanced proteolysis appears to be a general response to aneuploidy.
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| 24. |
- Lundberg, Lina E, 1982-, et al.
(författare)
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HP1a, Su(var)3-9, SETDB1 and POF stimulate or repress gene expression depending on genomic position, gene length and expression pattern in Drosophila melanogaster
- 2013
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Ingår i: Nucleic Acids Research. - : Oxford University Press (OUP). - 0305-1048 .- 1362-4962. ; 41:8, s. 4481-4494
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Tidskriftsartikel (refereegranskat)abstract
- Heterochromatin protein 1a (HP1a) is a chromatin-associated protein important for the formation and maintenance of heterochromatin. In Drosophila, the two histone methyltransferases SETDB1 and Su(var)3-9 mediate H3K9 methylation marks that initiates the establishment and spreading of HP1a-enriched chromatin. Although HP1a is generally regarded as a factor that represses gene transcription, several reports have linked HP1a binding to active genes, and in some cases, it has been shown to stimulate transcriptional activity. To clarify the function of HP1a in transcription regulation and its association with Su(var)3-9, SETDB1 and the chromosome 4-specific protein POF, we conducted genome-wide expression studies and combined the results with available binding data in Drosophila melanogaster. The results suggest that HP1a, SETDB1 and Su(var)3-9 repress genes on chromosome 4, where non-ubiquitously expressed genes are preferentially targeted, and stimulate genes in pericentromeric regions. Further, we showed that on chromosome 4, Su(var)3-9, SETDB1 and HP1a target the same genes. In addition, we found that transposons are repressed by HP1a and Su(var)3-9 and that the binding level and expression effects of HP1a are affected by gene length. Our results indicate that genes have adapted to be properly expressed in their local chromatin environment.
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| 25. |
- Mahmud, A. K. M. Firoj, et al.
(författare)
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Exploring a Drosophila Transcription Factor Interaction Network to Identify Cis-Regulatory Modules
- 2020
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Ingår i: Journal of Computational Biology. - : Mary Ann Liebert. - 1066-5277 .- 1557-8666. ; 27:8, s. 1313-1328
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Tidskriftsartikel (refereegranskat)abstract
- Multiple transcription factors (TFs) bind to specific sites in the genome and interact among themselves to form the cis-regulatory modules (CRMs). They are essential in modulating the expression of genes, and it is important to study this interplay to understand gene regulation. In the present study, we integrated experimentally identified TF binding sites collected from published studies with computationally predicted TF binding sites to identifyDrosophilaCRMs. Along with the detection of the previously known CRMs, this approach identified novel protein combinations. We determined high-occupancy target sites, where a large number of TFs bind. Investigating these sites revealed that Giant, Dichaete, and Knirp are highly enriched in these locations. A common TAG team motif was observed at these sites, which might play a role in recruiting other TFs. While comparing the binding sites at distal and proximal promoters, we found that certain regulatory TFs, such as Zelda, were highly enriched in enhancers. Our study has shown that, from the information available concerning the TF binding sites, the real CRMs could be predicted accurately and efficiently. Although we only may claim co-occurrence of these proteins in this study, it may actually point to their interaction (as known interaction proteins typically co-occur together). Such an integrative approach can, therefore, help us to provide a better understanding of the interplay among the factors, even though further experimental verification is required.
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| 26. |
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| 27. |
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| 28. |
- Nyberg, Markus, et al.
(författare)
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Modeling protein target search in human chromosomes
- 2021
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Ingår i: Physical Review Research. - : American Physical Society. - 2643-1564. ; 3:1
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Tidskriftsartikel (refereegranskat)abstract
- Several processes in the cell, such as gene regulation, start when key proteins recognize and bind to short DNA sequences. However, as these sequences can be hundreds of million times shorter than the genome, they are hard to find by simple diffusion: diffusion-limited association rates may underestimate in vitro measurements up to several orders of magnitude. Moreover, the rates increase if the DNA is coiled rather than straight. Here we model how this works in vivo in mammalian cells. We use chromatin-chromatin contact data from Hi-C experiments to map the protein target-search onto a network problem. The nodes represent DNA segments and the weight of the links are proportional to measured contact probabilities. We then put forward a diffusion-reaction equation for the density of searching protein that allows us to calculate the association rates across the genome analytically. For segments where the rates are high, we find that they are enriched with active gene starts and have high RNA expression levels. This paper suggests that the DNA's 3D conformation is important for protein search times in vivo and offers a method to interpret protein-binding profiles in eukaryotes that cannot be explained by the DNA sequence itself.
