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Characterization of regulators of CpG island activity in mouse stem cells
Get PDFCpG island (CGI) promoters control the expression of over two thirds of all mammalian genes, many which are highly active. Nevertheless, we are still lacking an understanding how their hallmark feature, the high frequency of unmethylated CpGs, contributes to promoter activity. A recent study showed that CpG density is necessary for transcriptional activity in mouse embryonic stem cells (mESCs), independent of transcription factors (TFs) that bind complex DNA motifs (Hartl et al., 2019). This thesis explores the contribution of proteins that bind unmethylated CpGs at CGIs and characterizes a distinct case in which binding of the TF BANP (BTG3 Associated Nuclear Protein) depends on promoter CpG density.
In the first part of this thesis, we aim to identify binders of unmethylated CpGs in mESCs. To that end, we employed oligo-affinity purification coupled with quantitative mass spectrometry (oligo-AP MS) utilizing three different promoters each with three different CpG densities. We identify common and specific hits (often representing specific protein complexes) that enrich with higher promoter CpG density. For selected targets we validate increased binding with higher CpG density in vivo (based on published ChIP-seq data), including the ZF-CxxC containing proteins KMT2B (Lysine Methyltransferase 2B) and KDM2B (Lysine Demethylase 2B) as well as ZBTB2 (Zinc Finger And BTB Domain Containing 2) and SAMD1 (Sterile Alpha Motif Domain Containing 1). Motif analysis, at least in the case of ZBTB2 and SAMD1, suggests that additional motif preferences beyond “CG” exist, offering an explanation for differential binding between the factors.
In order to explore how selected CpG binders contribute to promoter activity, we tested how transcription was affected upon their deletion in mESCs, particularly in the context of CpG density and binding. Upon genetic deletion we detect small trends at bound, CpG-rich sites towards a decrease of promoter activity for Kmt2b-/- and to a minor degree Zbtb2-/- cells as well as an increase for Kdm2b-/- and Samd1-/- cells. For Zbtb2-/- and Samd1-/- cells, we tested how stably expressed luciferase reporters with different promoter CpG densities behave in the mutant backgrounds compared to wildtype (WT), and the results align with the genome-wide trends. As the effect sizes at bound and CpG-rich promoters are relatively small, we generated combinatorial protein deletions to test for additional effects. Overall, we observe that most of the transcriptional changes upon combinatorial deletions can potentially be explained as additions of the single deletions. Furthermore, transcriptional changes at commonly bound sites appear to be a mixture of what is expected based on the single deletions. Finally, we explored functional links based on bound proteins at CGIs close to genes affected by our targets of interest; we identify common patterns as well as target-specific hits.
The last part of this thesis explores the observation that binding of the TF BANP is dependent on CpG density. BANP is a strong activator in mESCs, allowing the generation of a promoter-luciferase reporter that is highly dependent on BANP (Grand et al., 2021). We generated BANP promoter constructs with five different CpG densities and determined their transcriptional activity, as well as the footprints they generate when stably expressed in cells with and without DNA methylation. We find that the BANP reporters strongly depend on CpG density with a strong decrease in activity at 50 % non-motif CpG density that is partially rescued in cells lacking DNA methylation. Single molecule footprinting (SMF) suggests that CpG density as well as DNA methylation contribute to DNA accessibility and BANP binding. Finally, we observe a disconnect between loss of reporter activity and loss of BANP binding, which requires further validation but speculatively suggests the contribution of additional factors to BANP promoter activity.
Taken together, this work identifies proteins that preferentially bind CGI promoters and show trends to either activate or inhibit transcription at their bound sites. Our study on CpG-binding proteins gives some insight into the regulation of their bound CGI promoters, offering a good framework to discover additional CGI-regulating factors and gain a better understanding of CGI regulation including complex associations and combinatorial binding. Similarly, the BANP reporter constructs provide a promising tool to disentangle the effects of CpG density, BANP binding, DNA accessibility, DNA methylation, and additional binders on promoter activity.
