VictorJuiz/thesis huggingface dataset:
The Role of Suppressor of Fused During Murine Cerebellar Morphogenesis

Suppressor of Fused (SuFu) is an intracellular mediator of the Hedgehog signaling pathway with diverse biological roles, including a tumour suppressor function. Recent data suggest an essential role for this gene in cell differentiation in various tissues and cancers. Despite its known role as a tumour suppressor in medulloblastoma, a neuroectodermal tumour of cerebellar origin, the functional importance of SuFu in the developing cerebellum remains unknown. In this thesis, using in vivo conditional knockout approaches and in vitro assays, I present evidence for a novel requirement for SuFu during mid-hindbrain specification, cerebellar patterning and cell differentiation. Prior to the establishment of the cerebellar territory, SuFu regulates the formation of mid-hindbrain structures via GLI3 repressor (GLI3R), a Hedgehog pathway effector with a transcriptional repressor function. SuFu controls cerebellar morphogenesis by promoting timely cell differentiation of specific neuronal types. SuFu is also required for proper regulation of Sonic hedgehog signaling activity, which drives cell proliferation in granule cell precursors and is dysregulated in medulloblastomas. By using in vivo approaches to target gene deletion to specific cell lineages, I further demonstrate an essential role for SuFu in neuronal differentiation of granule cells and radial precursors, respectively. SuFu governs proper cell cycle exit, migration and terminal differentiation in granule cell precursors in a GLI3R-dependent manner. In cerebellar radial precursors, which give rise to cerebellar GABAergic neurons and glia, SuFu acts to promote neuronal differentiation in vivo and in vitro. This SuFu function is in part mediated by GLI3R, suggesting a novel functional contribution for this Hedgehog pathway effector in the developing cerebellum. These insights will serve as a basis for future investigation into the molecular mechanisms by which SuFu functions to control cerebellar neurogenesis, a process central to our understanding of the pathogenesis of medulloblastoma and other developmental disorders of the human cerebellum.

Keywords: Cerebellum
Suppressor of Fused
Sonic hedgehog
Differentiation
Advisor: Rosenblum, Norman
Probing the Role Played by Magnetic Fields in Star Formation with BLASTPol

BLASTPol, the Balloon-borne Large Aperture Submm Telescope for Polarimetry, measures linearly polarized Galactic dust emission in order to map magnetic field morphology in nearby molecular clouds. The role played by magnetic fields in providing support against gravitational collapse of clouds and cores and in forming observed cloud morphology is a key outstanding question in our understanding of star formation. BLASTPol has the unique combination of sensitivity to large scale magnetic fields, and sub-arcminute resolution necessary to trace fields into prestellar cores and dense filaments. Thus BLASTPol provides a crucial link between the low resolution Planck all sky maps and the detailed but narrow field-of-view capabilities of ALMA. The telescope operates simultaneously in three bands at 250, 350 and 500 microns which span the SED peak of 10-20K cold dust. This thesis gives an overview of the instrument, with special focus on the telescope motor control and scanning strategy. BLASTPol has had two science flights, both launched from Antarctica. The first flight began in December 2010 and lasted for 9.5 days. The 2010 flight had a number of problems, most significantly a damaged IR blocking filter which degraded the optical performance of the telescope. The second BLASTPol flight began in December 2012 and had much better instrument performance resulting in 12.5 days of science data. The performance of the instrument is reviewed, with the goal of providing insight for future BLASTPol flights and balloon telescopes. Reduction of the 2010 flight data was challenging because of the systematics associated with the damaged IR blocking filter. A review is given of the data reduction pipeline, instrumental polarization characterization, and a series of null consistency tests designed to reject polarization pseudo-vectors contaminated by systematics. This pipeline was tested with simulated timestreams which show that the BLASTPol instrument can be used to reconstruct polarization morphology. The 2012 data is of better quality and preliminary data reduction products have been run through the 2010 analysis pipeline. Though the resulting maps are preliminary they highlight the potential of these data to provide the most detailed maps to date of magnetic field morphology in molecular clouds.

