This work investigates the influence of mutations at selected positions on the structure formation of the Alzheimer’s disease peptide amyloid β. Amyloid β is a member of the class of intrinsically disordered proteins that can aggregate into fibrils, which are characterized by a highly stable secondary structure, called cross-β structure. A central contact during fibrillation is the hydrophobic F19-L34 contact, which is located within the core of the cross-β structure. Modifications of this contact are known to influence the local molecular structure whereas the fibril morphology and the cross-β structure remain stable. In contrast, toxicity of amyloid β was completely lost for all previously investigated mutants of F19 and L34. This work characterizes the properties of this contact and answers the question what the minimally tolerated modifications are. To characterize the structure, structure formation process and biological activity of the Aβ variants a set of experiments was carried out. The local structure and dynamics were investigated using NMR experiments focusing on 13C-chemical shift changes and 1H-13C dipolar couplings, respectively. The fibril morphology and cross- β structure was verified by electron microscopy, circular dichroism spectroscopy and X-ray diffraction. Toxicity and biological activity was investigated using complementary cell culture experiments. The work was divided in three parts. First, L34 was substituted with three highly similar amino acids: the isomer isoleucine, valine that is one methylene group shorter but also a branched chain amino acid and the stereoisomer D-leucine. The L34 position proved to be important for the initiation of the structure formation, oligomer stability, fibril growth and the biological activity of amyloid β. These characteristics and properties were highly sensitive also to minor modifications but the different mutants showed no specific but qualitatively similar effects. The second part complemented previous mutation studies of the F19 position. Four new mutants were designed testing mild modification of the F19-L34 contact: phenylglycine and the homophenylalanine (S)-2-amino-4-phenyl-butyric acid change the length of the side chain, cyclohexyl-alanine eliminates the π-aromaticity of the ring system and increases the 3D steric demand, and (1-naphtyl)-alanine increases the 2D steric demand while maintaining the aromaticity. Mutations at the F19 position caused qualitatively similar effects as L34 modifications but proved to have quantitatively greater impact. Furthermore, they showed some specificity as steric constraints caused larger changes than modifications of the ring system. The third part investigates the influence of β-methylamino-L-alanine (BMAA) substitutions at positions F19, S8, and S26. The serine to BMAA substitutions were included because of their potential medical relevance. A F19BMAA substitution caused similar effects like other modifications at this position. Replacement of serine lead to a structural reorientation of the Aβ N-terminus and turn region. Furthermore, the pathways of the cell response changed from mitochondrial activity and plasma membrane integrity to apoptosis and neuronal stress reaction. Summarizing, it could be shown that, although the formation and structure of amyloid β fibrils is robust against different modifications the fibrillation kinetics, local structure and especially biological activity is highly sensitive and to some extend specific to even minor modifications.

