VictorJuiz/thesis huggingface dataset:
Regulation of CFTR function and stability by phosphorylation and its membrane environment
Keywords: channel activity
cystic fibrosis
cystic fibrosis transmembrane conductance regulator
lipids
phosphorylation
stability

Cystic Fibrosis is caused by mutations of the gene that codes for Cystic Fibrosis Transmembrane conductance Regulator (CFTR), an ATP-Binding Cassette anion channel. The major disease-causing mutation, deletion of phenylalanine at position 508 (F508del), results in misfolding, mistrafficking and loss of CFTR at the cell surface where CFTR normally functions to maintain proper hydration of epithelia. Protein kinase A (PKA) phosphorylation enhances CFTR function at the cell surface. However, not much is known regarding the structural consequences of full-length CFTR upon PKA phosphorylation. Towards this, I developed biophysical methods to study purified full-length CFTR and showed that PKA phosphorylation modifies its overall structure, possibly at the transmission interface between coupling helices and nucleotide binding domains. Channel function studies with disease-causing mutations at the transmission interface further support the role of transmission interface interactions for phosphorylation-mediated channel gating. Recent structural studies have found that PKA phosphorylation causes subtle changes in zebrafish CFTR structure. However, the channel remains closed. We propose that this may be due to the detergent environment of the protein. We developed a novel purification method that maintained CFTR: lipid interactions. We conducted ATPase and iodide efflux assays to show that purified CFTR associated with lipids resulted in significantly higher CFTR activity than purified CFTR in detergent micelles. These experiments suggest involvement of lipids in regulating CFTR function. Potentiator drugs, like ivacaftor, enhance CFTR function at the cell surface. I conducted biochemical experiments to find that high concentrations of ivacaftor non-specifically destabilized F508del-CFTR and other membrane proteins. This non-specific destabilization of ivacaftor appears to be associated with its lipophilicity which disrupts the membrane. This further highlights the importance of lipids for stabilizing and modifying CFTR function in the lipid bilayer. In summary, CFTR function and stability are regulated by PKA phosphorylation and lipids of the membrane environment of CFTR.
Scramble for Photographs of Empire: Complicating Colonial Visions of the Philippine American War (1899 - 1913)
Keywords: Colonial
Photography
Postcolonial
Visual Culture

This dissertation explores how a scramble for photographs of the Philippine American War helped capture a more complicated history of the United States empire in the Philippines from 1898 to 1913; preserving traces of Americans’ emotions, Filipinos’ resistance, and less common sights of colonial violence. As the war in the Philippines faded from American historical consciousness during the twentieth century, a few well-known colonial officials’ ethnographic photographs, aftermath images of dead Filipinos, and cliché phrases such as “civilize ‘em with a krag” or “the water cure” summarized it. In contrast, this dissertation’s case studies extend the work of photo theorists Laura Wexler, Tina Campt and others, to the Philippines, arguing that re-framing colonial representations reveals more complexity in colonial relations, same-sex sexual violence, and Filipino peoples’ resistance or a less overt refusal to accept colonial domination. The first case study re-situates one photograph of dead Filipino soldiers in the broader history of colonial relations on the ground in 1898 and 1899, demonstrating how the image documents same-sex sexual violence and what anti-imperialist Harvard professor of psychology William James denounced as American martial hysterics. Next, the dissertation goes beyond existing histories of photography and the Philippines to argue that many more individuals contributed to a multivalent, layered, visual discourse about the islands during an initial scramble for souvenirs and visual trophies of empire in 1898 and 1899. Looking at how images of the Philippines shifted increasingly towards depictions of progress and the continued need for American governance after 1899, the third case study considers what is often outside the frame of the U.S. government’s preferred depictions of benevolent assimilation. Undermining the social death implicit in racial typing, as Laura Wexler advocates in a different context, the final case study analyzes the colonial uncanny, or the secretly familiar resistance, refusal, and variation in American colonial structure on the ground. Moving beyond traditional analyses of how dominant gazes racialize, sexualize and objectify, this dissertation demonstrates how Americans scramble for photographs of empire created a more complicated visual discourse, recording Americans’ emotions, systemic and continuing violence, as well as different forms of Filipino resistance.
Le Truchement : Représentation d’une Instance de Médiation dans les Récits de Voyageurs Français du XVIe au XVIIIe Siècle
Keywords: Ancien Régime
Interprètes
Littérature de voyage
Médiation
Traduction
Truchements