Keywords: Star Formation
Balloon Astronomy
Magnetic Fields
Instrumentation
Submillimeter Astronomy
Interstellar Dust
Polarimetry
Advisor: Netterfield, Calvin Barth
Identification, Characterization and Redox Regulation of Carboxysomal Carbonic Anhydrases from Multiple Subtypes of the Cyanobacterial Bicarbonate Dehydration Complex

The photoautotrophic prokaryote, cyanobacteria possess a CO2 concentrating mechanism (CCM) necessary for growth. Carboxysomes are microcompartments essential for photosynthesis and the CCM in cyanobacteria. These organelles house Rubisco and carbonic anhydrase (CA) within a protein shell. In the cyanobacterium, Synechocystis PCC6803, CcaA is the functional CA and is complexed with two proteins, CcmM and CcmN, forming a HCO3- dehydration complex. CcmM acquires cytosolic HCO3- and delivers it to CcaA which catalyzes HCO3- dehydration within the carboxysome core. However, many beta-cyanobacterial strains such as Thermosynechococcus elongatus BP-1 lack a CcaA homolog. While the N-terminal region of CcmM is homologous to gamma CAs, this domain is inactive in Synechocystis. We identified and demonstrated that CcmM from T. elongatus, Nostoc PCC 7120, N.punctiforme, G.violaceus, Arthrospira and A.variabilis catalyzes 18O exchange in CO2, a hallmark of all CAs. Using truncated forms of CcmM, biochemical analysis and homology modeling, we defined the minimal gamma CA catalytic domain of CcmM to encompass the N-terminal 201 amino acids. CcmM201 is active, suggesting an important stabilizing role for a C194-C200 disulfide bond, despite these residues playing no direct role in the catalytic mechanism. Consistently, cysteine substituted mutants were less active than the wildtype. In addition, reducing agents inhibited the gamma CA activity of Nostoc PCC 7120 suggesting that the C194-C200 disulfide bond is critical for the oxidative activation (and potentially regulation) of the enzyme.

Keywords: Carbonic anhydrase
Carboxysome
Cyanobacteria
Carbon dioxide concentrating mechanism
Bicarbonate dehydration complex
Advisor: Espie, George S.
Positive and Negative Selection of Developing Antigen Specific B Cells in a Gut Associated Lymphoid Organ, the Bursa of Fabricius

Abstract Surface immunoglobulin (sIg) expression is a critical checkpoint in the early stages of B cell development. However it remains controversial as to whether sIg ligation plays a role in gut associated lymphoid tissues such as the bursa of Fabricius in chickens. To assess the role of antigen in B cell development, we constructed sIg related B cell receptors (BCRs) of defined specificity. Initial attempts to generate such BCRs by reverse engineering selected scFv expressing phage derived sequences into intact sIgM revealed a failure to maintain specificity due to loss of antigen binding or heavy chain:light chain pairing incompatibility in the full length immunoglobulin. In contrast, fusion of lamprey variable lymphocyte receptors (VLRs) of defined specificity, such as Phycoerythrin binding VLR, to the chicken sIgM constant region resulted in chimeric antigen specific BCRs that were expressed and supported the early stages of chicken B lymphopoiesis. While VLRPET&#61549;&#61472;expressing B cells specific for Phycoerythrin (PE) passed the sIg selection checkpoint and were detected in the bursa and the periphery, VLRHELT&#61549;&#61472;&#61506;&#61472;cells specific for the self-antigen Hen Egg Lysozyme (HEL) were not detected. Moreover, i.v. injection of PE into VLRPET&#61549; expressing chicks during embryogenesis induced deletion of splenic VLRPET&#61549; B cells. Furthermore, we demonstrated signaling dependent deletion of mCD8&#61537;:chIg&#61537; expressing B cells as a consequence of co-expressing TL, a natural ligand of mCD8&#61537;. While VLRPET&#61549; expressing B cells disappeared within the first 10 days after hatch in normal chicks, intrabursal application of PE sustained bursal and peripheral VLRPET&#61549; B cells. Migration of VLRPET&#61549; expressing B cells to the periphery was enhanced by intrabursal PE and included both short-lived and longer-lived peripheral B cells. This work represents the first evidence for positive and negative selection of B cells in avian gut associated B cell development. It demonstrates distinct B cell fates depending on the route of antigen administration and the requirement for BCR ligation to prolong the survival of bursal and peripheral B cells.

