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Mitochondrial Metabolic Suppression And Reactive Oxygen Species Production During Hypometabolism In Mammals, Jason Cl Brown

Electronic Thesis and Dissertation Repository

During hibernation, daily torpor, and fasting, mammals reduce metabolic rate (MR) up to 99%, 95%, and 30%, respectively, compared to resting levels. Mitochondrial metabolic suppression likely contributes to this MR reduction, and the first objective of this study was to determine the relative contributions of active, regulated inhibition and passive thermal effects as body temperature (T_b) falls, to mitochondrial metabolic suppression, and to examine the mechanisms involved using top-down elasticity analysis and novel statistical approach. The second objective of this study was to determine how mitochondrial metabolic suppression affects mitochondrial reactive oxygen species (ROS) production, a topic which …

Regulation Of G Protein Signaling By Goloco Motif Containing Proteins, Peishen Zhao

Electronic Thesis and Dissertation Repository

Signal transduction via heterotrimeric G proteins in response to transmembrane G protein-coupled receptors plays a central aspect in how cells integrate extracellular stimuli and produce biological responses. In addition to receptor-mediated activation of heterotrimeric G proteins, during the last few decades, accessory proteins have been found to regulate G protein activity via different mechanisms. Several proteins have been identified that contain multiple G protein regulatory domains. Using various molecular and biochemical approaches, we have characterized the effects of two such proteins, G18 and RGS14, on G protein activity. Both proteins contain a second G protein binding domain in addition to …

Metastatic Disease: Interactions Between Tumor Cells And Host Environment During Cancer Cell Spread, Jennifer M. Maclean

Electronic Thesis and Dissertation Repository

Tumor and metastasis formation are not cell autonomous phenomena, but rather an evolution of disease within and responding to the host environment. Metastatic spread from a primary tumor occurs as a result of a complex interplay between tumor cells and the host, wherein tumor cells must escape the primary tumor, enter the host vasculature, travel to and arrest in a distant tissue and survive and grow in that new organ. It is known that cells that progress through these stages must both escape and exploit host systems, yet the mechanisms used are not fully understood. Therefore, the goal of this …

The Regulation Of Cell Division And Neurogenesis By The Chromatin Remodeling Protein Atrx, Kieran L. Ritchie

Electronic Thesis and Dissertation Repository

Mutations in the ATRX gene cause alpha thalassemia mental retardation X-linked syndrome (ATR-X). Individuals with ATR-X syndrome show a broad range of developmental abnormalities including severe cognitive, behavioural, and learning deficits, seizures, and microcephaly, indicating that ATRX is essential for normal development of the central nervous system. Conditional inactivation of Atrx in the developing mouse forebrain leads to a reduction in cerebral cortical size, elevated levels of p53-dependent neuronal apoptosis, and dysgenesis of certain subcortical structures, including the hippocampus and dentate gyrus, confirming a requirement for Atrx in mammalian brain development. The mammalian ATRX gene encodes a member of the …

Role Of Rac1 In Myocardial Tnf-Alpha Expression In Sepsis, Ting Zhang

Electronic Thesis and Dissertation Repository

Tumor necrosis factor-a (TNF-alpha) is a pro-inflammatory cytokine and its high levels of expression in the heart leads to cardiac dysfunction in sepsis. However, the underlying molecular mechanisms of regulating myocardial TNF-alpha expression are not fully understood. The aim of this thesis was to investigate the role of Rac1 in myocardial TNF-alpha expression and cardiac dysfunction during sepsis. Studies were performed using cultured neonatal cardiomyocytes and a mouse model of endotoxemia.

PI3K-mediated Rac1 activation is required for induction of TNF-alpha mRNA and protein expression in cardiomyocytes and cardiac dysfunction during endotoxemia. Rac1 promotes TNF-alpha mRNA expression via NADPH oxidase/ERK1/2 and …

Dissecting The Molecular Role Of Distinct Binding Interfaces On The Retinoblastoma Tumor Suppressor In Growth Control And Tumorigenesis., Matthew J. Cecchini

Electronic Thesis and Dissertation Repository

The retinoblastoma tumor suppressor protein (pRB) functions to maintain proliferative control and act as a barrier to tumorigenesis. pRB is capable of regulating E2F transcription factors to mediate control of proliferation through transcriptional regulation of S-phase target gene expression. In addition, pRB can stabilize the CDK inhibitor p27 through an interaction with two ubiquitin ligase complexes. Further, pRB is capable of forming a unique interaction with E2F1 termed the ‘specific’ interaction that is capable of blocking E2F1 induced apoptosis. These functions of pRB are mediated by distinct binding interfaces and their contributions to the overall functionality of pRB are not …