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| 29. |
- Philip, Philge, et al.
(författare)
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CBP binding outside of promoters and enhancers in Drosophila melanogaster
- 2015
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Ingår i: Epigenetics & Chromatin. - : Springer Science and Business Media LLC. - 1756-8935. ; 8
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Tidskriftsartikel (refereegranskat)abstract
- Background: CREB-binding protein (CBP, also known as nejire) is a transcriptional co-activator that is conserved in metazoans. CBP plays an important role in embryonic development and cell differentiation and mutations in CBP can lead to various diseases in humans. In addition, CBP and the related p300 protein have successfully been used to predict enhancers in both humans and flies when they occur with monomethylation of histone H3 on lysine 4 (H3K4me1). Results: Here, we compare CBP chromatin immunoprecipitation sequencing data from Drosophila S2 cells with modENCODE data and show that CBP is bound at genomic sites with a wide range of functions. As expected, we find that CBP is bound at active promoters and enhancers. In addition, we find that the strongest CBP sites in the genome are found at Polycomb response elements embedded in histone H3 lysine 27 trimethylated (H3K27me3) chromatin, where they correlate with binding of the Pho repressive complex. Interestingly, we find that CBP also binds to most insulators in the genome. At a subset of these, CBP may regulate insulating activity, measured as the ability to prevent repressive H3K27 methylation from spreading into adjacent chromatin. Conclusions: We conclude that CBP could be involved in a much wider range of functions than has previously been appreciated, including Polycomb repression and insulator activity. In addition, we discuss the possibility that a common role for CBP at all functional elements may be to regulate interactions between distant chromosomal regions and speculate that CBP is controlling higher order chromatin organization.
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| 30. |
- Philip, Philge, et al.
(författare)
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Male X-linked genes in Drosophila melanogaster are compensated independently of the Male-Specific Lethal complex
- 2013
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Ingår i: Epigenetics & Chromatin. - : BioMed Central (BMC). - 1756-8935. ; 6:Article number: 35
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: In organisms where the two sexes have unequal numbers of X-chromosomes, the expression of X-linked genes needs to be balanced not only between the two sexes, but also between X and the autosomes. In Drosophila melanogaster, the Male-Specific Lethal (MSL) complex is believed to produce a 2-fold increase in expression of genes on the male X, thus restoring this balance.RESULTS: Here we show that almost all the genes on the male X are effectively compensated. However, many genes are compensated without any significant recruitment of the MSL-complex. These genes are very weakly, if at all, affected by mutations or RNAi against MSL-complex components. In addition, even the genes that are strongly bound by MSL rely on mechanisms other than the MSL-complex for proper compensation. We find that long, non-ubiquitously expressed genes tend to rely less on the MSL-complex for their compensation and genes that in addition are far from High Affinity Sites tend to not bind the complex at all or very weakly.CONCLUSIONS: We conclude that most of the compensation of X-linked genes is produced by an MSL-independent mechanism. Similar to the case of the MSL-mediated compensation we do not yet know the mechanism behind the MSL-independent compensation that appears to act preferentially on long genes. Even if we observe similarities, it remains to be seen if the mechanism is related to the buffering that is observed in autosomal aneuploidies.
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| 31. |
- Philip, Philge, et al.