Keywords: B cell development
GALTB lmphopoiesis
Comparetive immunology
Avian B lymphopoiesis
B cell Positive and Negative selection
BCR
BCR fusion
B cell receptor ligation
Bursa of Fabricious
VLR
Antigen
Advisor: Ratcliffe, Michael
Engendering Security: Lessons from Post-conflict Central America

Analyzing post-conflict contexts from a feminist perspective sheds light on an important, yet overlooked, fact in peacebuilding research: While human rights are officially recognized and codified by state and international institutions, the actual possibility of enjoying these rights often depends on whether they are respected or violated in private, often intimate, gendered relations. I therefore ask: How might peacebuilding efforts bridge the gap between the promise and practice of women’s rights and basic security in post-conflict contexts? In response, I argue that peacebuilding efforts must foster a feminist vision of positive peace that requires not only the absence of war, but also the elimination of unjust social relations. So long as gendered violence is largely excluded from mainstream peacebuilding theories and practices, a robust human rights regime will remain elusive - even in contexts that are otherwise regarded as peacebuilding "success stories". In theory, the protection and promotion of human rights as part of peacebuilding is not very controversial. The challenge, as I illustrate, is in finding ways to make these rights a meaningful reality (i.e. a behavioural practice) in the lives of a post-conflict population. Based on 92 interviews and extensive participant-observation research in El Salvador, Guatemala, and Nicaragua, I demonstrate how grassroots projects are well-positioned to: a) address the private relations in which gendered violence often occurs, and b) change the attitudes, beliefs and behaviours that sustain the gendered power relations and gendered roles subordinating women. I focus on three projects developed by Central American citizens to strengthen women’s rights and basic security: Women’s Police Stations, Nuevos Horizontes (New Horizons) a shelter for survivors of intimate partner violence, and the innovative telenovela or "social soap" Sexto Sentido (Sixth Sense). My findings show how these projects both empower women through a combination of resources, agency and achievements, and challenge traditional gendered roles and identities common in the region, such as machismo, that normalize violence against women. By challenging patriarchal power distributions and gendered identities/roles from the "bottom-up," my research offers baseline examples of how grassroots initiatives can improve women's security, consequently strengthening the rights-based culture necessary for positive peace.

Keywords: gender
peacebuilding
rights
Central America
Advisor: Nedelsky, Jennifer
The Association between Cardiac Energy Metabolism and Physical Activity during Adolescence and Early Adulthood

A greater understanding of the mechanistic associations among subclinical cardiac dysfunction, metabolism, and physical activity may facilitate the early identification and treatment of premature cardiovascular disease originating during late adolescence, thereby preventing the progression of cardiovascular disease in adulthood. The present set of studies involved the implementation of novel cardiac 31Phosphorus Magnetic Resonance Spectroscopy in adolescents and young adults at The Hospital for Sick Children. These pilot studies are the first to characterize cardiac energy metabolism in both healthy and overweight cohorts during adolescence and early adulthood, as well as the first to explore the relationships among physical activity, cardiac and skeletal muscle energy metabolism, and cardiac function. Notably, reproducibility of primary study measures, including cardiac (i.e., PCr/ATP-beta) and skeletal muscle energy metabolism (i.e., PCr recovery half-time), was explored (Study A). A positive association was demonstrated between cardiac energy metabolism and skeletal muscle energy metabolism, independent of moderate-to-vigorous physical activity levels and peak exercise performance (Study B). A positive association was also observed between novel measures of cardiac energy metabolism (resting cardiac PCr/ATP-beta) and cardiac function (i.e., resting LV torsion and longitudinal strain during a graded exercise protocol) in active healthy weight and overweight adolescents, independent of body mass index percentile and moderate-to-vigorous physical activity levels (Study C). A crossover study intervention, including nine consecutive days of remote ischemic preconditioning (RIPC) and high-intensity interval exercise, was not associated with improvements in peak exercise performance or cardiac energy metabolism (Study D). Larger, randomized clinical research studies ultimately need to be conducted to substantiate these study findings, as well as to investigate the longer-term effect of chronic RIPC and exercise on energy metabolism and cardiovascular health in both sedentary and clinical cardiology cohorts.

