Life Sciences Commons^™

Discovery And Characterization Of A Novel Class Of Metabolic Regulators In The Malaria Parasite Plasmodium Falciparum, Ann Marie Guggisberg

Arts & Sciences Electronic Theses and Dissertations

The malaria parasite, Plasmodium falciparum, infects hundreds of millions of people per year and causes hundreds of thousands of deaths. Within the host red blood cell, the parasite relies on glycolysis for energy and synthesis of essential biomolecules. One such anabolic fate of glucose is the synthesis of isoprenoids, a broad and essential class of compounds that participate in a variety of cellular functions. In the face of ever-evolving drug resistance, new inhibitors and better understanding of parasite metabolism are required. The antibiotic fosmidomycin (FSM) targets the methylerythritol phosphate pathway for isoprenoid synthesis and is a well-validated inhibitor of P. …

Genetic Mechanisms For The Maintenance Of Behavioral Mating Barriers In Drosophila, Kathleen M. Mortland

Arts & Sciences Electronic Theses and Dissertations

One of the most successful and diverse systems involved in the maintenance of behavioral barriers between closely related animal species is pheromonal communication. In the fruit fly, contact chemosensation input is especially important during sexual decision-making as it allows for the sensing of sex and species-specific non-volatile cuticular hydrocarbons (CHCs), which function as insect pheromones. However, how pheromonal systems support the maintenance of mating barriers is puzzling since any change in either pheromone ligands or their cognate receptors would carry a fitness cost, which should be eliminated by stabilizing selection. To resolve this evolutionary conundrum I hypothesized that pleiotropic genes …

Epigenomics Of Cell Fate In Development And Disease, Rebecca Faith Lowdon

Arts & Sciences Electronic Theses and Dissertations

Epigenetic features at regulatory elements provide instructive cues for transcriptional regulation during development. However, the particular epigenetic alterations necessary for proper cell fate acquisition and differentiation are not well understood. This dissertation explores the epigenetic dynamics of regulatory elements during development and uses epigenome annotations to document inappropriate transcriptional regulation in disease. First, I summarize my contributions to developing a new algorithm for detecting differential DNA methylation, M&M. I report the application of the M&M algorithm to identify distinct classes of DNA methylation dynamics in surface ectoderm (SE) progenitor cells and SE-derived lineages: epigenome alterations, and differential DNA methylation in …

Functions Of The Dna Polymerase Delta Replicase In Lagging Strand Replication, Joseph L. Stodola

Arts & Sciences Electronic Theses and Dissertations

The work described in this dissertation focuses on several aspects of DNA replication in the model organism Saccharomyces cerevisiae, with particular attention paid to the function of the replicative DNA polymerase delta (Pol ), and its functions in Okazaki fragment synthesis and maturation. The first major theme of this dissertation is investigating the role that metal binding motifs play in the structure and function of Pol and other budding yeast polymerases. First, I discuss the role that two metal binding motifs within the catalytic subunit of Pol play in creating the multi-subunit polymerase complex and in promoting crucial interactions with …

The Role Of Epidermal Enhancer 923 In The Chromatin Architecture And Transcriptional Regulation Of The Epidermal Differentiation Complex, Inez Oh

Arts & Sciences Electronic Theses and Dissertations

The epidermis covers the surface of the skin and provides a functional barrier across the entire body. Epidermal cells or keratinocytes proliferate in the innermost basal layer and migrate upwards into the suprabasal spinous and granular layers as they differentiate, and finally into the terminally differentiated outermost stratum corneum. Keratinocytes undergoing terminal differentiation are marked by tissue-specific concomitant expression of genes encoded in the Epidermal Differentiation Complex (EDC) locus. The EDC genes are organized into four gene families - S100, Sprr, Lce, and Flg-like, which are coordinately expressed upon activation of the terminal differentiation program in keratinocytes. The molecular mechanisms …