The Effects Of Superovulation And Embryo Culture On Genomic Imprinting In A Mouse Model System, Brenna A. M. Velker

Electronic Thesis and Dissertation Repository

Genomic imprinting is a specialized transcriptional mechanism resulting in the unequal expression of alleles based on their parent-of-origin. Imprinted genes are critical for embryonic and fetal development and their dysregulation is linked to a group of human diseases called imprinting disorders, including Beckwith-Wiedemann Syndrome, Angelman Syndrome and Silver-Russell Syndrome. Two critical phases of genomic imprinting exist. The acquisition phase occurs in developing germ cells, asynchronously for different imprinted loci, while the maintenance phase takes place during preimplantation development, while the rest of the genome is undergoing demethylation. Increased frequencies of human imprinting disorders are observed in children following the use …

Extent, Characteristics And Downstream Effects Of Stream Enclosure In Southwestern Ontario, Katie L. Stammler

Electronic Thesis and Dissertation Repository

In a 16000 km² area of southwestern Ontario, almost 15% of all streams have been enclosed (buried) largely for agricultural purposes. ArcGIS was used to characterize the natural features of catchments and to calculate enclosedness (proportion of stream network enclosed; x̅ = 16.5%, n = 10106). Catchments with the highest enclosedness received >990 mm of precipitation annually and had high drainage density (>1.9 km/km²), while catchments with the lowest enclosedness receivedannually, were characterized by clay and undrumlinized till plains and had relatively shallow water tables (m). These natural features influence where enclosure is likely to occur, …

Regional Specification Of The Xenopus Lateral Plate Mesoderm, Steven J. Deimling

Electronic Thesis and Dissertation Repository

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Successful patterning of the embryo, from establishing the three primary axes to the regional specification of tissue progenitors is essential to generating a viable embryo. The three germ layers in the early embryo undergo patterning through slightly different mechanisms. The tissue of interest to this study is the lateral plate mesoderm (LPM), which will give rise to the lineages of the cardiovascular system and is essential for regional specification of adjacent germ layers. …

Transactivation By Human Adenovirus Early Region 1a-Conserved Region Three, Jailal Ng Ablack

Electronic Thesis and Dissertation Repository

One of the critical functions of human adenovirus (hAd) early region 1A (E1A) protein is to activate transcription of the early viral genes. The largest isoform of E1A contains a unique region termed conserved region 3 (CR3), which includes a Cysteine-4 (C4) zinc finger domain. This region activates viral gene expression by interacting with and recruiting cellular transcription machinery to the regulatory regions of early viral genes. Although this process has been studied at length with hAd type 5 E1A, far less is known about how the E1A proteins from other hAd types activate transcription. There are dramatic differences in …

Serum Iron Uptake And Virulence In Staphylococcus Aureus, Federico C. Beasley

Electronic Thesis and Dissertation Repository

The high affinity iron scavenging glycoprotein transferrin sequesters trace amounts of serum Fe³⁺ to concentrations below what is required to sustain microbial life. Iron may be liberated from this important innate immune factor after interaction with molecules that chelate or reduce Fe³⁺. Organisms with cognate transport systems for these iron coordinating molecules can survive in the bloodstream using transferrin iron.

Staphylococcus aureus is an opportunistic bacterial pathogen. S. aureus executes numerous strategies for overcoming the innate immune barrier of iron deprivation in the bloodstream. In addition to specialized mechanisms for hemoglobin iron extraction, S. aureus can proliferate …

On The Early Onset Of Vascular Stiffening And Sexual Dimorphism Of Sympathetic Control In The Spontaneously Hypertensive Rat, Louis Mattar

Electronic Thesis and Dissertation Repository

The purpose of this thesis was to explore the role of the sympathetic nervous system (SNS) in hindlimb vasomotor control during the development of hypertension (HT). Using an animal model of essential HT (the spontaneously hypertensive rat [SHR]) we demonstrated that neuropeptide Y (NPY) and the Y₁ receptor (Y₁R) play a greater role in modulating hindlimb hemodynamics in the early stages of HT compared to normotensive controls (Wistar Kyoto [WKY]). Hindlimb vascular mechanics (compliance [C] and viscoelasticity [K]) were assessed using a modified Windkessel model developed in our laboratory. The hindlimb mechanics did …

Dissociable And Dynamic Components Of Cognitive Control: A Developmental Electrophysiological Investigation, Matthew Waxer

