VictorJuiz/thesis huggingface dataset:
Behaviour-dependent activity and synaptic organisation of septo-hippocampal GABAergic neurons

Animal behaviour is coordinated by biological rhythms over multiple timescales. In the temporal cortex, rhythmic oscillatory activity in the 5 to 12 Hz theta frequency range supports behaviour through temporal windows of changes in excitability and circuit computation. Subcortical inputs, including those from the medial septum are thought to generate the theta oscillatory activity. However, the rules of innervation of identified types of cortical cells in different cortical regions by a diverse population of septal neurons are largely unknown. During my DPhil studies, I tested the hypothesis that medial septal GABAergic neurons which exhibit distinct activity patterns would express distinct molecular marker combinations and have distinct synaptic partners.
In Chapters 1 and 2, I formulate my research questions and explain the methodology used. My main findings are presented in chapters 3 to 7. In Chapter 3, I present the strategy used to identify four populations of rhythmic medial septal neurons based on their spike-train dynamics in the mouse in vivo. In Chapter 4, I present a detailed analysis of the spike-timing and molecular parameters of a specialised population of rhythmic neuron, the Teevra cells. Teevra cells are PV+, do not change their firing rate from rest to running and fire short bursts of action potentials preferentially at the trough of both CA1 theta and slow irregular activity recorded in stratum pyramidale. In Chapter 5, I show that Teevra cells selectively innervate axo-axonic cells and CCK-immunopositive interneurons in hippocampal area CA3, bypassing CA1, CA2, and the dentate gyrus. In Chapter 6, I show that Teevra cell axons terminate in restricted septo-temporal segments of CA3 according to their rhythmicity. Finally, in Chapter 7, I present the Komal cells, which fire long bursts of action potentials at the peak of theta, increase their firing rate during running, and preferentially innervate extrahippocampal cortices. I discuss the broad relevance of my observations in Chapter 8.
Overall, my results show an unexpected specialisation in the organisation of medial septal GABAergic inputs to the temporal cortex. Teevra cells fire maximally at the trough and preferentially innervate the CA3 whereas Komal cells fire maximally at the peak and target extrahippocampal cortices, thereby coordinating the timing of converging glutamatergic inputs to CA1 via disinhibition. This cortical-region-specific GABAergic innervation by physiologically distinct septal neuronal types suggests a general mechanism for the coordination of network activity, both during regular theta oscillations and irregular large amplitude waves, providing windows of computation through synchronisation of neuronal activity in the cortex.

Advisor: Somogyi, P
Dupret, D
Interactions of perihperal membrane proteins with phosphatidylinositol lipids: insights from molecular dynamics simulations

Interactions between proteins and membranes are central to many signalling pathways and other cellular processes. Phosphatidylinositol phosphates (PIPs) are a family of lipids often acting as second messengers and targeted by peripheral proteins in these processes. A pipeline was developed combining the molecular dynamics (MD) approaches of umbrella sampling and coarse-grain modelling, and used to quantify and compare the interactions with PIP-containing model membranes of 13 pleckstrin homology (PH) domains, a common lipid-binding domain found in many proteins showing varied affinities and specificities for PIPs. Lipid selectivity generally agreed with previous observations. Several membrane-binding modes were identified, revealing PIP interactions through a secondary site are more common than suggested experimentally and appear to be related to overall affinity. Results suggest that simultaneous binding of multiple PIP lipids is required to achieve the high affinities characteristic of PH domains. Multiscale MD, combining coarse-grain binding simulations and atomistic refinement, was used to investigate PTEN, a tumour suppressor catalysing interconversion of PIPs and associated with many cancers and other disorders. Regions often ignored in previous studies were revealed to favour productive binding, largely via electrostatics. PIP clustering by bound PTEN and membrane insertion in the productive mode were demonstrated. Existence of an N-terminal PIP-binding site was supported, with this region appearing disordered, rather than helical as previously suggested. Changes in interdomain orientation when bound and with the clinically-relevant R173C mutation further suggest the importance of the interdomain interface for productive binding. Together, this work demonstrates the important contributions MD can make towards understanding protein/membrane interactions, particularly in the context of managing the diseases caused by their disruption.