Human infants rely on social interactions to acquire culturally relevant knowledge about their environment. Aside from active participation (“first-party perspective”), infants encounter social interactions through third-party observation (“third-party perspective”). Despite the absence of own involvement, the mere observation of others’ interactions represents an essential source of social learning opportunities. The overarching aim of this dissertation was to deepen our understanding of the foundations of infants’ observational learning from third-party interactions. This was achieved by investigating (a) social attentional developments and motivational influences driving infants’ attention toward third-party interactions (Study 1 & 2), and (b) factors influencing infants’ attention and memory while observing third-party interactions (Study 3). Study 1 investigated how infants’ attentional orienting to third-party interactions develops in parallel with their active social attention behavior. In Experiment 1, 9.5- to 11-month-old infants looked longer than 7- to 8.5-month-olds at videos showing two adults engaging in a face-to-face interaction, when simultaneously presented with a non-interactive back-to-back scene showing the same people acting individually. Moreover, older infants showed higher social engagement (including joint attention) during parent-infant free play. Experiment 2 replicated this age-related increase in both measures and showed that it follows continuous trajectories from 7 to 13 months of age. These findings suggest that infants’ attentional orienting to others’ social interactions coincides with developments in their social attention behavior during own social interactions. Study 2 examined the incentive value of social interactions as a proximal driver of infants’ attentional orienting to third-party interactions. In a gaze-contingent associative learning task, two geometrical shape cues were repeatedly paired with two kinds of target videos showing either a dyadic face-to-face interaction or a non interactive back-to-back scene. We found that 13-monthold infants performed faster saccadic latencies and more predictive gaze shifts toward the cued target region during social interaction trials. This suggests that social interaction targets can serve as primary reinforcers in an associative learning task, supporting the view that infants find it intrinsically rewarding to observe others’ social interactions. Study 3 investigated infants’ object encoding in the context of observed social interactions. In Experiment 1, 9-month-old infants were presented with four types of videos showing one object and two adults. The scenarios varied regarding the eye contact between the adults (eye contact or no eye contact) and the adults’ object directed gaze (looking toward or away from the object). Infants showed increased object encoding, but only when seeing two adults looking at an object together, following mutual eye contact. We found an identical pattern of results in a matched first-party design during which 9-month-old infants were directly addressed by one single adult on screen (Experiment 2). Together, these findings suggest that the capacity to learn about novel objects by observing third-party interactions emerges in the first postnatal year, and that it may depend on similar factors as infants’ learning through direct social interactions at this age. The findings of all three studies are integrated in a general discussion. In summary, the results of this thesis suggest that, throughout the first year after birth, infants develop abilities and preferences enabling them to approach and efficiently learn from third-party social interactions.:General Introduction 1 1.1 The Infant As an Active Learner 2 1.2 Social Attentional Requirements of Infants’ Learning From Social Interactions 3 1.3 Motivational Mechanisms Affording Opportunities to Learn From Social Interactions 16 1.4 Infants’ Learning From Social Interactions 21 1.5 Research Gaps 26 1.6 Focus of This Dissertation 27 Study I 33 2.1 Introduction 34 2.2 Experiment I 38 2.3 Experiment II 45 2.4 General Discussion 50 Study II 55 3.1 Introduction 56 3.2 Methods 57 3.3 Results 63 3.4 Discussion 64 Study III 69 4.1 Introduction 70 4.2 Experiment I 74 4.3 Experiment II 81 4.4 General Discussion 86 General Discussion 91 5.1 Summary of Results 91 5.2 Research Contributions 93 5.3 Limitations and Future Directions 101 5.4 Overall Conclusion 114 References 115 Appendix A – Supplementary Materials Study I 137 Appendix B – Supplementary Materials Study II 148 Appendix C – Supplementary Materials Study III 152 Curriculum Vitae 167 Scientific Publications and Conference Contributions 169 Contributions of Authors 171 Declaration of Authorship 175

Der Hirschkäfer ist der größte und markanteste heimische Käfer und in vielen Regionen sehr selten geworden. Diesen typischen Bewohner alter Laub- und Obstbäume sowie ihrer Stubben und Wurzeln stellt die reich bebilderte Broschüre vor. Sie macht auf die Gefährdungsursachen aufmerksam und enthält zahlreiche Hinweise, wie Waldbesitzer und Eigentümer oder Nutzer von Grundstücken zur Erhaltung des Lebensraums und damit auch gleichzeitig zum Schutz des Hirschkäfers und vieler anderer Tier- und Pflanzenarten beitragen können. Redaktionsschluss: 02.02.2022

Cancer is the second leading cause of death in females. According to the American Cancer Society, there are 327,660 new cases in breast and gynecological cancers estimated in 2014, placing emphasis on the need for cancer prevention and new cancer treatment strategies. One important approach to cancer prevention involves phytochemicals, biologically active compounds derived from plants. A variety of studies on the impact of dietary compounds found in cruciferous vegetables, green tea, and spices like curry and black pepper have revealed epigenetic changes in female cancers. Thus, an important emerging topic comprises epigenetic changes due to the modulation of noncoding RNA levels. Since it has been shown that non-coding RNAs such as microRNAs and long non-coding RNAs are aberrantly expressed in cancer, and furthermore are linked to distinct cancer phenotypes, understanding the effects of dietary compounds and supplements on the epigenetic modulator non-coding RNA is of great interest. This article reviews the current findings on nutrition-induced changes in breast and gynecological cancers at the non-coding RNA level.