Electronic Thesis and Dissertation Repository

One standard task used to investigate the development of cognitive control is the Dimensional Change Card Sort (DCCS). Performance and patterns of brain activity associated with the DCCS show continued age-related advances into early adolescence. According to many theoretical accounts, the DCCS places demands on a single underlying executive control process. Three experiments examined the possibility that the DCCS places demands on multiple control processes that follow distinct developmental trajectories. In Experiment 1, rule switching and conflict processing made orthogonal contributions to DCCS performance. Rule switching was associated with a cue-locked late frontal negativity (LFN) event-related potential (ERP) and conflict …

Development Of New Platforms For Efficient Production And Purification Of Recombinant Protein In Higher Plants, Reynald Tremblay

Electronic Thesis and Dissertation Repository

The world-wide demand for recombinant proteins continuously increases as new medical and industrial applications are developed. Higher plants have the potential to help meet this rising demand as green bioreactors. A major hurdle, however, is low recombinant protein yields in higher plants and, as with many production systems, high cost associated with downstream purification and the production of short peptides. The goal of this research was to address each of these problems to further increase the utility of plant bioreactors. The short antigenic peptide 277 (p277) from heat shock protein 60, which has the potential for use in the prevention …

The Role Of Granulocyte Colony-Stimulating Factor In The Intestinal Immune System And The Response To Probiotics, Andrew J. Martins

Electronic Thesis and Dissertation Repository

Loss of immunoregulation in the intestine results in inflammation, such as is observed in the chronic, relapsing, inflammatory bowel diseases (IBD), Crohn's disease and ulcerative colitis. However, our understanding of this process remains incomplete. The cytokine granulocyte-colony stimulating factor (G-CSF) is an important stimulator of granulopoiesis in steady-state and during inflammation, but has also been shown to have anti-inflammatory effects and is a potential therapeutic for IBD. In addition, certain strains of bacteria, termed probiotics, have been postulated to have anti-inflammatory effects in the intestine. The mechanisms by which these strains of bacteria elicit anti-inflammatory effects are not fully understood. …

The Hiv-1 Tat Protein And Adverse Drug Reactions: A Model System Utilizing Jurkat T Cells And Sulphamethoxazole-Hydroxylamine, Kaothara Adeyanju

Electronic Thesis and Dissertation Repository

In 2009 approximately 2.6 million people became infected with the Human Immunodeficiency Virus (HIV). In addition to the estimated 33.3 million people currently living with the virus, this makes HIV/ AIDS an epidemic of unprecedented scale in modern times. Treatment of HIV infection requires antiretroviral agents as well as a number of other drugs such as antimicrobials. Hypersensitivity adverse drug reactions (ADRs) to a variety of drugs are common in HIV-infected individuals, but the antimicrobial Sulphamethoxazole remains a major culprit. Hypersensitivity ADRs cause significant morbidity, with the skin and liver most commonly affected and are among the top causes of …

The Life Cycle Of Pannexins: Trafficking, Cell Surface Dynamics, Turnover And Degradation, Ruchi Gehi

Electronic Thesis and Dissertation Repository

Pannexins (Panxs) are a recently identified family of channel-forming glycoproteins, comprised of three members: Panx1, Panx2 and Panx3. Panxs were classified as gap junction (GJ) proteins based on their shared sequence homology to the invertebrate GJ protein, innexins. While no sequence homology exist between Panxs and the GJ proteins, connexins (Cxs), they share similar topology with four transmembrane domains, two extracellular loops, an intracellular loop and cytoplasmic exposed amino and carboxy terminal tails. In this study, we investigated if Panx1 and Panx3 exhibit unique or shared properties of cell surface delivery, mobility, cytoskeletal dependency, internalization and degradation, to the well …

The Role Of The Mcm2 Subunit In Regulating The Activities Of The Mcm2-7 Helicase, Brent E. Stead

Electronic Thesis and Dissertation Repository

The transmission of genetic information from parental to daughter cells requires the faithful duplication of an organism’s genome. Uncontrolled DNA replication can result in proliferative diseases, such as cancer. DNA replication requires a single-stranded DNA template to be produced from duplex DNA. In eukaryotes, DNA unwinding for replication is performed by the heterohexameric replicative helicase complex comprised of the minichromosome maintenance proteins 2 through 7.