Keywords: Phosphatidylinositol phosphate lipids
Molecular dynamics
Peripheral membrane proteins
Advisor: Sansom, M
Kalli, A
Security and privacy in app ecosystems

Smartphones are highly-capable mobile computing devices that have dramatically changed how people do business, interact with online services, and receive entertainment. Smartphone functionality is enhanced by an ecosystem of apps seemingly covering the entire gamut of functionality. While smartphone apps have undoubtedly provided immeasurable benefit to users, they also contribute their fair share of drawbacks, such as increases in security risks and the erosion of user privacy. In this thesis, I focus on the Android smartphone operating system, and pave the way for improving the security and privacy of its app ecosystem.
Chapter 3 starts by doing a comprehensive study on how Android apps have evolved over a three-year period, both in terms of their dangerous permission usage and the vulnerabilities they contain. It uncovers a trend whereby apps are using increasing numbers of dangerous permissions over time and at the same time becoming increasingly vulnerable to attack by adversaries.
By analysing the Google Play Store, Android's official app marketplace, Chapter 4 shows that many general-purpose apps can be replaced with functionallysimilar alternatives to the benefit of the user. This confirms that users still wield power to improve their own security and privacy. Chapter 5 combines this insight with real-world data from approximately 30,000 smartphones to understand the actual risk that the average user faces as a result of their use of apps, and takes an important first step in measuring the improvements that can be made.
Users, however, are not always aware of the risks they face and thus Chapter 6 demonstrates the feasibility of a classification system that can transparently and unobtrusively identify and alert users to the presence of apps of concern on their devices. This classification system identifies apps from features in the network traffic they generate, without itself analysing the payload of their traffic, thus maintaining a high threshold of privacy.
While the work presented in this thesis has uncovered undesirable trends in app evolution, and shows that a large fraction of users are exposed to non-trivial risk from the apps they use, in many cases there is suficient diversity in the offerings of general-purpose apps in the Google Play Store to empower users to mitigate the risks coming from the apps they use. This work takes us a step further in keeping users safe as they navigate and enjoy app ecosystems.

Keywords: app
privacy
security
smartphone
ecosystem
Advisor: Martinovic, I
Dynamics of temporal anticipation in perception and action

The selective deployment of attention over time optimises our perception and action at the moments when relevant events are expected to happen. Such "temporal orienting" to moments when something is going to happen is especially useful when this information can be combined with predictions about where and what events are likely to occur. A large body of research has already established how temporal predictions dynamically influence our perception and action, but questions remain regarding the neural bases of these attentional mechanisms. In this thesis I present three magnetoencephalography (MEG) studies that I conducted to investigate anticipatory neural dynamics associated with spatial-temporal orienting of attention for perception and action. I also investigate and discuss how such anticipatory dynamics change with ageing and neurodegeneration in Parkinson's disease (PD), and how these anticipatory neural dynamics behave in situations where a complex, hidden spatial-temporal structure is present. In Chapter 1, I introduce the topic of this thesis by reviewing the literature on temporal orienting of attention and by introducing my specific research questions. In Chapter 2, I present an MEG study on anticipatory neural dynamics of joint spatial-temporal orienting of attention in the visual domain, in younger and older adults. This study shows that neural dynamics with spatial, temporal and spatial-temporal orienting are all differentially affected by ageing. In Chapter 3, I describe an MEG experiment that investigates anticipatory neural dynamics during spatial-temporal motor preparation and compares PD participants to healthy control participants. This study reveals that both behavioural and neural dynamics with temporal orienting are affected in PD. In Chapter 4, I describe an experiment that explores how an implicit spatial-temporal structure is utilised to predict and prepare for upcoming actions. This study shows that motor cortical excitability is dynamically modulated in anticipation of the location and timing of events, even when such expectations are hidden in complex visual-motor sequences that remain largely implicit. In Chapter 5, the General discussion, I place these results in their wider context and discuss limitations and future directions.