(författare)
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Sequence signatures involved in targeting the male-specific lethal complex to X-chromosomal genes in Drosophila melanogaster
- 2012
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Ingår i: BMC Genomics. - : BioMed Central. - 1471-2164. ; 13, s. 97-
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: In Drosophila melanogaster, the dosage-compensation system that equalizes X-linked gene expression between males and females, thereby assuring that an appropriate balance is maintained between the expression of genes on the X chromosome(s) and the autosomes, is at least partially mediated by the Male-Specific Lethal (MSL) complex. This complex binds to genes with a preference for exons on the male X chromosome with a 3' bias, and it targets most expressed genes on the X chromosome. However, a number of genes are expressed but not targeted by the complex. High affinity sites seem to be responsible for initial recruitment of the complex to the X chromosome, but the targeting to and within individual genes is poorly understood.RESULTS: We have extensively examined X chromosome sequence variation within five types of gene features (promoters, 5' UTRs, coding sequences, introns, 3' UTRs) and intergenic sequences, and assessed its potential involvement in dosage compensation. Presented results show that: the X chromosome has a distinct sequence composition within its gene features; some of the detected variation correlates with genes targeted by the MSL-complex; the insulator protein BEAF-32 preferentially binds upstream of MSL-bound genes; BEAF-32 and MOF co-localizes in promoters; and that bound genes have a distinct sequence composition that shows a 3' bias within coding sequence.CONCLUSIONS: Although, many strongly bound genes are close to a high affinity site neither our promoter motif nor our coding sequence signatures show any correlation to HAS. Based on the results presented here, we believe that there are sequences in the promoters and coding sequences of targeted genes that have the potential to direct the secondary spreading of the MSL-complex to nearby genes.
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| 32. |
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| 33. |
- Sobhy, Haitham, et al.
(författare)
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Highly interacting regions of the human genome are enriched with enhancers and bound by DNA repair proteins
- 2019
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Ingår i: Scientific Reports. - : Nature Publishing Group. - 2045-2322. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- In specific cases, chromatin clearly forms long-range loops that place distant regulatory elements in close proximity to transcription start sites, but we have limited understanding of many loops identified by Chromosome Conformation Capture (such as Hi-C) analyses. In efforts to elucidate their characteristics and functions, we have identified highly interacting regions (HIRs) using intra-chromosomal Hi-C datasets with a new computational method based on looking at the eigenvector that corresponds to the smallest eigenvalue (here unity). Analysis of these regions using ENCODE data shows that they are in general enriched in bound factors involved in DNA damage repair and have actively transcribed genes. However, both highly transcribed regions as well as transcriptionally inactive regions can form HIRs. The results also indicate that enhancers and super-enhancers in particular form long-range interactions within the same chromosome. The accumulation of DNA repair factors in most identified HIRs suggests that protection from DNA damage in these regions is essential for avoidance of detrimental rearrangements.
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| 34. |
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| 35. |
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| 36. |
- Stenberg, Per, 1974-, et al.
(författare)
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Buffering of segmental and chromosomal aneuploidies in Drosophila melanogaster
- 2009
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Ingår i: PLOS Genetics. - : Public Library of Science (PLoS). - 1553-7390 .- 1553-7404. ; 5:5
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Tidskriftsartikel (refereegranskat)abstract
- Chromosomal instability, which involves the deletion and duplication of chromosomes or chromosome parts, is a common feature of cancers, and deficiency screens are commonly used to detect genes involved in various biological pathways. However, despite their importance, the effects of deficiencies, duplications, and chromosome losses on the regulation of whole chromosomes and large chromosome domains are largely unknown. Therefore, to explore these effects, we examined expression patterns of genes in several Drosophila deficiency hemizygotes and a duplication hemizygote using microarrays. The results indicate that genes expressed in deficiency hemizygotes are significantly buffered, and that the buffering effect is general rather than being mainly mediated by feedback regulation of individual genes. In addition, differentially expressed genes in haploid condition appear to be generally more strongly buffered than ubiquitously expressed genes in haploid condition, but, among genes present in triploid condition, ubiquitously expressed genes are generally more strongly buffered than differentially expressed genes. Furthermore, we show that the 4th chromosome is compensated in response to dose differences. Our results suggest general mechanisms have evolved that stimulate or repress gene expression of aneuploid regions as appropriate, and on the 4th chromosome of Drosophila this compensation is mediated by Painting of Fourth (POF).
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| 37. |
- Sundell, David, et al.
