VictorJuiz/thesis huggingface dataset:
Evaluating the Interaction of HIV and the Immune System in Mucosal Tissues

90% of Human Immunodeficiency Virus (HIV) infections are acquired across the genital or gastrointestinal mucosa, and infection leads to profound depletion of CD4+ lymphocytes. Antiretroviral therapy can restore blood CD4+ T cells. However, immune dysfunction and defects in mucosal antimicrobial defence persist. Some CD4+ T-subsets, particularly antimicrobial Th17 cells, show enhanced susceptibility to HIV infection and are also preferentially depleted in the course of HIV infection; the latter may allow microbial translocation into the bloodstream. Genital infections have been shown to have direct mucosal immune effects and to increase susceptibility to HIV; however, the effect of systemic infections, such as Malaria (which is holo-endemic in some HIV prevalent regions) is unknown. Understanding the relationship between HIV, highly susceptible immune cells, immune activation and malaria infection on mucosal tissues has been the main focus of my thesis. In HIV-infected individuals, I explored whether HIV antiretroviral therapy restores gut Th17 populations and improves gut antimicrobial defences. Therapy restored gut Th17 populations in some, but not all individuals, but antimicrobial defence remained impaired. I then piloted a novel mucosal-optimized PCR assay to measure cervical immune gene responses, as standard mucosal assays are inadequate. I succeeded in measuring mitogen-induced, but not HIV-specific, cervical immune responses in HIV-infected individuals. Next, using this PCR platform I examined mitogen-induced cervical immune responses in individuals demonstrating reduced susceptibility to HIV, and found that they had reduced production of both Th17-associated and pro-inflammatory cytokines from cervical cells. Finally, in a murine model I found that malaria caused genital and gastrointestinal mucosal immune activation, and increased both the expression of mucosal HIV susceptibility immune markers, and mucosal T cell immune activation. In summary, insufficient gastrointestinal Th17 cells restoration does not underlie persistent mucosal immune activation and microbial translocation in HIV-infected people on therapy. A reduced frequency of highly susceptible Th17 cells in the cervix of HIV-exposed but uninfected individuals was identified as a correlate of reduced HIV susceptibility. Malaria, a common systemic infection in HIV-endemic countries, may enhance susceptibility to HIV through increasing putative immune markers of HIV susceptibility and immune activation in potential mucosal sites of HIV exposure.

Keywords: HIV
Mucosa
Advisor: Kaul, Rupert
The Role of Intercellular Contacts in EpithelialL-mesenchymal/-myofibroblast Transition

Epithelial mesenchymal/-myofibroblast transition (EMT/EMyT) has emerged as one of the central mechanisms in wound healing and tissue fibrosis. The main feature of EMyT is the activation of a myogenic program, leading to the induction of the α-smooth-muscle actin (SMA) gene in the transitioning epithelium. Recent research suggests that intercellular contacts are not merely passive targets, but are active contributors to EMT/EMyT. Indeed, our group showed previously that contact uncoupling or injury is necessary for TGFβ to induce EMyT (two-hit paradigm). Further, our previous work also revealed that Smad3, the main TGFβ-regulated transcription factor, binds to the Myocardin Related Transcription Factor (MRTF), the prime driver of SMA promoter, and inhibits MRTF’s transcriptional activity. During EMyT, Smad3 eventually degrades, which liberates the MRTF-driven myogenic program. However the mechanisms whereby cell contacts regulate the fate of Smad3 and MRTF during EMyT are poorly understood. Accordingly, the central aim of my studies was to explore the role of intercellular contacts, in particular that of Adherens Junction (AJs) in the induction of the myogenic reprogramming of the injured epithelium. This thesis describes two novel molecular mechanisms through which AJs impact EMyT. In the first part, we show β-catenin, an AJs component and transcriptional co-activator counteracts the inhibitory action of Smad3 on MRTF. Moreover we reveal that β-catenin is necessary to maintain MRTF stability via protecting MRTF from proteasomal degradation. Thus, β-catenin is an indispensable permissive factor for SMA expression. In the second part, we demonstrate that contact injury and TGFβ suppress the expression of the phosphatase PTEN. EMyT-related reduction or absence of PTEN potentiates Smad3 degradation. EMyT is associated with enhanced phosphorylation of the T179 residue in Smad3 linker region, and this event is necessary for Smad3 degradation. PTEN silencing increases the stimulatory effect of contact uncoupling and TGFβ on SMA promoter activity and SMA protein expression. Thus, the integrity of intercellular contacts regulates the level of PTEN, which in turn controls Smad3 stability through impacting on T179 phosphorylation. This new knowledge holds promises for targeted therapies and more effective prevention of the currently incurable fibroproliferative and fibrocontractile diseases.