Use Of Proteomics To Probe Dynamic Changes In Cyanobacteria, Amelia Yen Nguyen

Arts & Sciences Electronic Theses and Dissertations

Cyanobacteria are unicellular photosynthetic microorganisms that capture and convert light energy to chemical energy, which is the precursor for feed, fuel, and food. These oxygenic phototrophs appear blue-green in color due to the blue bilin pigments in their phycobilisomes and green chlorophyll pigments in their photosystems. They also have diverse morphologies, and thrive in terrestrial, marine water, fresh water, as well as extreme environments. Cyanobacteria have developed a number of protective mechanisms and adaptive responses that allow the photosynthetic process to operate optimally under diverse and extreme conditions. Prolonged deprivation of essential nutrients, such as nitrogen and sulfur, commonly found …

Perceiving Oldness In Parietal Cortex: Fmri Characterization Of A Parietal Memory Network, Adrian Gilmore

Arts & Sciences Electronic Theses and Dissertations

The manner in which the human brain recognizes certain stimuli as novel or familiar is a matter of ongoing investigation. The overarching goal of this dissertation is to improve our understanding of how this may be accomplished. More specifically, work contained herein focuses on a recently described "parietal memory network" (PMN; Gilmore et al., 2015) that shows opposite patterns of activity when perceiving novel or familiar stimuli: deactivating in response to novelty, and activating in response to familiarity. Critically, our understanding of this network is based on explicit memory tasks, in which subjects are deliberately instructed to learn or remember …

Mechanisms Of Extracellular Vesicle Biogenesis, Charles Eldon Jackson

Arts & Sciences Electronic Theses and Dissertations

Extracellular vesicles (EVs) are important mediators of intercellular communication. Different types of EVs are released from cells by either fusion of late endsomal multivesicular bodies with the plasma membrane (exosomes) or direct budding from the plasma membrane (ectosomes). Topologically equivalent processes including intralumenal vesicle formation for degradation in the endosomal pathway and virus budding from the plasma membrane depend on the ATPase VPS4 and its Endosomal Sorting Complex Required for Transport (ESCRT)-III substrates for membrane fission and release. Whether this machinery is generally required for EV biogenesis has, however, been the subject of debate. Studies of the EV proteome from …

Actin-Based Feedback Circuits In Cell Migration And Endocytosis, Xinxin Wang

Arts & Sciences Electronic Theses and Dissertations

In this thesis, we study the switch and pulse functions of actin during two important cellular processes, cell migration and endocytosis. Actin is an abundant protein that can polymerize to form a dendritic network. The actin network can exert force to push or bend the cell membrane. During cell migration, the actin network behaves like a switch, assembling mostly at one end or at the other end. The end with the majority of the actin network is the leading edge, following which the cell can persistently move in the same direction. The other end, with the minority of the actin …

Regulation Of Transcription And Stress Response By Card In Mycobacteria, Ashley Louise Garner

Arts & Sciences Electronic Theses and Dissertations

Mycobacterium tuberculosis, the causative agent of Tuberculosis, infects over one third of the world's population. To control this epidemic, we must develop new chemotherapeutic strategies for treatment, which requires further insight into the physiology of this bacterium. Previous studies have identified CarD as a transcriptional regulator essential during both acute and persistent infection. Depletion of CarD sensitizes strains to a diverse panel of stresses and deregulates several hundred genes and ribosomal RNA (rRNA) which suggested that CarD may be a transcriptional regulator. Chromatin immunoprecipitation sequencing experiments showed that CarD was localized to promoters throughout the genome, suggesting that CarD regulates …

Genetic Imputation: Accuracy To Application, Shelina Raynell Ramnarine

Arts & Sciences Electronic Theses and Dissertations

Genotype imputation, the process of inferring genotypes for untyped variants, is used to identify and refine genetic association findings. This body of work focuses on assessing imputation accuracy and uses imputed data to identify genetic contributors to mentholated cigarette preference.