Keywords: Parkinson's disease
Ageing
Temporal orienting
MEG
Neural oscillations
Attention
Advisor: van Ede, F
Nobre, K
Targeted cell-type specific gene expression profiling in a transgenic mouse model of Parkinson’s

Parkinson's disease is a neurodegenerative disorder that is pathologically characterised by the preferential cell death of vulnerable dopaminergic neurons (DAn) in the midbrain. Multiplications of the human alpha-Synuclein (aSyn) gene locus give rise to autosomal-dominant Parkinson’s and elevated aSyn expression has been linked to an increased risk to develop sporadic Parkinson's. Thus, aSyn provides an important link between familial and sporadic forms of the disease. This thesis brings together cutting-edge transgenic mouse models to examine early translational changes in the specifically vulnerable DAn. Translating ribosome affinity purification (TRAP) methodology was combined with a mouse model of Parkinson’s that over-expresses aSyn from the complete human SNCA locus. Immunoprecipitation of translated mRNA from midbrain DAn enabled the investigation of perturbations in gene expression early during disease progression. Next generation sequencing analysis of translated mRNAs determined the effect of aSyn over-expression on the translational profile of the "at-risk" population of DAn. This translatome analysis in THTR mice which express an eGFP-tagged ribosomal transgene (eGFP-L10a) under the endogenous mouse promoter of tyrosine hydroxylase (TH) identified a mitochondrial deficit in aged transgenic mice. In addition, a novel Cre-conditional TRAP line expresses eGFP-L10a inCre expressing cells upon crossing to a DAT-Cre driver. Pilot TRAP experiments successfully isolated translated mRNA from soma (midbrain) and axons (striatum) of DAn. Overall, data generated with the transgenic TRAP/Parkinson’s mouse lines will allow to obtain a more complete understanding of the underlying pathological pathways and translational changes during Parkinson's.

Advisor: Wade-Martins, R
Identification of novel Runx1 targets involved in HSC development

Haematopoietic stem and progenitor cells (HSPCs) are de novo generated within in the ventral aspects of the embryonic dorsal aorta (DA). Cells of this haemogenic endothelium (HE) will eventually undergo an endothelial to haematopoietic transition (EHT) that involves cell budding out of the aortic wall. Despite the detailed description of the cellular events, the exact haemogenic lineage path and the underlying molecular mechanism that establish full haematopoietic competence are still not entirely understood.
The transcription factor Runx1 is critical for the emergence of HSPCs and shows expression in the zebrafish HE as early as 24 hpf. To facilitate a detailed analysis of the transient HE population I generated a TgBAC(runx1P2:Citrine) reporter line under the control of the endogenous runx1 promoter on a bacterial artificial chromosome (BAC). Double-transgenic reporter lines for runx1 and the endothelial marker kdrl allowed us to isolate specifically cells of the DA away from the whole endothelial population, which could be further sub-divided into HE and non-haemogenic cells. Genomewide expression analysis within the respective tissues and upon Runx1 loss of function enabled the identification of HE-specific Runx1-regulated genes. Hereby, the gfi1ab gene appeared as the functional homologue of the murine Gfi1. I show that in zebrafish, EHT is orchestrated through a conserved Runx1-Gfi1-Lsd1 axis. The cellular functions of the remaining Runx1 targets imply that maturation into fully functional HSCs depends on epigenetic regulation due to the up-regulation of de novo DNAmethyltransferases, as well as on factors that allow the developing HSCs to respond to extrinsic cues from haematopoietic niches.
Lastly, it became evident that the early HE expresses dll4 at similar levels to the rest of the aortic endothelium, indicating a common lineage path. In the absence of RUNX1 the HE remains essentially arterial and persists as an integrated part of the DA.