Each of the Mcm2-7 subunits likely has a unique role in DNA binding and unwinding by the Mcm2-7 complex. The present study examines the role of the Saccharomyces cerevisiae Mcm2 subunit in regulating the activities …

Identification Of Regions Responsible For The Open Conformation Of S100a10 Using Chimaeric S100a11/S100a10 Proteins, Liliana Santamaria-Kisiel

Electronic Thesis and Dissertation Repository

S100A11 is a dimeric, EF-hand calcium-binding protein. Calcium binding to S100A11 results in a large conformational change that uncovers a broad hydrophobic surface used to interact with phospholipid-binding proteins (annexins A1 and A2), and facilitate membrane vesiculation events. In contrast to other S100 proteins, S100A10 is unable to bind calcium due to deletion and substitution of calcium-ligating residues. Despite this, calcium-free S100A10 assumes an “open” conformation that is very similar to S100A11 in its calcium-bound state (Ca2+-S100A11). To understand how S100A10 is able to adopt an open conformation in the absence of calcium, seven chimeric proteins were constructed where regions …

Perception Meets Action: Fmri And Behavioural Investigations Of Human Tool Use, Kenneth F. Valyear

Electronic Thesis and Dissertation Repository

Tool use is essential and culturally universal to human life, common to hunter-gatherer and modern advanced societies alike. Although the neuroscience of simpler visuomotor behaviors like reaching and grasping have been studied extensively, relatively little is known about the brain mechanisms underlying learned tool use.

With learned tool use, stored knowledge of object function and use supervene requirements for action programming based on physical object properties. Contemporary models of tool use based primarily on evidence from the study of brain damaged individuals implicate a set of specialized brain areas underlying the planning and control of learned actions with objects, distinct …

Effects Of Acute Synchronous Whole-Body Vibration Exercise, Tom J. Hazell

Electronic Thesis and Dissertation Repository

Recently, whole-body vibration (WBV) exercise has been gaining interest in the health/fitness community for its reported beneficial outcomes. However, most of these have not been evaluated scientifically leading to some reservation in promoting this new exercise modality. The purpose of this dissertation was to assess the viability of WBV exercise to enhance several selected indices of health.

Study 1 demonstrated that the addition of WBV to an isometric semi-squat in young healthy men (n = 8, 25±2.6 y, 177±7.0 cm, 84±12.1 kg) resulted in increases in femoral artery blood flow and leg skin temperature vs the same exercise without vibration …

Structural Insights Into Dna Replication And Lesion Bypass By Y Family Dna Polymerases, Kevin N. Kirouac

Electronic Thesis and Dissertation Repository

Y family DNA polymerases are specialized enzymes for replication through sites of DNA damage in the genome. Although the DNA damage bypass activity of these enzymes is important for genome maintenance and integrity, it is also responsible for DNA mutagenesis due to the error-prone nature of the Y family. Understanding how these enzymes select incoming nucleotides during DNA replication will give insight into their role in cancer formation, aging, and evolution. This work attempts to mechanistically explain, primarily through X-ray crystallography and enzymatic activity assays, how Y family polymerases select incoming nucleotides in various DNA replication contexts. Initially, we sought …

Regulation Of Akt And Wnt Signalling By The Dopamine D2 Receptor And Metabotropic Glutamate Receptor 2/3, Laurie P. Sutton

Electronic Thesis and Dissertation Repository

Akt and the Wnt pathway, two cascades that regulate GSK-3, have been implicated in schizophrenia and antipsychotic drug action. Although it is known that antipsychotic drugs alleviate psychosis by blocking the dopamine D2 receptor (D2DR) and that metabotropic glutamate receptor 2/3 (mGluR2/3) agonists may improve some of the symptoms of schizophrenia, it is unclear if both classes of drugs exert their effects through Akt, GSK-3 and/or the Wnt pathway or if changes in these pathways are mediated through the D2DR and mGluR2/3 respectively. In addition to antipsychotics, mood stabilizers and antidepressants also target GSK-3, suggesting that there must be something …

Cardiac Adaptation To Chronic Blockade Of Voltage-Gated, L-Type Calcium Channels In The Sarcolemma, Ji Zhou

Electronic Thesis and Dissertation Repository

L-type Ca²⁺ channels (dihydropyridine receptors, DHPRs) in the sarcolemma are essential to cardiac excitation-contraction (E-C) coupling. Thus, Ca²⁺ influx through DHPRs upon cardiomyocyte excitation triggers Ca²⁺ release from the sarcoplasmic reticulum (SR) through ryanodine receptors (RyRs) to initiate myofilament activation and muscle contraction. Muscle relaxation occurs upon sequestration of Ca²⁺ back into the SR lumen by sarco/endoplasmic reticulum calcium-ATPase (SERCA) in the SR. As a treatment option for hypertension, long-term use of DHPR blockers is associated with increased risk of heart failure; underlying mechanisms are unknown. This research used male Wistar rats treated with verapamil (subcutaneously, …