Keywords: exercise
magnetic resonance spectroscopy
Advisor: McCrindle, Brian
Development of Cell Culture Processes on Digital Microfluidic Platforms

In vitro microenvironments used for culturing and studying living cells have remained virtually unchanged for the last five decades. Mammalian cells are routinely seeded as monocultures onto rigid, homogeneous, two dimensional substrates &ndash; systems with limited physiological relevance. Microfluidics has the potential to significantly improve cell models to better mimic native tissue or disease states in addition to a host of other benefits such as improved throughput, reduced consumable requirements and seamless integration with a number of analysis techniques. Digital microfluidics, a fluid handling technique which manipulates discrete droplets over micro-electrode patterned surfaces, may be a valuable tool for a number of cell applications incorporating all of the above potential advantages. Cell culture and analysis is a new application for digital microfluidics &ndash; the first report of such was in 2008. As a result, a number of technical impediments must be addressed before cells can be effectively and routinely studied on these microfluidic devices. These impediments include: a) rapid device failure due to protein biofouling on hydrophobic device surfaces, b) the unexplored possibility of detrimental effects on cell fitness arising from the electrokinetic manipulation of droplets and c) the lack of robust iii systems capable of long term automated culture and integrated analysis of cells. The aim of the work described in this thesis is to advance digital microfluidics for biologically relevant cellular culture and analysis by addressing each of these challenges. The technical advancements gained from these studies can then be used to develop a proof-of-concept digital microfluidic platform for modeling liver tissue. In summary, this work describes advances towards physiologically-relevant culture and analysis of living cells on digital microfluidics, a technology which has the potential to become a valuable tool for the biomedical research community.

Keywords: microfluidics
cell culture
digital microfluidics
biomedical
Advisor: Wheeler, Aaron
A Single-Channel Acoustic Method for Portable Diagnosis of Sleep Apnea

Background: Sleep apnea is a very common disease with serious health consequences. It affects approximately 85% of adults, most of whom remain undiagnosed, which puts them at risk of car accidents, because of sleepiness, and hypertension, heart failure, and stroke. The objective of this thesis was to develop a single-channel device for portable monitoring of sleep apnea that uses breath sounds collected via a microphone placed in front of the nose and mouth. Methods: Project 1: Frequency characterization was used to identify the basic components of the respiratory cycle, i.e., inspiration and expiration. Project 2: Inspiratory sounds were used to determine upper airway narrowing by means of Linear Predictive Coding (LPC), which was validated against objective measures of upper airway resistance. Project 3: An algorithm to calculate the frequency of apneas and hypopneas per hour (apnea-hypopnea index or AHI) was developed and validated against polysomnography in 50 subjects during sleep. Project 4: Finally, a self contained device was then developed and evaluated in 49 subjects in the home setting, 11 of whom used the device on 2 different nights. Results: Project 1: Inspiratory and expiratory phases had characteristically different spectra that allowed them to be distinguished with up to 97% correct classification. Project 2: LPC coefficients were found to be modulated by the ensuing of upper airway narrowing. Project 3: AHI determined by acoustic analysis showed up to 94% correlation with PSG and up to 90% diagnostics accuracy. Project 4: In the home unattended setting, the overall rating for ease-of-use was excellent and the success rate of independent use was 94%. The portable device showed excellent performance with very high signal-to-noise ratio of 31.7 dB. The intra-subject 2-night AHI scores were reproducible in 9 out of 11 (82%) home subjects, which is well within reported inter-night variability. Conclusion: Acoustic analysis of breath sounds is a powerful tool for characterization of respiratory patterns including upper airway narrowing and accurate identification of apneas and hypopneas as compared with the current gold standard. This technology can be packaged in a compact device that can be used independently and reliably in the home environment.