Keywords: cell contacts
myofibroblast
myogenic program
EMT
Advisor: Kapus, Andras
Freight Market Interactions Simulation (FREMIS): An Agent-based Modelling Framework

Freight transport is the output of an economic market, which converts commodity flows into vehicle flows. Interactions in this market influence vehicle flows and since freight market characteristics (product differentiation and economies of scale/scope) violate perfect competition conditions, the output of this market cannot be predicted directly, unless these interactions are represented in the forecasting models. Traditional freight modelling frameworks do not consider these interactions and consequently they may provide inaccurate freight flow forecasts. In this dissertation, a freight modelling framework is proposed using simulation of freight agent interactions in the economic market to forecast freight flows. The framework is named FREMIS (FREight Market Interactions Simulation). The FREMIS framework consists of two demand models to represent shipper decisions (bundling of shipments and carrier selection) in the market and functions based on profit maximizing behaviour to simulate carrier proposals for contracts. Besides that, learning models are proposed to simulate agent learning processes based on their interactions. The framework was developed aiming to create a realistic representation of freight markets using feasible data collection methods. To illustrate the feasibility of the data collection, a customized web survey was implemented with shippers and carriers in a freight market. Two probabilistic models were developed using the data. The first model, a shipment bundling model was proposed combining a probabilistic model and a vehicle routing algorithm. The results of the probabilistic model are presented in this dissertation, where the locations of shipments (origin and destination) influence the probability of bundling them. Second, three carrier selection models were developed aiming to analyse the nonresponse bias and non-attendance problem in the survey. All of these models assumed heteroskedasticity (different scale or variance) in shipper behaviour. In all models, the hypothesis of agents’ heteroskedasticity cannot be rejected. Besides that, nonresponse bias and non-attendance problem were identified in the survey. In conclusion, the models obtained from the survey were consistent with their behavioural assumptions and therefore they can be adopted during FREMIS implementation.

Keywords: Freight Modelling
Agent-Based Modelling
Advisor: Roorda, Matthew J.
Transitions between Marine and Freshwaters in Fishes: Evolutionary Pattern and Process

Evolutionary transitions between marine and freshwater habitats are rare events that can have profound impacts on aquatic biodiversity. The main goal of my thesis is determining the processes involved in transitions between marine and freshwater biomes, and the resulting patterns of diversity using phylogenetic approaches. To test hypotheses regarding the geography, timing, frequency, and mechanisms regulating biome transitions I generated multi-locus time-calibrated molecular phylogenies for groups of fishes that include both exclusively marine and freshwater species. My analysis of anchovies demonstrated that Neotropical freshwater anchovies represent a monophyletic radiation with a single origin in South American freshwaters. I used a phylogeny of herring and allies (Clupeiformes) to investigate the evolution of diadromy, a migratory behavior in which individuals move between oceans and freshwater habitats for reproduction and feeding. These analyses do not support the hypothesis that differences in productivity between marine and freshwater explain the origins of diadromous lineages. Diadromy has been considered an evolutionary pathway for permanent biome transitions, however I found that diadromy almost never produces a fully marine or freshwater clade. Marine lineages often invade continental freshwaters during episodes of marine incursion. In South America, the rich diversity of marine derived fish lineages invaded during Eocene marine incursions from either the Pacific or the Caribbean, and Oligocene marine incursions from the Caribbean. I falsified the highly cited Miocene marine incursion hypothesis, but found that the Pebas Mega-Wetland catalyzed diversification in some marine derived lineages. Using diversification analyses, I investigated the evolutionary processes that have generated disparate patterns of diversity between continents and oceans. I found that freshwater silversides have higher speciation and extinction rates than marine silversides. Lineages accumulation plots suggest ecological limits are not regulating clade growth in either marine or freshwater biomes. Overall, biome conservatism is a widespread pattern among fishes, and this pattern is largely driven by competition in clades that are physiologically capable of biome transitions. Biome transitions are facilitated by rare paleogeographic events, such as marine incursions. Finally, a difference in net diversification rate is the macroevolutionary mechanism that best explains the difference in diversity between continents and oceans.