In contrast to mammals, zebrafish possess a high regenerative capacity even within the central nervous system including the retina. In the retina, lesions result in a loss of neurons and the subsequent activation of Müller glia, the retinal stem cells. Müller glia-derived progenitors differentiate into all types of retinal neurons. Eventually the anatomical tissue architecture is restored within a few weeks or months depending on the degree of lesion. However, little is known about how retinal lesions impair vision functionally, as well as how and to what extent visual function is restored during the course of regeneration, in particular in adult animals. Here, quantitative behavioral assays were applied to assess visual function during homeostasis and regeneration in adult zebrafish. While the dark-light preference test has been identified to be too variable to assess visual function, a novel visiondependent social preference test as well as an automated two level OKR-based vision test have proved to be more suitable. Two different lesion paradigms have been compared in this study: light-induced lesions that target photoreceptor cells and neurotoxin-mediated lesions that act in a concentration-dependent manner either affecting only inner retinal neurons or all retinal cell types. It could be shown that visual function was significantly impaired after light lesion but fully though gradually recovered within 28 days. Vision for easy conditions with high contrast and low level of detail, as well as color vision, was restored around 7–10 days post lesion while vision under more demanding conditions - with low contrast and high level of detailwas regained only from 14 days post lesion onwards. The corresponding retinal morphology was monitored in vivo using optical coherence tomography and revealed a close correlation between structure and function. Injection of neurotoxins resulted in a multitude of morphological changes that impeded the analysis of functional recovery. In general, restoration of visual function after neurotoxin-mediated lesion was limited and highly dependent on the initial degree of damage. As suggested by the behavioral studies, visual functions are, at least partially, restored after lesion. However, no conclusions can be drawn on the restoration of the underlying retinal circuitry. Hence, large-scale electrophysiological recordings using multielectrode arrays have been established for the adult zebrafish retina allowing the recording of spontaneous and light-driven activity. This technique can be exploited to analyze the restoration of signal processing of the retina in future studies. In addition, the established tools can be applied to characterize visual mutants and models of retinal degenerative diseases both morphologically and functionally as exemplarily demonstrated here for the blind atoh7 mutant. It was shown that the adult atoh7mutant largely resembles the larval phenotype and displays distinct alterations in optic tectum structure as well as impairments of social behavior. Altogether, the established vision-dependent behavioral assays suggest that the degree and kinetics of functional recovery are highly dependent on the lesion paradigm as well as quality of morphological restoration. In combination with the established multielectrode array recordings, the field is provided with new readouts to study functional recovery that can be used in addition to existing criteria to assess regeneration of the adult zebrafish retina. The established toolbox can furthermore be exploited to analyze retinal function not only in a regeneration context but also in visual mutants, disease models or ageing. Eventually, a deeper understanding of the regenerative processes in zebrafish will provide new perspectives for the development of therapies for retinal degenerative diseases.