Sous l’Ancien Régime, le terme « truchement » renvoie aux interprètes et médiateurs intervenant lors des contacts entre les Européens et les peuples étrangers. Toutefois, malgré leur rôle fondamental dans les échanges, ces personnages restent dans l’ombre et sont à peine mentionnés dans les textes de la plupart des voyageurs. À travers une analyse d’ensemble et diachronique du corpus viatique français, cette thèse répond à deux objectifs principaux. D’une part, elle vise à fournir un portrait global des interprètes à travers les informations présentes dans les récits. D’autre part, elle cherche à identifier les procédés employés pour dépeindre la relation qui s’établit entre les voyageurs et leurs truchements afin de mieux comprendre la représentation complexe de cette relation. Après une réflexion sur la façon dont les interprètes sont recrutés par les voyageurs lors de voyages au long cours, nous analysons les différentes méthodes employées dans la formation linguistique des médiateurs. Ceci nous permet d’aborder la question du niveau de compétence des interprètes. En suivant le point de vue du voyageur qui, seul, détermine l’écriture de la relation, nous analysons les différentes interactions, satisfaisantes ou non, qui influencent la représentation des interprètes. En effet, l’image qui nous est fournie de ces personnages fantomatiques donne aussi un portrait révélateur des voyageurs, dont la tendance à l’auto-valorisation et à l’héroïsation est déterminante pour leurs relations avec leurs interprètes. Lorsque la situation ne se prête pas à l’héroïsation, une autre stratégie prend le dessus : la représentation des voyageurs comme victimes méritant la pitié des lecteurs. Ces postures, qui fonctionnent en miroir l’une de l’autre, déterminent les procédés d’écriture mis en place dans l’établissement du portait des truchements. Ainsi, en ciblant les médiateurs, cette thèse fournit également un aperçu des enjeux de la traduction et du processus d’écriture du genre viatique durant l’Ancien Régime.
Three-dimensional Hyaluronan-based Hydrogels Designed for Breast Cancer Spheroid Formation
Keywords: Biomaterials
Cancer
Drug Screening
Hyaluronan
Hydrogel
Oxime

In vitro culture of cancer cells is traditionally performed with two-dimensional (2D) culture, which fails to model the in vivo tumour cell phenotype and microenvironment. Hyaluronan (HA) is upregulated in breast cancer stroma and thus an excellent starting material in which to culture breast cancer cells; however, HA itself is a viscous liquid. To create a 3D gel for the prolonged culture of breast cancer cells we invented a crosslinked hyaluronan hydrogel comprised of HA-aldehyde and HA-ketone crosslinked with poly(ethylene glycol)-oxyamine. In designing this hydrogel, we considered hydrogel stability, transparency, and mechanical and chemical tunability. These HA-oxime crosslinked gels enabled the long-term culture of breast cancer spheroids (CS). First, hydrogel stiffness was decoupled from the immobilization of cell-adhesive peptides using orthogonal oxime and Diels-Alder click chemistry. Breast CS formation on bifunctional HA hydrogels is dependent on hydrogel stiffness and immobilized peptide. Breast cancer cells treated with the chemotherapeutic doxorubicin were less sensitive compared to cells grown on Matrigel™, a complex protein extract without HA. Breast CS expressed multidrug resistance protein 1, a drug efflux transporter on bifunctional HA hydrogels, and showed decreased doxorubicin drug penetration. Secondly, HA-oxime hydrogels were designed with tunable gelation to encapsulate breast cancer cells. We compared the culture of 5 different cell lines in vitro to in vivo xenografts in mice, thereby benchmarking the gene profile of in vitro cultured cells in our HA-oxime hydrogels to xenografts and relative to more traditional culture strategies of 2D and in Matrigel™. Using these gene expression profiles, we probed drug sensitivity and demonstrated greater predictability in HA-oxime hydrogels than other strategies. Moreover, we demonstrated the capacity to grow patient-derived cells in these HA-oxime hydrogels, paving the way for future opportunities in personalized medicine. Collectively, in these studies we developed and benchmarked in vitro breast cancer models and demonstrate the importance of cell microenvironment when investigating chemotherapeutics.
Let’s Liberate the Bullers: Queer Activism and Community-making in the Anglophone Caribbean
Keywords: Caribbean
homoimperialism
human rights
LGBTIQ+
queer
transgender

