VictorJuiz/thesis huggingface dataset:
Spectral tuning of Archaerhodopsin 3 in Escherichia coli

Microbial rhodopsins are proteins that act as light driven proton pumps located in the plasma membrane of eukarya, bacteria and archaea. These proteins facilitate the movement of protons by light-induced isomerisation of a central retinal chromophore found in the transmembrane core of the opsin protein. Here we detail an extensive study of amino acid substitutions in the retinal binding pocket of archaerhodopsin 3 (AR3) and their effect on the absorbance spectrum of the protein for use in biotechnological applications.
In this thesis, protein engineering was used to generate a mutant library that produced hypsochromic and bathochromic shifts when purified from an E. coli model into a detergent environment. The largest shifts reported here (-25 and +7 nm compared to wtAR3) were achieved through strategic mutation of residues interacting with the retinal chromophore. The S151C mutant produced in this report perturbed the photocycle kinetics in numerous detergent systems tested, resulting in a delayed photo-intermediate decay and return to ground state.
The faster photocycle kinetics of AR3 compared to other rhodopsins makes it a promising target for both optogenetic and bio-sensory applications. For its use as a single molecule bio-sensor, a strategic cysteine mutation was incorporated into the intracellular EF loop of the protein to allow for its future immobilisation onto a conductive surface. With monomeric AR3 adsorbed onto a gold substrate, it can be assessed for its ability to generate a detectable proton current in response to activation by light of a wavelength specific to each mutant. For applications in optogenetics, distinctly shifted mutants can allow for targeted activation of different mutants in the same neural system, thus allowing for the assaying of more complex pathways. Red-shifted mutants are of particular interest in this field as they allow for non-invasive light penetration in the brain.

Advisor: Watts, A
Coherent combination of mismatched fibre lasers - steps towards real world applications

Large arrays of coherently-combined Quasi-Continuous-Wave (QCW) fibre lasers have been identified as a plausible solution to creating high-repetition rate (~1 kHz), high-wall-plug efficiency lasers capable of driving a wakefield for accelerator applications. The coherent combination of four near-identical Photonic Crystal Fibre (PCF) lasers to produce laser pulses with pulse energies of 5.7 mJ and peak powers of 22 GW has been successfully demonstrated by other groups. Extensions of the four PCF experiment also demonstrated the coherent combination of 8 photonic crystal fibres to attain pulse energies of 12 mJ and peak powers of 35 GW. However, these PCFs were near-identical in length, core size, fibre structure and dopant concentration. Work done in Oxford focused on the coherent combination of fibre lasers which were unmatched in length, core size, internal structure and dopant concentration. The aim was to identify the point at which the fibre differences caused the combining efficiency to degrade. Potentially, this information would be used to identify the tolerance levels required for the large-scale manufacture of the fibres needed to construct a facility-scale laser system for laser-wakefield acceleration.
This thesis covers the coherent combination of mismatched PCFs, a scenario which could occur during the construction of a facility-scale terawatt laser system. Here, hundreds or potentially thousands of PCFs could be purchased from different manufacturers, each of whom use their own methods of manufacture and tolerances. An investigation into the challenges associated with combining two highly-mismatched PCFs was performed, with the PCFs chosen to be very different to exaggerate any effects which could degrade their coherent combination. This work resulted in the successful coherent combination of these highly-mismatched PCFs, with a measured degree of linear polarisation of 85.9 ± 0.5 % and a fraction of available beam power of 93.0 ± 0.5 %. Also, supporting work investigated the difference between simulation and measurement in explaining the observed degree of linear-polarisation of the combined-beam.

Keywords: Filled aperture combination
Ultrafast Laser
Photonic crystal fibre
Laser
Advisor: corner, L
Walczak, R
Characterisation of an “eat-me” signal on dying and HIV-1-infected T cells

The clearance of dying cells by phagocytes, termed efferocytosis, is regulated by a combination of “eat-me” and “don’t eat-me” signals expressed on the dying cell surface. Efferocytosis is essential for ontogeny and tissue homeostasis, but can also mediate the spread of pathogen infection, and its dysregulation leads to inflammatory disorders. The Sattentau laboratory recently reported that the macrophage efferocytic uptake of HIV-1-infected CD4+ T cells drives efficient infection of macrophages. However, the signals involved in the macrophage capture of the infected T cell remain undefined, but cell death appears to be an important component of this recognition.
In this study a novel efferocytic pathway which is based on the cleavage of the heavily sialylated glycoprotein CD43 acting as a potent ‘don’t eat-me’ signal for macrophages is investigated. Data presented in this thesis imply that; i) CD43 shedding is mediated by ADAM10 which is activated by T cell death, and ii) cleavage of CD43 as a don’t eat-me signal from the surface of dying and infected T cells triggers their engulfment by macrophages.
In the context of HIV-1infection, interventions in this pathway may have the potential of reducing targeted macrophage infection. Also, the knowledge of this novel efferocytic pathway has general implications for understanding and potentially treating unwanted inflammatory disorders.