Mental Blocks: The Behavioural Effects And Neural Encoding Of Obstacles When Reaching And Grasping, Craig S. Chapman

Electronic Thesis and Dissertation Repository

The ability to adeptly interact with a cluttered and dynamic world requires that the brain simultaneously encode multiple objects. Theoretical frameworks of selective visuomotor attention provide evidence for parallel encoding (Baldauf & Deubel, 2010; Cisek & Kalaska, 2010; Duncan, 2006) where concurrent object processing results in neural competition. Since the end goal of object representation is usually action, these frameworks argue that the competitive activity is best characterized as the development of visuomotor biases. While some behavioural and neural evidence has been accumulated in favour of this explanation, one of the most striking, yet deceptively common, demonstrations of this capacity …

Pharmacokinetics And Therapeutic Uses Of Mesna, Murray J. Cutler

Electronic Thesis and Dissertation Repository

In the early 1980s, significant advancement in the safety of ifosfamide therapy was achieved by co-administrating mesna (sodium 2-mercaptoethane sulfonate) to prevent dose-limiting hemorrhagic cystitis. Mesna exerts its protective effect within the urine, where its free sulfhydryl group is able to conjugate cytotoxic metabolites. Within the circulation, however, mesna exists primarily as its inactive disulfide, dimesna. Dimesna is currently undergoing clinical development as a prodrug (BNP7787) to treat cisplatin-induced nephrotoxicity. Remarkably, chemoprotection is achieved without attenuation of efficacy of co-administered anti-cancer agents. This is widely attributed to the kidney-specific disposition and stability of dimesna.

We sought to evaluate the role …

Stopover Biology Of Migratory Landbirds In A Heavily Urbanized Landscape, The New York Metropolitan Area, Chad L. Seewagen

Electronic Thesis and Dissertation Repository

Migration routes of many Nearctic-Neotropical landbirds pass through the most urbanized regions of North America. Migrants use urban habitat fragments as stopover sites and commonly occur in cities at exceptional density. Yet, knowledge of migrant stopover biology and refueling opportunities in such places is severely limited. This dissertation examined several aspects of migrant stopover biology in the New York metropolitan area to gain a more holistic understanding of how migratory landbirds utilize urban stopover sites, and ultimately to assess the quality of urban habitats as stopover sites. I first generated morphometric predictive models using salvaged bird specimens to allow me …

The Lotus Japonicus Cytokinin Receptor Gene Family And Its Role In Nitrogen-Fixing Symbiosis, Mark A. Held

Electronic Thesis and Dissertation Repository

Nitrogen is the most abundant element in our atmosphere, yet has become increasingly limited in agricultural lands. Legume plants offer a possible solution to this problem due to their innate ability to symbiotically interact with nitrogen-fixing bacteria called rhizobia. In particular, a histidine kinase cytokinin receptor from the model legume Lotus japonicus (LHK1) has been clearly placed at the core of these interactions. Loss-of-function mutants in LHK1 fail to initiate timely cortical cell divisions in response to abundant bacterial infection, and gain-of-function mutations in the same locus cause L. japonicus plants to form spontaneous nodules in the absence of rhizobia, …

Mitotic Regulation Of Protein Kinase Ck2, Nicole A. St. Denis

Electronic Thesis and Dissertation Repository

Protein kinase CK2 is a serine/threonine kinase with a multitude of substrates and roles in many cellular processes, including mitosis. CK2 is constitutively active, yet we hypothesize that CK2 is indeed regulated in mitosis through subtle means, enabling CK2 to perform its functions unique to cell division. Our aims were to examine the roles of mitotic phosphorylation, subcellular localization, and interplay with mitotic kinases in the regulation of CK2 activity.

We first examined the role of four highly conserved mitotic phosphorylation sites located in the unique C-terminus of CK2α. Phosphospecific antibodies generated against the sites show that CK2α phosphorylation is …

The Roles Of Nitric Oxide Synthases (Nos) In Endochondral Bone Formation, Qian Yan

Electronic Thesis and Dissertation Repository

Longitudinal growth of endochondral bones is controlled by the cartilage growth plate. Chondrocyte proliferation and hypertrophy, vascular invasion, formation of ossification centers and cartilage replacement by bone tissue are all important processes required for normal growth. These biological processes have to be tightly regulated or disturbances will lead to skeletal diseases. A large number of genes, growth factors and hormones have been implicated in the regulation of growth plate biology, however, less is known about the intracellular signaling pathways involved. Nitric oxide (NO) has been identified as a regulator of cellular proliferation, differentiation, migration, survival and metabolism in multiple cell …