Grazing by large herbivores has the potential to provide multiple ecosystem services, depending on multiple factors. Through a range of interdisciplinary methods, including literature reviews, case studies, and semi-structured interviews, I seek to assess how grazing can contribute to sustainable landscape management, as well as how sustainable practices can be incentivised. In chapter one, I focus on the role of grazing in wildfire prevention. Through a systematic literature review, I uncover new insights into the effectiveness of large herbivores in mitigating wildfires. I find that large herbivores can reduce wildfire frequency by promoting grass-dominated landscapes and reduce fire intensity by consuming vegetation and creating landscape features that reduce fuel loads. However, the effectiveness of large herbivores depends on the type of vegetation and diet preferences of the animals, and management practices associated with livestock grazing can also influence fire ignition. In chapter two, I investigate the relationship between grazing management and ecosystem services through a literature review, focusing on synergies and trade-offs between them. I identify management practices that are beneficial to multiple ecosystem services as well as policy mechanisms that can incentivise them. In the third chapter, I use case studies to investigate the role of domestic animals in rewilding projects. I describe how domestic and semi-wild herbivores can contribute to rewilding projects and make them more inclusive for rural and indigenous communities. In the fourth chapter, I aim to identify the challenges faced by land users in performing sustainable grazing management through semi-structured interviews with 88 land users from eight case studies in Europe. I gain insights into land users' determinants of behaviour towards sustainable grazing practices using the Behaviour Change Wheel framework. Overall I suggest that grazing can be a potential solution to the challenges of the Anthropocene, but only when done in the right way. I emphasize the importance of an interdisciplinary approach in grazing research and considering socio-ecological systems. I also highlight the potential of rewilding and semi-wild grazing systems especially in areas undergoing land abandonment. The results imply a shift in meat production and consumption and potential new pathways for human-livestock relations. Finally, agricultural policies such as the Common Agricultural Policy (CAP) can play a crucial role in incentivizing sustainable grazing management and should be improved to support extensive grazing and extensification.:Table of Contents i. General Introduction 6 i.i Context 7 i.i.i European grazed landscapes can provide high biodiversity and supply various ecosystem services 7 i.i.ii Challenges for European grazed systems 9 i.ii Thesis methods and outline 13 i.ii.i Literature reviews (chapters 1 & 2) 13 i.ii.ii Case studies (chapters 3 and 4) 13 i.ii.iii General approach and methodological considerations 18 i.ii.iv Research aims and questions 18 i.ii.v Thesis outline 19 i.iii Graphical abstract 21 1. Chapter One 22 1.1 Introduction 23 1.2 Literature search 25 1.3 Overall effects of herbivores on wildfire risks 26 1.3.1 Effects of herbivores on wildfire frequency 26 1.3.2 Effects of herbivores on fire intensity or severity 30 1.4 Effects of herbivores on fuel loads and fire hazard 30 1.5 Effects of vegetation characteristics on fire regimes 33 1.6 Effects of grazing-associated management on wildfires 33 1.7 Implications for management 34 1.8 Implications for policy 36 1.9 Conclusions and future challenges 37 2. Chapter Two 38 2.1 Introduction 39 2.2 Literature review: general methods 42 2.3 Results 44 2.3.1 Effects of grazing on biodiversity 44 2.3.2 Effects of grazing on climate change and its mitigation 46 2.3.3 Effects of grazing on soil quality 48 2.3.4 Effects of grazing on wildfire mitigation 49 2.3.5 Effects of grazing on cultural ecosystem services 51 2.3.6 Synergies and Trade-offs of different grazing practices and ecosystem services 52 2.3.7 Recommendations: optimizing ecosystem multifunctionality in grazed systems 55 2.4 Outlook 59 3. Chapter Three 60 3.1 Introduction 61 3.2 The Broader Theory of Rewilding and Potential Role for Livestock 64 3.2.1 Stochastic Disturbance Regimes 64 3.2.2 Trophic Complexity 65 3.2.3 Interacting Processes 66 3.3 General Case Studies 67 3.3.1 Reindeer Engineer in Swedish Lapland 67 3.3.2 Livestock Fire Brigade and Free Running Horses in the Côa Valley, Portugal 68 3.3.3 Ecotourism and Sustainable Meat at Knepp Estate, England 70 3.3.4 Oostvaardersplassen: The “Wild Experiment” 71 3.4 Conclusions 73 4. Chapter Four 77 4.1 Introduction 78 4.2 Methods 80 4.2.1 Interviews 80 4.3 Findings from interviews: Determinants of sustainable grazing decisions 81 4.3.1 Physical capability: the impact of the ‘rural exodus’ 81 4.3.2 Psychological capability: access to knowledge and understanding of administrative rules influences how land users conduct their management 82 4.3.3 Physical opportunity 83 4.3.4 Social opportunities 85 4.3.5 Motivation 85 4.4 Discussion 87 4.4.1 Intervention functions to support sustainable grazing, and associated policies 88 4.5 Conclusions and outlook 92 5. Discussion 93 5.1 Importance of the scientific, social and political context of the research 93 5.2 Societal relevance of the thesis 93 5.3 Policy relevance of grazing research 94 5.4 The value of transdisciplinary research 94 5.5 Contribution of the research 95 5.5.1 Contribution of chapter 1: large herbivores and wildfire mitigation 95 5.5.2 Contribution of chapter 2: grazing and multiple ecosystem services, trade-offs and synergies in a European context 96 5.5.3 Contribution of chapter 3: domestic livestock and rewilding 97 5.5.4 Contribution of chapter 4: drivers of decision making for sustainable grazing 98 5.5.5 Interdisciplinary approach to grazing research, grazing areas as socio-ecological systems 99 5.6 Limitations 100 5.6.1 The framework of ecosystem services 100 5.6.2 Limitations to rewilding research 101 5.6.3 The challenge of addressing different socioecological and administrative scales 101 5.6.4 Practical limitations 103 6. Outlook 104 6.1 Future research needs 104 6.2 Pathways for sustainable grazing 104 7. Conclusion 107 References 110 Summary 135 Zusammenfassung 139 Appendix 143 Appendix Chapter One 144 Appendix Chapter Two 145 Appendix Chapter Four 149 Acknowledgements 153 Author contribution statement 155

The effect of six heavy metals, namely, silver (Ag), lead (Pb), palladium (Pd), copper (Cu), nickel (Ni), and chromium (Cr), on phosphorus (P) uptake by yeast was investigated by single-cell analysis using inductively coupled plasma time-of-flight mass spectrometry (SC-ICP-TOF-MS). It was found that the P content in cells with 1.55 g L1 P feeding after P starvation was increased by 70% compared to control cells. Heavy metals at 10 ppm, except Cu, had a negative impact on P accumulation by cells. Pd reduced the P content by 26% in single cells compared to control cells. Metal uptake was strongest for Ag and Pd (0.7 1012 L cell1) and weakest for Cr (0.05 1012 L cell1). Exposure to Cr markedly reduced (50%) Mg in cells and had the greatest impact on the intrinsic element composition. The SC-ICP-TOF-MS shows the diversity of elemental content in single cells: for example, the P content under standard conditions varied between 12.4 and 890 fg cell1. This technique allows studying both the uptake of elements and sublethal effects on physiology at a single-cell level.

