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- Ueberschär, Malin, 1991-
(författare)
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BEN-solo proteins in genome architecture and function
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Eukaryotic genomes are organized in a complex manner inside the nucleus of each cell. Chromatin insulators are DNA elements that help to fold the linear DNA fiber in three-dimensional space and partition regulatory gene units. However, the mechanisms by which chromatin insulators and their associated proteins, insulator binding proteins (IBPs), determine genome organization and modulate gene expression remain elusive. This thesis investigates these mechanisms by examining the roles of a stage-specific family of insulator proteins in Drosophila, the BEN-solo proteins. The BEN proteins regulate gene expression and cell specification in animal development. We characterized the genomic binding sites of the three BEN-solo proteins Elba1, Elba2 and Insensitive (Insv), and the Elba3 protein that acts as adapter for the heterotrimeric ELBA complex. ELBA and Insv share a set of common targets at the time of their ubiquitous expression – throughout the major wave of zygotic genome activation – but also regulate distinct loci. We found that ELBA and Insv not only repress their direct target genes, but also co-localize and act in redundancy with other known insulator binding proteins. Importantly, ELBA and Insv insulate closely-spaced transcription units ensuring the correct levels of promoter transcripts.The regulatory state of paused RNA Polymerase II (Pol II) has been suggested to comprise a mechanism for insulator function. To understand how paused promoters may block enhancer interaction, we assessed a series of paused and non-paused promoters in a transgenic reporter assay. Although we observed a high likelihood of paused promoters to act as enhancer blockers, there were several exceptions. Using a paused representative promoter, we dissected the contributions of Pol II and promoter-proximal IBPs to enhancer-blocking. We found that Pol II is dispensable for insulation at this site, while the IBPs and their motifs are essential, arguing against a general function of paused Pol II as enhancer-blocking mechanism.Insulators are known to set barriers between active and silent chromatin domains. We found that chromatin accessibility at chromatin boundaries in elba and insv mutants is frequently altered, suggesting an involvement of the proteins in chromatin boundary formation. Remarkably, shifted boundaries are accompanied by a deregulation of nascent transcripts. We further assessed the initial establishment of heterochromatin domains. Using a reporter embedded in the heterochromatin of the Drosophila Y chromosome, and the GFP-tagged heterochromatin protein 1a (HP1a), we found that ELBA contributes to heterochromatin silencing while Insv appears to limit HP1a clustering on a global scale. We hypothesize that ELBA and Insv contribute to boundary formation at some common sites, while diverging in their contributions to heterochromatin domain formation.Taken together, our results identify multiple roles of the BEN-solo family as chromatin factors, and highlight the intrinsic properties of insulator elements and IBPs as the main mechanism in enhancer blocking.
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| 2. |
- Örkenby, Lovisa, 1992-
(författare)
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Small non-coding RNA in early fly development : plasticity, interactions and improved bioinformatic tools
- 2023
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- At fertilization, the male and female pronuclei undergo a transformation from germline to pluripotency as they fuse, marking the beginning of Drosophila embryogenesis. As the parental contributions decrease, the zygote takes control of its genome in a process called the maternal-to-zygotic transition (MZT). Several small non-coding RNAs (sncRNAs), a very large and diverse group of RNAs, have regulatory roles during this transition. This includes for example microRNAs (miRNAs) and piwi-interacting RNAs (piRNAs). Regulation by miRNAs mainly occurs through mediating maternal mRNA degradation, while piRNAs operate by repressing transposable elements (TEs) and regulating the nanos-induced embryonic body axis determination.In this thesis, the complex and dynamic field of early Drosophila embryogenesis and sncRNAs are put in relation to the included papers. In Paper I, I explored the most stress-sensitive embryonic period and found that stress before the midblastula transition retains maternal miRNAs. These miRNAs impact zygotic gene activation by modulating the boundary factor Elba1, leading to compromised transcription control. Paper III examines the piRNA population during MZT. I find differences of unique piRNA sequences in embryos of different ages but not in target preferences, potentially highlighting the importance of constant repression of certain TEs. Paper II addresses specific difficulties with sncRNA seq data analysis and presents a bioinformatic framework to improve these analyses using a sequence-based strategy.This thesis highlights the intricate interplay of sncRNAs in the critical period of early Drosophila embryogenesis and offers insights into their regulatory roles.
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| 3. |
- Öst, Anita, et al.
(författare)
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Paternal Diet Defines Offspring Chromatin State and Intergenerational Obesity
- 2014
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Ingår i: Cell. - : Elsevier (Cell Press). - 0092-8674 .- 1097-4172. ; 159:6, s. 1352-1364
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Tidskriftsartikel (refereegranskat)abstract
- The global rise in obesity has revitalized a search for genetic and epigenetic factors underlying the disease. We present a Drosophila model of paternal-diet-induced intergenerational metabolic reprogramming (IGMR) and identify genes required for its encoding in offspring. Intriguingly, we find that as little as 2 days of dietary intervention in fathers elicits obesity in offspring. Paternal sugar acts as a physiological suppressor of variegation, desilencing chromatin-state-defined domains in both mature sperm and in offspring embryos. We identify requirements for H3K9/K27me3-dependent reprogramming of metabolic genes in two distinct germline and zygotic windows. Critically, we find evidence that a similar system may regulate obesity susceptibility and phenotype variation in mice and humans. The findings provide insight into the mechanisms underlying intergenerational metabolic reprogramming and carry profound implications for our understanding of phenotypic variation and evolution.
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