Keywords: macroevolution
biogeography
phylogenetics
fishes
Advisor: Lovejoy, Nathan
Mathematics Coaching to Improve Teaching Practice: The Experiences of Mathematics Teachers and Coaches

The purpose of the study is to determine how coaching can be used effectively to improve instruction and student achievement while exploring teachers’ specific emotions during mathematics education reform initiatives that challenge the teacher’s beliefs about teaching and learning in mathematics. It also examines how teachers incorporate the reform changes into their practice in order for the new instructional practices to have the expected effect. I explored teacher learning which refers to the correct use of reform strategies by mathematics teachers so that they have the intended effects on student achievement with the support of a coach during reform initiatives. Through questionnaires, interviews, observations and archival material, the study determines the relationship between teachers’ specific emotions, teacher learning and teacher coaching in secondary school mathematics classrooms. As a result, the study highlights the issues associated with the implementation of mathematics education reform initiatives and implications. The findings show that mathematics education reforms produce emotional responses that can be described as both negative and positive. For example, some emotions include pride, joy, fear, feeling drained and ineffective. The four teachers in the study experienced these emotions because of factors such as a lack of knowledge of how to implement mathematics reform, beliefs about teaching and learning in mathematics that were inconsistent with the reform initiatives, the nature of coaching, and gains in student achievement and engagement. They also experienced negative emotions because of favorable in-school factors such as an administration that supported teacher efforts to implement mathematics reforms. The study shows that: a) coaching may not help teachers reconstruct their professional self-understanding when it fails to address their self-image issues; b) teacher learning may occur even when the teacher’s beliefs are inconsistent with reform initiatives; and c) even when teacher learning results from coaching, reforms do not present themselves as expected in the classroom. Coaches experienced positive and negative emotions as a result of how well the reforms were being implemented by teachers. The experiences of the two coaches during mathematics reforms indicate a need to support coaches as they help teachers use the reform strategies. The directions for future research are described.

Keywords: Teacher Emotions
Mathematics
Advisor: McDougall, Douglas Emerson
Mapping Genetic Interaction Networks in Yeast

Global quantitative analysis of genetic interactions provides a powerful approach for deciphering the roles of genes and mapping functional relationships amongst path-ways. Using colony size as a proxy for fitness, I developed a method for measuring ge-netic interactions from high-density arrays of yeast double mutants generated by synthet-ic genetic array (SGA) technology. I identified several experimental sources of systematic variation and developed normalization strategies to obtain accurate fitness measurements. I used this scoring method to map quantitative genetic interactions among 5.4 million yeast double mutants and generated the first functionally unbiased genetic interaction map of a eukaryotic cell. My map produced an unprecedented view of the cell in which genes of similar biological processes cluster together in coherent subsets and functionally interconnected bioprocesses map next to each other. We discovered several physiological and evolutionary gene features that are characteristic of genetic interaction hubs, and explored the relationship between genetic and protein-protein interaction networks. In particular, by comparing quantitative single and double mutant phenotypes, we identified specific cases of positive genetic interactions, termed genetic suppression, and constructed a global network of suppression interactions among protein complexes. I also demonstrated that an extensive and unbiased mapping of genetic interactions provides a key for interpreting chemical-genetic interactions and identifying drug targets. In addition, I used genome-wide SGA data to map profiles of genetic linkage along all sixteen yeast chromosomes. These linkage profiles recapitulated previously identified recombination patterns and uncovered an unexpected correlation between chromosome length and the extent of centromere-related recombination repression. These findings suggest a chromosome size-dependent mechanism for ensuring proper chromosome segregation and highlight the SGA methodology as a unique approach for systematic analysis of yeast meiotic recombination.