(författare)
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FlexTaxD : flexible modification of taxonomy databases for improved sequence classification
- 2021
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Ingår i: Bioinformatics. - : Oxford University Press. - 1367-4803 .- 1367-4811 .- 1460-2059. ; 37:21, s. 3932-3933
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Tidskriftsartikel (refereegranskat)abstract
- The Flexible Taxonomy Database framework provides a method for modification and merging official and custom taxonomic databases to create improved databases. Using such databases will increase accuracy and precision of existing methods to classify sequence reads.
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| 38. |
- Zare, Aman, 1987-
(författare)
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Regulation of gene expression in fruit flies : how does it start, and will it be remembered?
- 2018
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- One of the most distinctive features of eukaryotic chromosomes is the bundling of DNA together with functionally associated RNA and proteins in chromatin. This allows huge amounts of DNA to be packed inside the very tiny space of the nucleus, and alterations in the structure of chromatin enable access to the DNA for transcription (“reading” genes by production of RNA copies). Much of the current knowledge of chromatin structure and regulation comes from studies of Drosophila melanogaster. When the chromatin structure is open the transcription of a gene can start after recruitment of the necessary factors. The main enzyme for gene transcription is Polymerase II (Pol II). For successful gene transcription, Pol II must not only be recruited to the gene’s promoter, but also escape from a pausing state which occurs soon after transcription initiation. CBP/P300 is one of the co-activators involved in transcriptional activation. In the studies this thesis is based upon, my colleagues and I (hereafter we) discovered a new function for CBP in transcription activation. Using high throughput sequencing techniques, we found that CBP directly stimulates recruitment of Pol II to promoters, and facilitates its release from the paused state, enabling progression to the elongation stage of transcription.For cells to remember their identity following division during development, the transcriptional state of genes must be transmitted. Intensively studied players involved in this memory are the Polycomb group (PcG) proteins, responsible for maintaining the repressed state of important developmental genes. The core members are Polycomb repressive complex 1 and 2 (PRC1 and PRC2), which are recruited in flies through poorly known mechanisms to target genes by so-called Polycomb response elements (PREs). Using Drosophila mutant cell lines, we showed that (in contrast to previous models) some PREs can recruit PRC1 even when PRC2 is absent. We also observed that at many PREs, PRC1 is needed for recruitment of PRC2 and concluded that targeting PRC complexes to PREs is a much more flexible and variable process than previously thought.Some phenotypic effects of environmental changes can be transferred to subsequent generations. Previous efforts to identify the mechanisms involved have focused on material (mainly, but not only, DNA) transferred through germ cells. However, organisms’ microbiomes are also transferred to the next generation. Thus, to investigate possible contributions of microbiomes to such transfer, we used fruit flies as the microbiomes they inherit can be easily controlled. We altered some parents’ environmental conditions by lowering the temperature, then grew offspring that received microbiomes from cold-treated and control parents in control conditions and compared their transcriptional patterns. Our results suggest that most of the crosstalk between the microbiome and the fly happens in the gut, and that further investigation of this previously unsuspected mode of inheritance is warranted.
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| 39. |
- Zare, Aman, et al.
(författare)
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The gut microbiome participates in transgenerational inheritance of low temperature responses in Drosophila melanogaster
- 2018
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Ingår i: FEBS Letters. - : John Wiley & Sons. - 0014-5793 .- 1873-3468. ; 592:24, s. 4078-4086
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Tidskriftsartikel (refereegranskat)abstract
- Environmental perturbations induce transcriptional changes, some of which may be inherited even in the absence of the initial stimulus. Previous studies have focused on transfers through the germ-line although microbiota is also passed on to the offspring. Thus, we inspected the involvement of the gut microbiome in transgenerational inheritance of environmental exposures in Drosophila melanogaster. We grew flies in the cold versus control temperatures and compared their transcriptional patterns in both conditions as well as in their offspring. F2 flies grew in control temperature while we controlled their microbiota acquisition from either F1 sets. Transcriptional status of some genes was conserved transgenerationally, and a subset of these genes, mainly expressed in the gut, was transcriptionally dependent on the acquired microbiome. This article is protected by copyright. All rights reserved.
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