This dissertation interrogates the work being done by activists and non-governmental organizations in the Anglophone Caribbean, and theorizes that current interventions fail to adequately address the complicated ways that queer people negotiate and resist homophobia and transphobia in the region. In this work I draw on transnational feminist, Black queer theory, and Caribbean studies frameworks to posit that queer sexual praxes extend beyond dominant human rights tropes of disease, mortality and the imperative to escape a violently homophobic region. I am also particularly interested in the ways that working class gay men and trans people create communities of exile within the region as a radical praxis of space making despite being deeply affected by politically and culturally sanctioned homophobia and transphobia. My multi-sited ethnographic study is based on fieldwork conducted between 2016 and 2018 in Barbados, Guyana, Jamaica and Trinidad and Tobago. In these sites I conducted extensive online ethnography and participant observation, as well as semi-structured interviews with representatives from prominent queer advocacy groups. I also collected life histories from trans and gender non-conforming persons in order to better understand how they make life in the region. My scholarship offers an interdisciplinary intervention that engages and extends transnational feminist theory, critical race theory, human rights theory, queer theory, anthropology, and Caribbean studies by focusing explicitly on the under-interrogated area of queer sexual and gender politics in the Anglophone Caribbean. This project starts by examining the nature of queer human rights organizing by activists from Toronto’s Caribbean diaspora, aided by financial and administrative support from human rights organizations in Canada. I then interrogate the ways that such activities materialize in the four research sites, and some disconnects between the work being done by funders and activists located in Toronto, and the dynamics on the ground in each space. Finally, I explore three important moments across the four research sites that disrupt human rights narratives of a violently homophobic Anglophone region.
Investigations of Tau-metal Interactions
Keywords: Alzheimer's Disease
Electrochemistry
Metal Interaction
Microtubule Binding Repeats
Protein Aggregation
Tau Protein

This thesis describes the investigation of metal-tau interaction and speculates about its possible role in the pathogenesis of Alzheimer’s disease. The investigation started by developing a surfaced-based electrochemical sensor to monitor the conformational changes of htau39 on the gold surface as a consequence of Fe2+ and Fe3+ interaction, which is the first account of monitoring the conformation changes of tau film using electrochemistry. This work depicts the ability of surface-based electrochemistry to study the biomolecular interaction including metal-protein and protein-protein interactions. This technique was also applied to study the influence of phosphorylation on the iron-tau interaction, which showed the possible role of tyrosine in Fe2+ interaction. The ESI-MS experiments provide substantial evidence for the metallation of the longest isoform of tau with Cu2+, Zn2+, Fe2+, and Fe3+, which further support our findings from surface-based electrochemistry. This work also confirms the impact of phosphorylation on metal interaction. The possible role of metal ions in Alzheimer’s was reinforced by the formation of tau aggregates in the presence of metal ions, and ROS generation in the case of Cu2+, and Fe2+. Focusing on the interaction of metal ions with full-length peptides of the microtubule binding repeats R1 – R4 was an opportunity to gain more insight into the possible implication of d-block metal ions in the interaction with the tau protein. In particular, the involvement of Cys-based redox chemistry in the interaction of Cu2+ with R2 and R3 leads to the ROS formation and fibrils formation, which could not be prevented by ascorbate and glutathione as the most abundant anti-oxidants in neurons. This thesis also discusses the conformational and structural changes of R1 and R4 upon interaction with Zn2+, Fe2+, and Fe3+, and while ES-MS spectra confirmed the complexation of Zn2+ and Fe2+, there is no such evidence for Fe3+. Findings obtained from this thesis reinforces the idea that metal ions play a role in tau aggregation and ROS formation, which may contribute to cell death. These findings provide new insights for the design of new potential therapeutic agents.
Reciprocal Learning Partnerships between Elementary Mathematics Teachers: A Partnership between Canada and China
Keywords: Canada-China
Comparative Education
Cross-Cultural Collaboration
Elementary Mathematics
Narrative Inquiry
Professional Development

This study investigates the Reciprocal Learning Partnerships (RLP) between two pairs of elementary school mathematics teachers where each pair is comprised of one teacher from Changchun, China and one teacher from Toronto, Canada. These teachers are part of a seven-year reciprocal learning project focused on understanding the cultural and educational perspectives of Canadian and Chinese mathematics educators. Furthermore, this study explores how long term cross-cultural reciprocal learning partnerships can be used as a form of professional development for mathematics teachers. Data for this study was collected through interviews, classroom observations, videos, documents, journals and e-communication transcripts. These exploratory case studies focus on the teacher’s professional development needs and their personal narratives as they participate in reciprocal learning partnerships. Findings suggest Canadian and Chinese teachers have complementary strengths that make ideal reciprocal learning partners (RLP). Some findings include: (1) subject specific teacher knowledge, pedagogical knowledge, and knowledge of the learner are main areas of focus for Canada-China RLPs; (2) Canadian teachers were able to learn content knowledge from their reciprocal learning partners; (3) differentiated learning, especially for students with exceptionalities, is a common area of interest for both Canadian and Chinese teacher’s due to recent curriculum reforms in both nations; (4) cultural misconceptions are still present in Canadian and Chinese schools, and they can be challenged and corrected through reciprocal learning; (5) long-term RLPs can increase teacher efficacy; (6) RLPs are developed through three main stages where teachers learn about each other, learn from each other, and then learn with each other; (7) student-involvement in in-service teacher development can increase teacher motivation and accountability; (8) administrative and peer support are advantages to motivating RLPs; (9) when translations are involved in RLP communications, it is important to use tools that allow participants to express their personality and emotions; (10) portability, ease of use, and accessibility are important factors to consider when selecting technology for in-service teacher development. This research suggests that mathematics teachers from Canada and China are willing and eager to learn from each other to increase their teacher knowledge. Recommendations for future research are also outlined.
Novel Insights into the eIF2 alpha Integrated Stress Response on Microbial Infection, Inflammation and Proteotoxic Stress
Keywords: HRI
infection
innate immunity
stress response