Keywords: genetic interactions
networks
yeast
Advisor: Boone, Charles
Bader, Gary D.
Validation and Functional Characterization of Novel Neurofibromin Interacting Proteins

Neurofibromin (NF1) is a 2,818aa protein encoded by the very large NF1 tumour suppressor gene located on chromosome 17q11.2. Loss of function mutations and deletions in NF1 underlie Neurofibromatosis type-1 (NF-1) - the most common inherited syndrome of the nervous system in humans with a birth incidence of 1:3,000. The most visible feature of NF-1 is the neoplastic manifestations known as neurofibromas, however, the syndrome is also characterized by pigmentary defects, peripheral motor dysfunction, learning disabilities and several developmental abnormalities. The molecular etiology of many of these non-neoplastic phenotypes remains unknown. Here we demonstrate that the Tubulin Binding Domain (TBD) of NF1 is a binding partner of the Leucine Rich Pentatrico Peptide Repeat motif-Containing protein (LRPPRC) and cytoplasmic Dynein Heavy Chain (DHC). The NF1-LRPPRC interaction is of high significance as it links NF-1 with Leigh’s Syndrome, French Canadian variant (LSFC) – an autosomal recessive neurodegenerative disorder that arises due to mutations in the LRPPRC gene. This interaction occurs as part of an RNA granule complex, and use of transgenic mouse models establishes an important role of NF1 and LRPPRC in peripheral nerve development. The NF1-DHC interaction is of importance in melanocytes where our studies suggest a possible role in melanosome localization, disruptions in which may underlie the abnormal pigmentary features known as café-au-lait macules that are commonly associated with NF-1. The validation of LRPPRC and DHC as novel NF1 interactors reveal new roles of NF1, which open the door to better understanding the molecular mechanisms that underlie the myriad of NF-1 manifestations.

Keywords: Neurofibromatosis Type 1
Leigh's Syndrome, French Canadian variant
Neurofibromin
Leucine Rich Pentatrico Peptide Repeat motif-Containing protein
Advisor: Guha, Abhijit
McGlade, Jane
Kaplan, David
Identification of a Novel G-protein Interactor, RADIL, and Functional Characterization of its Role in Cancer Cell Motility

Cell adhesion and migration play crucial roles in development of multicellular organisms, immune surveillance, wound repair and cancer metastasis. The Gβγ subunits of heterotrimeric G-proteins have been implicated in signalling activities that promote cell adhesion and migration but the molecular mechanisms are unclear. Using a mass-spectrometry based proteomic approach we identified a protein complex between Gβγ and Rap1a that is bridged by a novel Rap1 effector, Radil. Overexpression of constitutively active Rap1a, Gβγ or stimulation of cells with the GPCR ligand fMLP triggers recruitment of Radil to the plasma membrane. Exogenous expression of Radil promotes cell spreading through Rap1-dependent inside-out activation of integrins leading to enhanced cell-matrix adhesion. Structure function experiments demonstrated that the RA and PDZ domains of Radil are required for its ability to promote cell adhesion. Using phage-display and mass-spectrometry we identified the kinesin family protein KIF14 as a novel interacting partner for Radil. Both KIF14 and Radil colocalized on microtubules in a PDZ-dependent manner. Depletion of KIF14 or disruption of microtubules led to accumulation of Radil at the cell membrane. Functionally, KIF14 is a negative regulator of Radil signalling as its depletion increased cell spreading and integrin activation and both phenotypes are rescued by simultaneous knockdown of Radil. Knockdown of KIF14 affects focal adhesion dynamics, which we determined is due to delayed adhesion disassembly. Depletion of either KIF14 or Radil dramatically decreased breast cancer cell migration and invasion in vitro. Additionally, knockdown of Radil compromised the ability of cells to metastasize to the lung and reduced tu-mor growth in xenograft mouse models. Collectively, these studies describe a functional re-quirement for the Gβγ-Rap1a-Radil complex during GPCR signalling for the control of integrin-mediated cell adhesion, cell motility and cancer progression.