Inaccuracies in imputed data can distort the observed association between variants and a disease. Many statistics are used to assess accuracy; some compare imputed to genotyped data and others are calculated without reference to true genotypes. Prior work has shown that the Imputation Quality Score (IQS), which is based on Cohens kappa statistic and compares imputed genotype probabilities to true …

Role Of Atg16l1 In Uropathogenic E. Coli Pathogenesis, Jane Symington

Arts & Sciences Electronic Theses and Dissertations

Urinary tract infections (UTIs) are among the most common infectious diseases and are primarily caused by uropathogenic E. coli (UPEC). Given the greater incidence of antibiotic resistance among UPEC isolates, it is vital to determine factors and pathways important for an effective host response to UPEC in order to improve therapeutic options for combating UTIs. Autophagy is a cellular degradation pathway that plays important roles in pathogen control and modulation of innate immunity. One essential autophagy protein, ATG16L1, has been further implicated in controlling inflammation due to a common variant of ATG16L1 being associated with increased risk of Crohns disease, …

Metal Selectivity And Acquisition By The Yersiniabactin Metallophore System In Escherichia Coli, Eun-Ik Koh

Arts & Sciences Electronic Theses and Dissertations

Urinary tract infections (UTI) are one of the most common bacterial infections, of which, the majority are caused by uropathogenic Escherichia coli (UPEC). Many bacterial pathogens including UPEC synthesize and secrete chemically diverse metabolites called siderophores, which are classically defined by their ability of bind and deliver iron(III), an essential nutrient, to pathogens during infections. UPEC isolates can express multiple siderophore systems, of which, the virulence-associated siderophore yersiniabactin (Ybt) binds to both iron(III) and copper(II) during urinary tract infections. In this thesis we show that Ybt interacts with multiple physiologic transition metals and acts as a metallophore system to deliver …

Mechanism Of Calcium-Dependent Chloride Channel Activation By The Secreted Regulator Clca1, Zeynep Yurtsever

Arts & Sciences Electronic Theses and Dissertations

The calcium-activated chloride channel regulator (CLCA) proteins are key signaling molecules, which are implicated in various diseases through their tissue-specific expression. Human CLCA1 protein, overexpressed in airway epithelia under pathophysiological conditions, is centrally involved in the manifestation of IL-13-driven mucus cell metaplasia (MCM), a hallmark feature of asthma and chronic obstructive pulmonary disease (COPD), for which there are currently no available therapeutics. Elucidating the poorly understood molecular basis of CLCA1 function is thus required to design specific inhibitors of CLCA1 activity to treat MCM in asthma and COPD.

Originally misannotated as ion channels, CLCA proteins are secreted soluble proteins that …

A Quantitative Genetic Analysis Of Craniofacial Variation In Baboons, Jessica Lynn Joganic

Arts & Sciences Electronic Theses and Dissertations

This dissertation is an explication of baboon craniofacial variation and its genetic basis. Intraspecific variation is the result of input from and complex interactions among genetic information, functional demands, and developmental processes. The relative effect of each of these on craniofacial variation, as well as the degree of inter-trait covariance, determines whether traits can respond to selection and what that response might look like. Using a sample of pedigreed baboons, I quantify craniofacial variation to address specific questions regarding the distribution and magnitude of phenotypic, genetic, and environmental variation patterns. In addition, I identify regions of the genome containing genetic …

Structure And Dynamics Of A Small Multidrug Resistance Transporter, Emre, Chao Wu

Arts & Sciences Electronic Theses and Dissertations

EmrE is a small multidrug resistance transporter in E. coli. It effluxes a wide range of antibiotics, thus contributing to the evolving epidemic of drug resistance. Despite its small size, EmrE is a fully functional transporter making it an ideal model system for a comprehensive study of the multidrug transport mechanism. In the transport cycle, EmrE must alternate between outward- and inward-facing conformations upon substrate binding to translocate substrates across the membrane. High-resolution structures of EmrE in complex with substrates facing different sides of the membrane will shed light on the coupling mechanism between substrate binding and transport. However, the …

Modulation Of Human And Malarial Glucose Transporter Activity By Lipids And Small Molecules, Thomas E. Kraft

Arts & Sciences Electronic Theses and Dissertations

Glucose transport is a fundamentally important process for maintenance and regulation of cellular metabolism in all kingdoms of life. Despite their high importance, detailed examination of glucose transport proteins in humans and parasites through biochemical, biophysical and structural properties was greatly hampered by the inability to express, purify and reconstitute sufficient amounts of active transporters. This dissertation describes strategies that led to the first successful expression, purification, stabilization and functional reconstitution of active insulin-responsive GLUT4 transport protein. Furthermore, the work described herein establishes a requirement of anionic and conical lipids for full activity of the mammalian glucose transporters GLUT3 and …