The integrated stress response is an evolutionarily conserved pathway found in all eukaryotic cells, which allows for adaptation to environmental insults. At the same time, mammalian cells have intricate mechanisms to detect and respond to pathogens, however the interactions between the integrated stress response and innate immunity has become increasingly apparent. The findings in this thesis highlight novel interactions between the integrated stress response sensors GCN2 and HRI in innate immune signaling and cellular homeostasis. In Chapter 2, I highlight the impact of the integrated stress response by infection with several intracellular bacterial pathogens using cells which are stress irresponsive. I then compared the roles of the eIF2 alpha kinases GCN2 and HRI in inducing the integrated response and using the intracellular pathogens Shigella flexneri and Listeria monocytogenes as models, which revealed that although both sensors are activated during infection, they have different effects. While exploring these differences, I discovered that innate immune pathways which oligomerize require HRI for signaling and the mechanisms by which HRI mediates the signaling of these complexes is described in Chapter 3. Specifically, I found that the pattern recognition molecules NOD1, NOD2, TIFA, MAVs and NLRP3 all require HRI to induce inflammatory responses while TLR2, TL4R, TLR5 and STING-driven inflammation was unaltered. Interestingly, I found that the defect in HRI deficient cells to induce these signaling pathways is because of aberrant complex formation. I then determined that for oligomerizing pattern recognition molecules to assemble correctly, the chaperone HSPB8 is required. I also found that HSPB8 is regulated by the integrated stress response in an ATF4-HRI-eIF2 alpha dependent manner. I then explored the effect that HRI may have on more general proteotoxic stress in Chapter 4. I found that HRI is crucial in mediating protective responses to proteasome inhibition as well as against aggregating alpha-synuclein and TDP-43, which aggregate in Parkinson’s disease and Amyotrophic Lateral Sclerosis respectively. This body of work sheds light on a novel signaling pathway, which may lead to the development of alternate avenues for the treatment of autoinflammatory disease as well as neurodegenerative disease.
Vascularizing Tissues Ex Vivo: Engineering Microvascular Structures and Cellular Phenotypes
Keywords: bioprinting
endothelial cells
RNA-seq
stem cells
tissue engineering
vascularization

Numerous approaches to vascularizing engineered tissues ex vivo have been explored for relatively low-volume constructs; however, the development of the methods and biological understanding necessary to engineer a range of construct dimensions with physiologically-accurate microvasculature remains ongoing. This thesis focuses on several aspects of this challenge, including (i) the fabrication of vascular channels using 3D bioprinting, (ii) assessment of collagen, fibrin, and collagen-fibrin co-gels as scaffold materials, and finally (iii) exploring the suitability of iPSC-derived endothelial cells (ECs) for engineering vasculature. In the case of (i), a bioprinter was designed and sacrificial materials assessed for characteristics relevant to their use in patterning vascular channels within hydrogels (e.g., biocompatibility, mechanical properties). The most promising sacrificial materials, Pluronic F-127 (35% w/v) and gelatin-hyaluronan (5.25%, 3% w/v), had sufficient viscosity to create precisely patterned channels and exhibited no cytotoxicity. With regard to (ii), it was found that fibrin and co-gels facilitated EC sprouting, whereas collagen elicited migration of individual ECs, and that PKC-epsilon phosphorylation may play a role in regulating these differential responses. Overall, co-gels were found to be superior to either hydrogel alone as fibrin both enabled EC sprouting and impeded cell-mediated contraction, while increasing collagen within co-gels resulted in a progressively higher compressive modulus and enabled the formation of microchannels that could be perfused up to ~15 mmHg when composed of 2.5 mg mL-1 fibrin and 3 mg mL-1 collagen. In terms of (iii), RNA-seq was performed on iPSC-ECs and primary ECs (HUVEC, HIAEC, and HdMVEC) and revealed that iPSC-ECs have upregulated genes related to arterial hemogenic endothelium, indicative of possible immaturity. Additionally, differential expression of inflammation, hemostasis, shear-sensing and barrier function-related genes was discerned. Favourably, iPSC-ECs had an intermediate angiogenic capacity compared to native ECs. Overall, iPSC-ECs displayed promising similarities to native ECs, however, several key aspects of their function were identified as requiring further evaluation and improved maturation. Ultimately, this thesis has contributed to the advancement of biomaterials and the cell sourcing necessary for developing vascularized tissues and their concomitant clinical and scientific applications.