Keywords: Hematopoiesis
Runx1
Advisor: Patient, R
De Bruijn, M
Development of viral vectors for passive immunisation against influenza

Seasonal influenza is a major global health threat causing more than 500,000 deaths annually. Pandemic influenza can arise through antigenic shift, or when novel influenza strains escape from animal reservoirs, and can affect a large geographical area leading to a large number of deaths. Neither prior infection, nor current vaccines, provide lasting protection due to the rapid antigenic evolution of influenza virus. The discovery of broadly neutralising antibodies (bnAbs) against influenza has opened the possibility of passive immunisation, which could provide protection against multiple influenza strains. Passive immunisation by parenteral administration of purified antibody is challenging, hindered by high antibody production costs and the short half-life of antibody in the circulation leading to relatively transient protection; however the use of viral vectors to deliver bnAbs can begin to address these limitations. Studies presented in this thesis investigated the feasibility of a novel, lung-targeting lentiviral vector pseudotyped with the fusion (F) and haemagglutinin-neuraminidase (HN) coat proteins from Sendai virus (rSIV.F/HN) as a gene transfer agent for bnAb expression. Recombinant SIV.F/HN vector expressing a secreted reporter protein Gaussia luciferase showed expression in the mouse lung lumen 1 month after lung administration. A single dose of rSIV.F/HN encoding novel anti-influenza bnAb (T1-3B) in a single Open Reading Frame directed expression of T1-3B into the lung lumen and systemic circulation, and conferred partial protection against lethal influenza challenge. Intramuscular (IM) delivery of a recombinant adeno-associated virus vector (rAAV2/8), previously reported to be effective against influenza challenge in mice, directed levels of T1-3B that conferred complete protection against influenza challenge. Daily repeat administrations of rSIV.F/HN did not improve protection against influenza challenge, but the ability of this vector to be repeatedly administered without loss of efficacy may be an advantage in potential clinical applications. Engineering of the bnAb, into the IgA1 backbone greatly enhanced the potency of several bnAbs and might improve influenza protection mediated by rSIV.F/HN. In conclusion, prophylaxis provided by gene transfer during an influenza pandemic might be feasible, providing long-lasting, more cost-effective and time-responsive protection than traditional vaccines or parenteral administration of therapeutic antibody.

Advisor: Hyde, S
Gill, D
Homogeneous catalysts for the synthesis of oxygenated polymers

This thesis describes the synthesis and characterisation of novel mono and dinuclear homogenous [Zn(II)] and [In(III)] metal complexes. Their applications as catalysts for CO2/epoxide or epoxide/anhydride ring opening copolymerisation and lactide ring opening polymerisation to generate polycarbonates and polyesters, respectively, are also reported.
Chapter 3 reports the first indium phosphasalen catalysts for CO2/cyclohexene oxide ring opening copolymerization. The catalysts are active at 1 bar pressure of CO2 and are most effective without any co-catalyst. It is also possible to use the complexes to isolate and characterise the key intermediates in the catalytic cycle. Kinetic and spectroscopic analyses show that polymerisation proceeds via a rare cis-mononuclear coordination- insertion mechanism.
Chapter 4 describes a series of mono and dinuclear zinc macrocycle catalysts with very high activities for the racemic lactide ring opening polymerisation. In most cases, the dinuclear zinc catalysts significantly out-perform the mono-zinc homologue. In addition, kinetic and spectroscopic investigations suggest a role for the ligand conformation in mediating rate. The catalysts perform very well under immortal conditions and operate at low catalyst loading, whilst conserving high activities.
Chapter 5 presents four dinuclear zinc acetate salen catalysts for the ring opening copolymerisation of CO2/cyclohexene oxide and phthalic anhydride/cyclohexene oxide. The catalysts show moderate activities for CO2/epoxide copolymerisation but are highly active for epoxide/anhydride copolymerisation. Structure/activity relationship studies reveal that the more flexible and electron donating ligand displays the highest activity. Poly(ester-b-carbonate)s are also afforded using the most active catalyst in terpolymerisations of anhydride/epoxide/CO2.