Development And Application Of Comparative Gene Co-Expression Network Methods In Brachypodium Distachyon, Henry David Priest

Arts & Sciences Electronic Theses and Dissertations

Gene discovery and characterization is a long and labor-intensive process. Gene co-expression network analysis is a long-standing powerful approach that can strongly enrich signals within gene expression datasets to predict genes critical for many cellular functions. Leveraging this approach with a large number of transcriptome datasets does not yield a concomitant increase in network granularity. Independently generated datasets that describe gene expression in various tissues, developmental stages, times of day, and environments can carry conflicting co-expression signals. The gene expression responses of the model C3 grass Brachypodium distachyon to abiotic stress is characterized by a co-expression-based analysis, identifying 22 modules …

The Resolution Phase Of Nk Cell Proliferation And Ifn Production Following Viral Infection Are Highly Regulated Processes., Leslie Abigail Fogel

Arts & Sciences Electronic Theses and Dissertations

In response to MCMV infection, NK cells undergo three distinct phases of proliferation: the non-specific phase mediated by pro-proliferative cytokines; the specific phase mediated by recognition of an MCMV-encoded protein by an NK cell activating receptor, Ly49H; and the resolution phase, whose mechanism is unknown. MCMV infection of RAG mice, which lack all adaptive immune cells, results in prolonged proliferation of NK cells despite similar viral titers compared to wildtype mice. Interestingly, there are different kinetics for Ly49H+ and Ly49H- NK cells. We have identified several additional markers that may distinguish NK cells that have been specifically activated through their …

The Molecular And Cellular Basis For Cold Sensation, Daniel Brenner

Arts & Sciences Electronic Theses and Dissertations

The ability to sense changes in temperature is crucial to surviving harsh environments. Over the last decade several ion channels that have been proposed to be cold sensitive have been identified, most notably TRPM8 and TRPA1. Although these molecules have been extensively studied in vitro, their exact roles in cold sensation in vivo are still debated. This uncertainty is in large part due to problems with the standard methods of testing cold sensitivity in vivo, which often rely on subjective measures of cold responsiveness. Experiments using these subjective measures have been repeated by different groups and have yielded conflicting results, …

A Novel Ultrashort Pulsed Laser Method For Pathogen Inactivation, Shaw-Wei David Tsen

Arts & Sciences Electronic Theses and Dissertations

Emerging viral pathogens represent a constant and prominent threat to human health worldwide. There is a dire need to develop new pathogen inactivation techniques that can be used to safeguard the blood supply and to generate inactivated vaccines against these emerging viruses. Unfortunately, current techniques are inadequate because they introduce toxic/carcinogenic chemicals or alter the structure of the products, leading to loss of safety and efficacy. We have developed a novel, chemical-free ultrashort pulsed laser treatment system capable of inactivating a broad spectrum of both enveloped and non-enveloped viruses. We found that ultrashort pulsed laser treatment targets the vibrational modes …

The Role Of Dnmt3a In Acute Myeloid Leukemia Pathogenesis, Christopher Browning Cole

Arts & Sciences Electronic Theses and Dissertations

Loss of function mutations in the DNA methyltransferase DNMT3A are highly recurrent in acute myeloid leukemia (AML). DNMT3A and the highly homologous gene DNMT3B encode the two methyltransferases that are primarily responsible for mediating de novo methylation of specific DNA sequences during cellular differentiation. DNMT3A mutations are mutually exclusive of several translocations that create oncogenic fusion genes (PML-RARA, RUNX1-RUNX1T1, CBFB-MYH11, and MLL-X), suggesting that these fusions may require functional DNMT3A to initiate leukemogenesis. Using bone marrow cells from a constitutive Dnmt3a null mouse, we show that loss of Dnmt3a caused a striking loss of DNA methylation throughout the genome of …

Refining Associations Between Targeted Genes And The Development Of Substance Use Disorders, Emily Olfson