Advisor: Sattentau, Q
Russell, R
The role of enhancers in regulating the transcription cycle

Enhancers are key determinants in the spatiotemporal control of gene expression which underlies cellular differentiation and disease; however, the dominant mechanisms through which they exert their effects on genes remain poorly understood. In order to achieve fine control over nascent transcript levels, transcription is highly regulated at many stages. It has been suggested that enhancers may act on many specific stages in transcription to modulate transcript production. The idea that initiation of transcription is the principal barrier in achieving changes in gene expression has been challenged by the discovery of promoter proximal pausing of RNA polymerase II (Pol 2 pausing) at the majority of protein coding genes in addition to further downstream regulatory steps.
In this thesis I sought to determine the predominant mechanism by which enhancers bring about differential gene expression in vivo by examining knockout models of well-defined enhancers at the alpha and beta globin gene clusters. Using a wide variety of complementary methodologies, I demonstrate that these two enhancer clusters promote transcription initiation whilst having minimal effects on downstream transcriptional kinetics. In addition, I build on these findings to show that genome-wide differential transcription initiation best correlates with differential gene expression and is likely the predominant regulatory step in transcription. Pol2 pausing is detectable genome-wide but scales according to initiation frequency suggesting this process is an inherent kinetic property of mammalian promoters, but its role in gene regulation may be less significant than previously thought.
This study highlights the role that enhancers play in regulation of specific stages in transcription to achieve tissue-specific gene expression. These findings may be applicable to the broader study of metazoan transcription, given the highly conserved nature of transcriptional mechanisms and will enable a better understanding of how eukaryotic gene expression is altered development and disease.

Advisor: Hughes, J
Higgs, D
In situ study of atmospheric stress corrosion crack initiation in aluminum 6061-T6 and particulate SiC reinforced nanocomposite

This project aimed to study the atmospheric stress corrosion cracking (SCC) behavior of 6061 T6 aluminum alloy and a composite based on the same alloy, reinforced with SiC nano particles.
The materials were machined into taper-shape specimens, and on each specimen, three salt deposits were applied at the center and two ends. The SCC experiment was conducted at a temperature of 40 ºC and a relative humidity of 30%. The 6061 T6 aluminum alloy specimens and the composite specimens were tested at an applied constant load of 948±20 N and 1050±20 N respectively.
Cracks were only observed at the center salt deposit location of 6061 T6 aluminum alloy specimens. It is observed that pre-pitting accelerates the development of strain, but does not affect the final strain that is developed in the SCC test. There was no measurable strain change at the end location of all the 6061 T6 aluminum specimens, indicating that there is a threshold of applied stress or strain for stress corrosion cracks to develop.
For the aluminum based composite, no crack was found on the specimens. The applied strain of the center location of the composite specimen is equal to that of the end location of the aluminum specimen, within the error range, and it is concluded that the addition of SiC nano particles may make the composite material more susceptible to stress corrosion.
DIC technique was adopted to measure the displacement and strain change of specimens, and an effective method to monitor the initiation and propagation of small cracks has been developed. SEM and X-ray tomography were used to observe specimens after SCC testing, verifying the DIC result.

Advisor: Marrow, T
Galano, M
Security analysis of behavioural biometrics for continuous authentication

In recent years, behavioural biometrics have become increasingly popular, with many types of behaviour being explored for the purpose of user authentication. Some of the most common examples are keystroke dynamics, mouse movements, touchscreen inputs and human gait.
Unlike physiological biometrics (e.g., fingerprints), behavioural biometrics are often believed to be relatively hard for adversaries to collect, but nevertheless have been subject to active attacks, including presentation, signal injection and imitation attacks.
In this thesis, we take a holistic view on the design, evaluation and security analysis of behavioural biometric recognition systems. First, we underline their usefulness by designing a novel authentication system based on distinctive eye movement behaviour. We evaluate this system under different adversary models and show that eye movements can be used for both user authentication and judging a user's task familiarity. Drawing from insights gained from this project, we go beyond the state of the art to develop metrics and methodologies that more accurately reflect a system's real-world performance and security. This approach is centred around reflecting a biometric's systematic false negatives (i.e., attackers that consistently go undetected) more accurately.
A frequent focus of related work is how to present previously obtained biometric data to a behavioural authentication system (e.g., through imitation or mimicry attacks). However, the challenge of obtaining this data in the first place is far less explored. In this thesis, we perform a series of experiments to judge the usefulness of biometric data collected through a variety of sources. The idea is to measure the security impact of the plethora of biometric data that is involuntarily created through our day-to-day interactions with diverse systems.