Keywords: Stereoselectivity
Polyester
Zinc
CO2
Homogeneous catalysis
Salen
Ring Opening Copolymerization
Dinuclear
Polycarbonate
Lactide
Indium
Isoselectivity
Epoxide
Mononuclear
Ring Opening Polymerization
Phosphasalen
Mechanistic studies
Advisor: Williams, C
Placement of fluid viscous dampers to reduce total-building seismic damage

Nonstructural damage has been found to critically influence economic losses and building downtime following earthquakes. Attaining a target level of seismic performance mandates the harmonisation of structural and nonstructural performance. Retrofitting buildings with fluid viscous dampers (FVDs) can improve interstorey drifts and floor accelerations, parameters which characterise seismic demands. The distribution of dampers within a building and the amount of supplemental damping are critical decisions. Many placement methods have been proposed, however no conclusive optimal method has been identified. The objectives of the thesis included benchmarking the performance of code-compliant buildings, investigating the optimal amount of damping, and comparing major damper placement methods. The seismic performance of Eurocode-compliant concentric braced frame buildings was benchmarked. Structures were modelled in OpenSees and the FEMA P-58 procedure was used to assess seismic performance in repair costs, a comprehensive measure of total-building performance. Buildings may be demolished following an ultimate limit state earthquake due to high repair costs. Storeys satisfying the Eurocode drift limit for nonstructural protection nevertheless experienced drift- and acceleration-sensitive damage. Most damage was attributed to nonstructural systems. Acceleration-sensitive damage is of comparable or greater consequence than drift-sensitive damage. These findings should be more appropriately reflected in structural design procedures. The optimal amount of damping to minimise repair costs was identified as 30-40%, larger than previously suggested levels based on structural parameters. Six damper placement methods were evaluated using linear FVDs. No method produced optimal results for both drifts and accelerations. Iterative methods that purport to optimise performance did not achieve that objective: local rather than global parameters are considered, and optimising for a single parameter may worsen another that impacts damage. The storey shear strain energy method and uniform damping produced repair costs more favourable than, or equal to, the other placement methods. Damper placement optimisation may be successful for high-rise or irregular structures.

Advisor: Williams, M
Blakeborough, T
An investigation into the structural role of the CCR4-NOT complex in mRNA stability

The CCR4-NOT complex is a global regulator of gene expression which controls mRNA levels by removing the poly-(A) tail, a step known as deadenylation and one that constitutes the rate-limiting step in mRNA decay. The human complex is comprised of eight stably associated CNOT subunits where CNOT1 forms the scaffold onto which CNOT2-11 bind. Although much has been learnt about the CCR4-NOT complex, questions still remain. Thus, this study focused on a number of sub-complexes of CNOT subunits and associated proteins to determine the mode of interaction with a hope to explore the mechanism of deadenylation by the CCR4-NOT complex. Firstly, the complex of CNOT10:CNOT11, found only in higher eukaryotes, was reconstituted for the first time using recombinant proteins. Crystallisation trials, limited proteolysis and mass spectrometry were used to isolate novel interaction regions between CNOT10 and CNOT11 which may provide direction for future structural and functional studies. Secondly, the interaction between CNOT9 and TTP was characterised. TTP is a RNA-binding protein which targets inflammatory mRNAs for deadenylation by recruiting the CCR4-NOT complex. This study highlights novel interactions between TTP and both CNOT2 and CNOT9. Moreover, BioLayer interferometry (BLI), peptide arrays and site-directed mutagenesis identified that TTP interacts with CNOT9 in a tryptophan-mediated manner. These findings change the known interface between TTP and the CCR4-NOT complex. Lastly, the MultiBac system was used to reconstitute a number of human CNOT sub-complexes, one of which was shown to effectively degrade a poly-(A) substrate, demonstrating it is enzymatically active. This achievement provides a tool for the future study of the CCR4-NOT complex. In summary, this study highlights novel interactions and characterises previously unknown binding mechanisms between CCR4-NOT subunits which expands our current understanding of the complex.

Keywords: Expression of protein complexes
Deadenylation
Inflammatory gene expression
Protein expression
Advisor: Burgess-Brown, N
Yue, W
Dean, J