Keywords: Sleep Apnea
Portable Diagnosis
Acoustics
Medical Device
Signal Processing
Algorithm
Upper Airway
Breathing Phases
Usability
Embedded Module
Advisor: Fernie, Geoff R
Bradley, T Douglas
The Role of H-NS in Bacterial Evolution and Factors that Modulate its Silencing Activity

Horizontal gene transfer (HGT) is the primary source of genetic variability among free-living bacteria. Genes acquired by HGT typically display a lower AT content than the host genome average, but the reason for this bias is unknown. H-NS is a pleiotropic regulatory protein that is thought to facilitate HGT by silencing the expression of foreign genes with a lower AT-content than the host genome. In Salmonella enterica serovar Typhimurium H-NS represses 12% of the genome and loss of H-NS function imposes severe growth penalties on hns mutant cells. Here I applied an experimental evolution approach to examine the underlying basis for the hns mutant fitness defects. Genetic adaptions that improved hns mutant growth included deletion of the horizontally acquired Salmonella pathogenicity island 1 (SPI-1) and missense mutations that enhanced the gene silencing properties of the H-NS paralogue protein StpA. My findings support a role for H-NS in maintaining genomic integrity and suggest that the determinants responsible for the functional distinctions between StpA and H-NS reside in their self-association domains. The repression of many foreign sequences requires a second regulatory molecule called Hha. Hha directly interacts with the N-terminal dimerization domain of H-NS and augments silencing at a subset of the H-NS regulon. To explore this further we solved the first x-ray crystal structure of Hha in complex with the N-terminal dimerization domain of H-NS. Our complete structural/functional analysis of the Hha/H-NS complex supports a model whereby Hha enhances the silencing properties of H-NS by remodeling the nucleoprotein filament structure. In addition to Hha, H-NS also interacts with a protein encoded by the T7 bacteriophage called 5.5. The 5.5 protein was previously shown to activate transcription at model H-NS repressed E. coli and T7 phage promoters by an unknown mechanism. I demonstrate that the 5.5 protein antagonizes the oligomerization properties of H-NS by binding to its central dimerization domain. My work on the 5.5 protein further highlights the importance of higher order nucleoprotein complexes for H-NS mediated repression of foreign DNA. Overall, the studies presented in this dissertation establish the macromolecular structure of H-NS/DNA complexes as a key determinant of transcriptional silencing activity and shed light on mechanisms that modify these nucleoprotein structures.

Keywords: horizontal gene transfer
transcriptional silencing
Salmonella
DNA binding proteins
Advisor: Navarre, William
Multiliteracies Theory Into Practice: An Inquiry Into Junior Level Literacy Classrooms

The purpose of this research was to investigate the extent to which junior level literacy teachers enacted a pedagogy of multiliteracies in their classroom program. This qualitative study employed semi-structured interviews to investigate participants' perceptions and implementation of multiliteracies theory. The researcher also assumed the role of a participant using self-study research methodology, recording personal reflections from her experiences as a Grade 4 literacy teacher over a one-year period in addition to responding to the interview questions. Findings emerged in five areas. First, the participants demonstrated broad conceptualizations of literacy that aligned with the principles and components of multiliteracies theory. Secondly, the data revealed the use of cross-curricular integration, allowing these teachers to provide literacy instruction across the various subject areas. Third, there was a strong presence of digital literacies, highlighting both positive outcomes of technology use and many obstacles that the participants worked to overcome. The fourth theme presents transformed practice activities that occurred through in-class activities and small group initiatives running parallel to participants' literacy programs. Finally, participants' current use of supports versus desired supports for their literacy programs are discussed. Based on the findings, implications and recommendations are provided for supporting teachers, teacher education programs, and future research.

Keywords: Adolescent
Classroom
Literacy
Multiliteracies
Qualitative
Technology
Advisor: Kosnik, Clare