Keywords: security
eye movements
authentication
biometrics
Advisor: Martinovic, I
Rasmussen, K
15th-century printed Italian editions of Aesopian texts

My thesis provides the first comprehensive and detailed account of the impact of the introduction of movable-type printing on the production and circulation of Aesopian texts in Italy in the second half of the 15th century.
It highlights how the introduction of the new medium did not correspond, at least within the first decades, to a standardisation in the written circulation of the rich Aesopian corpus; on the contrary, it fostered the composition of new versions, translations and rewritings, contributing to enhance its varied and multiform nature.
The goal of my thesis is to unveil this process, and to examine the parallel development of textual and figurative traditions of Aesopian texts in the earliest editions produced in Italy, in order to reassess their place in the wider context of the circulation of Aesopian texts in print.
Chapter I introduces the main features of the manuscript transmission of those among Aesopian texts, which ended up in print in the first years after the introduction of the new medium. It also focuses on the cultural and historical circumstances of the production of the first editions in the European context.
Chapter II presents, in chronological order, all the texts included in the Italian incunable editions, contextualising their authors, their main textual and stylistic features and the circumstances of their composition.
Chapters III and IV respectively examine the evolution of the tradition of the different Aesopian texts in print and the evolution of the figurative and iconographic cycles of images illustrating Italian incunable editions of the Vita and the Fabulae. They present a synthesis and assessment of the first-hand data gathered in the Catalogue included in the appendix.
The Catalogue in appendix offers a detailed and complete overview of the characteristics of all seventy Aesopian editions printed in Italy in the 15th century. Each edition is analysed with special attention to its bibliographic and textual features, its illustrations, and provenance information concerning the copies examined.
The Conclusions focus on the originality of certain texts created specifically for the press, on the people who played an important role in the composition and printing of Aesopian texts, on the identification of new lines of transmission in print, on the distinction and analysis of different iconographic lines of descent in the illustrated editions, and on the discovery of noteworthy cases of provenance evidence, which suggest how the consumption of Aesopian Italian incunables transversely engaged people at different social and cultural levels.

Advisor: Dondi, C
Risk and care in an aging United States

In the United States, an ‘aging crisis’ has been brewing around the rising number of older persons and the relations of care their longer lives require. New forms of neoliberal governance, which seek to manage this aging disaster, task elders with the individual responsibility to prepare and to control their aging futures. Through an ethnographic case study at Hollenbeck Palms, a Continuing Care Retirement Community in Los Angeles, this dissertation investigates how a risk framework shapes the everyday, affective experiences of aging through the examination of contracting care, rewriting kin relations, mobility practices, and playing bingo. It explores how strategies of care at once reflect this regime of risk and also defy and transcend it.
Through the analysis of elders’ and caregivers’ lives at Hollenbeck Palms, this dissertation contributes three insights: a logic of risk does not control aging futures, but instead generates even more uncertainty and insecurity in elders’ lives; uncertainty is not only an indelible part of aging, but is also actively generated by elders; and contrary to the notion that the uncertainty of aging renders elders and caregivers into passive victims, elders and caregivers can learn to live alongside the uncertainty of aging care-fully. This dissertation rethinks aging in the twenty-first century United States as a site of generative possibility, instead of a site of danger.

Advisor: Pratten, D
Investigating the role of Dis2 phosphatase in RNA Polymerase II transcription termination in fission yeast

The end of the RNA polymerase II (Pol II) transcription cycle is a tightly regulated process that requires: (i) dissociation of elongation factors, (ii) recruitment of the 3' end processing machinery that is essential for premRNA cleavage and polyadenylation, and (iii) association of termination factors to dislodge Pol II from the DNA template. A characteristic feature of termination factors is that they directly interact with the phosphorylated C-terminal domain (CTD) of the largest subunit of Pol II (Rpb1). The CTD consists of heptad repeats (Y1S2P3T4S5P6S7) where Tyr1, Ser2, Ser5, Ser7 and Thr4 can be reversibly phosphorylated during the transcription cycle. The work presented here reveals that Thr4 phosphorylation levels are enriched at the 3' ends of protein-coding genes in fission yeast and identies Dis2 as a Pol II-CTD phosphatase that dephosphorylates Thr4 both in vitro and in vivo. Dis2 activity is required for Thr4 dephosphorylation at the polyadenylation site, for recruitment of termination factors Pcf11, Rhn1 and Seb1 and for normal Pol II termination. Interestingly, Seb1 cotranscriptional recruitment is shown to be dependent not on Dis2 activity on CTD but on its ability to dephosphorylate the elongation factor Spt5, suggesting that Dis2 regulates the elongation-to-termination transition by targeting multiple substrates.