Understanding how CpG density contributes to (CGI) promoter activity constitutes one major step in ultimately predicting gene expression in development and disease
Motives and needs for physical activity among forcibly displaced people living in a refugee camp: A qualitative study
No full textThe living conditions of forcibly displaced persons are marked by traumatic experiences and persistent stress factors, necessitating simple and cost-effective health support methods. At present, many mental, physical, and social health problems are treated through sports activities. Examining the impact of the SALEEM sports program on their daily lives and well-being is crucial, considering personal successes, disappointments, and improvement suggestions for a comprehensive understanding of the program's effectiveness and potential enhancements. In the qualitative study, 38 participants of the SALEEM study were interviewed in a semi-structured self-recorded interview and the data was subsequently analyzed using content analysis. The sports program led to enhanced physical well-being, pain alleviation, and aided in weight loss. Additionally, the program also works to foster emotional well-being, decreasing depression, increase in happiness, and improving stress management. It can be said that the overall result was improved mental health. More so, it also promoted social competence by improving interactions, multicultural unity, and friendships, thereby alleviating loneliness. The program acted as a coping strategy by combating monotony, interrupting rumination, offering hope, and managing stress. Participants appreciated the available sports facilities but sought better conditions and daily training. The trainers' positive impact was acknowledged. The program bolstered self-confidence, structured daily routines, and motivation for well-being and academics, but faced challenges like interruptions to family duties affecting participation. In conclusion, the study highlights multiple effects of the sports program on participants: increased physical well-being, emotional well-being, improved social skills, behavioral adjustments, and proven coping strategies
Environmental enrichment promotes sparse coding in hippocampus via increased dendritic inhibition
No full textEnvironmental enrichment has been shown to improve hippocampus-dependent spatial learning and memory. However, how enrichment influences hippocampal network activity remains still largely unclear.
Here we used cFos labeling, in vivo Ca2+ imaging and in vitro electrophysiology to study the effect of environmental enrichment onto hippocampal network activity. First, we showed that exploration of novel context leads to increased cFos activity in CA1 and DG in animals housed under both standard and enrichment conditions. Remarkably, the number of cFos cells during exploration was lower in enriched animals. Additionally, we recorded Ca2+ responses from CA1 pyramidal cells with miniature microscope from mice exploring familiar and novel contexts. The analysis of cell responses revealed lower firing frequency in PCs of enriched animals. This indicates that enrichment does not increase overall hippocampal activity but improves sparse coding in hippocampus.
To understand the underlying mechanisms, we explored the possibilities that inhibitory interneurons are involved. We found that enrichment leads to increased activation of dendrite-targeting somatostatin-expressing (SOM) interneurons during exploration, which was linked to changes in the activity levels of principal neurons in CA1. This is not due to changes in intrinsic properties of SOM interneurons, but due to larger number of glutamatergic synapses onto SOM interneurons. At the functional level, it is accompanied by increased synaptic connectivity between pyramidal cells and SOM interneurons, as we show by performing whole-cell patch-clamp recordings from SOM interneurons while stimulating pyramidal cell axons in brain slices from control and enriched mice. Furthermore, we observed increased lateral feedback inhibition in enrichment in vitro.
Using simultaneous in vivo Ca2+ imaging from pyramidal cells and optogenetic silencing of SOM interneurons in CA1 in mice during spatial exploration, we found that SOM interneurons have a big impact on the firing frequency of PCs and the number of active cells. Most importantly, the effect of silencing on the number of active cells is stronger in enrichment.