Keywords: G protein
Cell motility
Radil
Cancer
Gbetagamma
Rap1
Advisor: Angers, Stephane
Phenotypic Classification of Paediatric Inflammatory Bowel Disease

This thesis explores aspects pertinent to the phenotypic classification of paediatric patients with inflammatory bowel disease (IBD). In the current era it has never been more important to have rigourous phenotypic classification to facilitate genotype-phenotype correlation studies as well as to optimize design of clinical trials of emerging therapies, where frequently response may differ according to phenotype of disease. The first study examined the reliability of the Montreal Classification for classifying paediatric IBD patients. This is the first study exploring the reliability of phenotypic classification in a paediatric population. The reliability of assigning an overall diagnosis of type of IBD was good, but not excellent. Amongst Crohn’s disease patients, reliability of assigning disease behaviour was excellent, while the reliability of assigning disease location categories varied from fair to good. The percentage agreement when describing disease extent for ulcerative colitis was high. The second study described the evolution of disease phenotype in a cohort of paediatric IBD patients. Similar to observations in adult-onset IBD, disease location was found to be relatively stable, while Crohn’s disease behaviour evolves from an inflammatory to a stricturing and/or penetrating phenotype in 20% of patients by 5 years of follow-up. The final study explored the association between 2 polymorphisms in the NOD2 gene with the requirement for intestinal resection (a surrogate marker of complicated disease) in models that included and excluded disease duration. Although no difference was found, this may have been influenced by data quality, which was suboptimal. In conclusion, this thesis has demonstrated that imprecision exists in the phenotyping of paediatric IBD patients, in whom phenotypic characteristics evolve over time. It is pertinent that disease duration be considered in any study attempting to make phenotypic correlations.

Keywords: Inflammatory bowel disease
paediatric
classification
phenotype
Advisor: Urbach, David Robert
School Travel Mode Choice Behaviour in Toronto, Canada

Interest in school transportation has emerged in response to concern over the reduced levels of physical activity among children and youth. Recent Canadian policies emphasize population health intervention to encourage active travel among this younger population; urban planners and public health professionals have also highlighted the importance of the neighbourhood built environment. However, this “child-youth friendly” turn in policy and planning practice, particularly in Canada, has arguably occurred in advance of conclusive and generalizable knowledge about the relationship between the built environment and children’s travel. Within this context, this thesis is the first quantitative research that examines school transportation mode choice behaviour in the largest Canadian city, the City of Toronto. At first, a Behavioural Model of School Transportation was outlined. This exercise was followed by three empirical studies that explored school travel by children and youth, using travel data from Transportation Tomorrow Survey. The first of these three studies investigated the association between the built environment and the likelihood of walking or being driven, for journeys to and from school. The next study focused on a measurement issue; the potential influence of the modifiable areal unit problem on statistical modelling of the built environment - mode choice relationship was examined. Lastly, the potential influence of travel interactions among household members, the built environment, and unobservable spatial dependency (i.e., spatial auto-correlation), on school travel outcome of children and youth, were examined. The results indicated that caregiver availability may influence travel mode choice. The built environment near both home and school locations was associated with the likelihood of walking. In addition, the correlates of mode choice were different between children and youth, which perhaps, reflects a child’s cognitive development with age as an independent traveller. These findings suggest the need for neighbourhood-wide improvement in the built environment, and age-specific population health interventions at schools and in the communities.

Keywords: Built environment
Active travel
Children
School transportation
Physical activity
Intra-household interaction
Advisor: Buliung, Ron
Hess, Paul M.