Advisor: Vasilieva, L
The spatio-temporal regulation of Polo-like kinase 1 in DNA repair

The G2/M cell cycle kinase Polo-like kinase 1 (PLK1) was recently discovered to play an active role in DNA repair by phosphorylating the RAD51 recombinase to aid homologous recombination. This occurs at a much shorter timeframe than the well-characterised downregulation of PLK1 activity via the DNA damage checkpoint. In this thesis, I expand upon this by investigating the regulation of PLK1 during this earlier timeframe post-damage. Laser microirradiation experiments observe fluorescently-tagged PLK1 localising to DNA damage sites within 15 minutes, in both S/G2 and, unexpectedly, G1 cells. In S/G2, this recruitment is dependent on PLK1's polo-box domain binding sites primed by the cyclin-dependent kinase CDK2. Activity of the DNA damage response kinase ATM is also required for PLK1's localisation to damage in both phases of the cell cycle. Meanwhile, a proteomics approach identified novel DNA repair-related PLK1 binding partners, including the E3 ubiquitin ligase UBR5 and the protein kinase DYRK2. DYRK2 increases in association with PLK1 within 30 minutes post-damage and can induce PLK1 activation by phosphorylation of its T-loop. These findings shed light onto how PLK1 is regulated to carry out its repair function, open up new avenues of future investigation into this topic, and even suggest the possibility of PLK1 playing further as-yet-uncharacterised roles in DNA repair.

Advisor: Esashi, F
'Sur le patron de plusieurs bons autheurs italiens’: poetics, performance and politics in French comic drama (1540–1580)

This thesis is the first study of the reinvention of French comic theatre from 1540–1580 and of the productive and at times antagonistic relationship of French playwrights with Italian paradigms. Using close textual and historical analysis, my study documents the move from classical comedy regarded as a scholarly exercise to drama revived in print and performance, which was anything but a smooth transition. I employ as a framework for this exploration the three material conditions of poetics, performance, and politics, which connect and overlap to create an intertextual and cross-cultural survey of the changing status of French comedy and of the key role played in its reinvention by Italian sources. In terms of poetics, I trace the difficulties faced by France’s earliest playwrights and theoreticians in their bid to prove comedy to be a viable mode of humanist literary production, examining the apathetic and sometimes contentious discussions of their contemporaries regarding comic theatre and its Italian exemplars. Moving to performance, I consider the effects of Italian comic performances in France throughout the mid-century, demonstrating how these productions acted as persuasive evidence to French writers that comedy was worthy of a place within their national literature. My enquiry into politics evaluates how the reverence of French playwrights towards their Italian sources turned into resentment across the course of the century. I argue that as French writers grew increasingly skilled at composing Italianate comedy, they began to use the genre against the Italians themselves, employing theatre not only as a means of resisting the influx of Italian culture and politics into France but also as a canny means of replacing the Italians as arbiters of European literature. Ultimately I challenge the conveniently linear narrative of early modern comic theatre created by previous scholars, contending that the development of comedy in France — along with its reliance on the Italian model — was fraught with complexities and tensions which have long been overlooked.

Advisor: Cooper, R
McLaughlin, M
A genetic modifier of B6.YPOS gonadal sex reversal on chromosome 13

Sex determination is the process by which the embryonic gonad differentiates into either a testis or an ovary. When the mammalian gonad first arises in the embryo, the cells comprising it are bipotential: they can adopt a testiscular or ovarian fate. Such commitment is governed chromosomally, principally by the presence or absence of the Y chromosome. Testis determination in the mouse is initiated by the expression of the Y-linked gene, Sry. The timing of Sry expression is critical: if there is any delay in the onset of expression, such that a threshold level is not reached before 11.5 days post coitum (dpc), then the outcome will be XY ovotestis or ovary development. Disrupted Sry expression and gonadal sex reversal can arise due to mutations in genes encoding key regulators (WT1, GATA4, MAP3K4, GADD45γ etc.) and also from a putative functional ‘mismatch’ between the Y chromosome and the autosomes/X chromosome. Such a situation arises following introgression of the domesticus Y chromosome of Mus poschiavinus into the C57BL/6J (B6) genetic background, resulting in so-called B6.YPOS sex reversal. While examining the molecular basis of B6.YPOS sex reversal, a region of chromosome 13 in the B6.YPOS colony at MRC Harwell was found to be heterozygous for sequences not derived from B6. This region modifies the B6.YPOS sex reversal phenotype, promoting testis development and male fertility. This thesis describes experiments aimed at understanding this locus, attempting to identify its origins, and elucidating the mechanism by which it acts. It also reports on an investigation into the genes within the region and their possible role in sex determination.

Advisor: Greenfield, A