Taken together, our data show that environmental enrichment decreases the size of hippocampal cell assemblies via enhanced recruitment of dendrite-targeting SOM interneurons
Le ragioni dell’espressione sintattica del topic, tra anteposizione sintattica, dislocazioni e costruzioni passive. Registro, testualità, struttura frasale. In analisi testi divulgativi contemporanei: un approccio qualitativo
No full textVom Musikalisch-Schönen zur Philosophie der neuen Musik. Spuren der Hanslick-Rezeption bei Theodor W. Adorno
No full textGestaltung eines virtuellen Museumsraumes unter Berucksichtigung positiver Einflussfaktoren auf die Intention zum physischen Museumsbesuchs. Eine Proof-of-Concept-Arbeit am Beispiel des sic! Elephanthouse in Luzern
No full textCultivating Citizenship through Cultural Terrains: A Study of the NEOJIBA Project’s Complementary Registers of Emancipation in Bahia, Brazil
No full textThis study investigates NEOJIBA, a socio-pedagogical music initiative in Bahia, Brazil, analyzing its impact on urban development and the emancipation of socially vulnerable youth. Employing a multidisciplinary approach, the research examines how NEOJIBA’s blend of cultural and material practices promotes emancipation through orchestral music, critical pedagogy, and community engagement, exploring themes of marginality, territory, culture, and socialization in citizenship development. Central to the analysis is NEOJIBA's operational structure and its diverse activities, which are focused on promoting autonomy, collaboration, and critical engagement with one’s territory, assessing their influence on participants' trajectories. Conclusively, the thesis illustrates how socially and culturally engaged practices, finely attuned to specific territorial contexts, facilitate the attainment of a more substantive citizenship that transcends its legal framework. It thus contributes to scholarly debates on the spatial and cultural construction of citizenship, advocating for a conceptualization based on everyday practices and ideas of urban agency, in line with perspectives from southern urban theorists like Holston and Caldeira. Furthermore, it informs discussions on urban policies and infrastructures aimed at mitigating youth marginalization, emphasizing how different material, social, and cultural supports act in complementarity to foster emancipation
Dynamic environmental conditions recorded by the trace fossil Teichichnus and event beds during deposition of the basal Vaca Muerta Formation in the central Neuqu´en Basin (Argentina)
Get PDFThe mud-dominated part of the basal Vaca Muerta Formation in the central Neuquén Basin accumulated mainly under dysoxic conditions as indicated by the absence of both fine lamination and large burrows, typical of anoxic and oxic conditions, respectively. Event beds and large, strata-bound concretions preserving a nearly uncompacted sediment fabric document rather discontinuous deposition, in particular as concretions must reside within the zone of carbonate supersaturation for a prolonged period of time to form. The mudrock hosts numerous ash layers. Among them is a bed up to 40 cm-thick that was deposited by storm-induced currents and later became calcified. It contains abundant, winding Teichichnus up to 220 cm-long. The trace producers partly reworked their own spreite and probably applied a gardening and sequestrichnial nutritional strategy (that is collecting and stowing of nutritional material in the burrow for later utilization). The spreite fill documents depositional processes that delivered argillaceous sediment and provided oxygen. The ash originally constituting the Teichichnus bed originated from the volcanic arc west of the basin. In contrast, the erosive surface limiting the Teichichnus bed is covered by mud, implying sediment delivery from the east or south where rivers entered the basin while the climate was (at least seasonally) humid, whereas aridity increased towards the north. Consequently, an estuarine circulation likely developed and prevailed during the initial transgression of the lower Vaca Muerta Formation while cold Palaeo-Pacific water flowed into the subsiding and expanding basin. Later, during prevailing dry climate, an anti-estuarine circulation was probably established
A high-resolution monitoring approach of urban CO2 fluxes. Part 1 - bottom-up model development
No full textMonitoring carbon dioxide (CO2) emissions of urban areas is increasingly important to assess the progress towards the Paris Agreement goals for climate neutrality. Cities are currently voluntarily developing their local inventories, how-ever, the approaches used across different cities are not systematically assessed, present consistency issues, neglect the biogenic fluxes and have restricted spatial and temporal resolution. In order to assess the accuracy of the urban emis-sion inventories and provide information which is useful for planning local climate change mitigation actions, high res-olution modelling approaches combined or evaluated with atmospheric observations are needed. This study presents a new high-resolution bottom-up (BU) model which provides hourly maps of all major components contributing to the local urban surface CO2 flux (i.e. building emissions, traffic emissions, human respiration, soil respiration, plant respi-ration, plant photosynthetic uptake) and can therefore be used for direct comparison with in-situ atmospheric obser-vations and development of local scale atmospheric inversion methodologies. The model design aims to be simple and flexible using inputs that are available in most cities, facilitating transferability to different locations. The inputs are primarily based on open geospatial datasets, census information, road traffic monitoring and basic meteorological pa-rameters. The model is applied on the city centre of Basel, Switzerland, for the year 2018 and the results are compared to a local inventory. It is demonstrated that the model captures the highly dynamic spatiotemporal variability of the urban CO2 fluxes according to main environmental drivers, population activity dynamics and geospatial information proxies. The annual modelled emissions from buildings and traffic are estimated 14.8 % and 9 % lower than the respec-tive information derived by the local inventory. The differences are mainly attributed to the emissions from the indus-trial areas and the highways which are beyond the geographical coverage of the model