Focused ultrasound (FUS) is a non-invasive technique producing a variety of biological effects by either thermal or mechanical mechanisms of ultrasound interaction with the targeted tissue. FUS could bring benefits, e.g., tumour sensitisation, immune stimulation, and targeted drug delivery, but investigation of FUS effects at the cellular level is still missing. New techniques are commonly tested in vitro on two-dimensional (2D) monolayer cancer cell culture models. The 3D tumour model—spheroid—is mainly utilised to mimic solid tumours from an architectural standpoint. It is a promising method to simulate the characteristics of tumours in vitro and their various responses to therapeutic alternatives. This study aimed to evaluate the effects of FUS on human prostate and glioblastoma cancer tumour spheroids in vitro. The experimental follow-up enclosed the measurements of spheroid integrity and growth kinetics, DNA damage, and cellular metabolic activity by measuring intracellular ATP content in the spheroids. Our results showed that pulsed FUS treatment induced molecular effects in 3D tumour models. With the disruption of the spheroid integrity, we observed an increase in DNA double-strand breaks, leading to damage in the cancer cells depending on the cancer cell type.

Orderliness, speed, and rhythm in biochemistry are vital for cellular function. In order to achieve this, cells implement compartmentalization via several methods, one of which is the formation of membrane-less compartments. These compartments, often referred to as “biomolecular condensates”, are understood to be formed by separation of proteins and other biomolecules into dense and dilute phases. While the formation of the resulting protein-rich condensates is fundamental for spatiotemporal organization of biochemistry within the cell, a vast majority of proteins found to phase separate in vitro do so at a concentration an order of magnitude above their endogenous expression levels. Recently, a theoretical study has shown that membrane binding of phase separating proteins can result in phase separation spatially occurring at the membrane well below bulk saturation concentrations. However, much remains unknown about the formation mechanism and function of these condensates. To that end, for my doctoral project, I used a synthetic system composed of supported lipid bilayers decorated with lipid-bound NTA(Ni) to allow for coordination and thus membrane binding of the well-characterized protein FUS via a C-terminal His-tag. Through this model system I found that 2D phase separation of FUS could occur an order of magnitude below the experimentally determined bulk saturation concentration. FUS was able to form dense and dilute phases in 2D and the transition point to form these phases could be controlled by modulating buffer conditions. Additionally, membrane-bound FUS condensates were able to further recruit FUS from the bulk to form a multilayer of protein through a prewetting transition. With this characterization of 2D phase separation of FUS, I then explored a physiologically relevant protein in the form of PAR-3, a fundamental protein of the PAR polarity system, which is necessary for the establishment of polarity in the C. elegans zygote. I found that full-length PAR-3 was able to phase separate under physiological salt conditions with a Csat of 100nM. Further, I identified a C-terminal predicted prion-like domain to act as a driver for phase separation. Additionally, I determined PAR-3’s affinity and specificity for PI(4,5)P2 and found that it could form 2D condensates upon binding to the membrane at physiological concentrations. Furthermore, these condensates were able to recruit PAR-6 alone and PAR-6 in complex with PKC-3 to the membrane, ultimately resulting the reconstitution of the anterior PAR complex which is known to exist in a condensed clustered form in vivo. Taken together, this work provides insight into a mechanism where phase separation can be locally triggered by membrane binding under sub-saturation concentration, offering a robust and potentially universal mechanism by which cells can spatially control phase separation and pattern cellular membranes.

Der Hirschkäfer ist der größte und markanteste heimische Käfer und in vielen Regionen sehr selten geworden. Diesen typischen Bewohner alter Laub- und Obstbäume sowie ihrer Stubben und Wurzeln stellt die reich bebilderte Broschüre vor. Sie macht auf die Gefährdungsursachen aufmerksam und enthält zahlreiche Hinweise, wie Waldbesitzer und Eigentümer oder Nutzer von Grundstücken zur Erhaltung des Lebensraums und damit auch gleichzeitig zum Schutz des Hirschkäfers und vieler anderer Tier- und Pflanzenarten beitragen können.