Arts & Sciences Electronic Theses and Dissertations

Recent genome-wide association studies (GWAS) provide strong evidence for the contribution of a few specific genes to alcohol and nicotine dependence. Chapter 2 explores numerous previously identified candidate genes for alcohol dependence using a publicly available GWAS. I found that many candidate loci do not replicate, highlighting the utility of GWAS for focusing on disease associated genes. Chapters 3-5 dissect associations between three genome-wide significant genes and substance use disorders. Chapter 3 focuses on a functional variant in alcohol dehydrogenase (ADH) 1B. Through examining 1,550 adolescent drinkers in the Collaborative Study on the Genetics of Alcoholism (COGA), I extended adult …

Granulocyte-Colony Stimulating Factor Reprograms The Bone Marrow Microenvironment To Suppress B Lymphopoiesis, Ryan Brent Day

Arts & Sciences Electronic Theses and Dissertations

The production of hematopoietic cells in the bone marrow is tightly and dynamically regulated in response to environmental stimuli. In response to infection, the bone marrow increases granulopoiesis at the expense of lymphopoiesis. The mechanisms mediating this shift are poorly understood. We show that treatment with granulocyte-colony stimulating factor (G-CSF), which is often induced during infection, results in marked decline of B lymphocytes at multiple stages of bone marrow B cell development. Transgenic mouse models show that G-CSF acts in a non-cell intrinsic fashion through cells of the monocyte-macrophage lineage to suppress B lymphopoiesis by downregulating important B trophic factors …

Omcp Mediated Cowpox Virulence And Its Dependence On The Immune Receptors Nkg2d And Fcrl5., Michel Muzi Sun

Arts & Sciences Electronic Theses and Dissertations

Viruses with large DNA genomes, such as cowpox virus, encode many open-reading frames involved in the modulation of the host immune system, facilitating escape from immune detection or downregulation of specific aspects of the host immune response. Investigation of virally-encoded immunoevasins has been instrumental in understanding host-pathogen interactions. Here, we focus on the cowpox virus immunoevasin Orthopoxvirus MHC Class I-like Protein (OMCP) and demonstrate for the first time that OMCP facilitates cowpox virus virulence in vivo. We have previously documented that OMCP binds the activating receptor NKG2D on NK cells as well as the orphan receptor FCRL5 on innate B …

Engineering Cre Recombinase For Genome Engineering, Chi Zhang

Arts & Sciences Electronic Theses and Dissertations

Cre recombinase recombines its DNA target, loxP sites, without help of accessory proteins or DNA repair systems. The simplicity of Cre-lox system has been widely utilized for genome editing, especially in mouse genetics. The goal of this dissertation is to constructCrerecombinase variants that will operate uponrecombination target sites (RTs) present within the genome, instead of perturbing the genome by inserting wildtype RTs for subsequent genome engineering. In general, the desired RTs native to the genome are asymmetric. However, the loxP sequence is pseudo-palindromic, requiring a homotetrameric formation of Cre recombinase. As a first step, I broke the symmetry of Cre …

Iron Regulation Of Macrophage Responses To Uropathogenic E. Coli, Nana Kwame Owusu-Boaitey

Arts & Sciences Electronic Theses and Dissertations

Uropathogenic E. coli (UPEC) are the principal cause of urinary tract infections (UTIs), one of the most common infections globally. Given the rising incidence of antibiotic resistance among UPEC strains, there is an increasing need to better understand the host response to UPEC and to develop ways to harness the bladder innate immune response that clears infection. In response to infection, the host attempts to limit the ability of UPEC to access iron, a metal critical to UPEC survival. Innate immune cells known as macrophages are known to regulate iron homeostasis through the production of pro-inflammatory cytokines, though it remains …

Superoxide Dismutase 1: Novel Insights On Disease Models And Tissue Specificity In Amyotrophic Lateral Sclerosis, Matthew James Crisp

Arts & Sciences Electronic Theses and Dissertations

Mutations in superoxide dismutase 1 (SOD1) are known to cause dominantly-inherited amyotrophic lateral sclerosis (ALS), a rapidly-fatal adult-onset neurodegenerative disorder defined by motor neuron loss and progressive paralysis. In the past twenty years, research into the disorder has been driven by the creation of numerous transgenic animal models that have yielded multiple theories on the pathogenesis of the disease. Patients and animal models with SOD1 mutations express the defective protein in every cell, yet the disease only affects tissues in the neuromuscular axis. In this dissertation, I present original work exploring two aspects of SOD1 ALS. The first details the …

Application Of Genomic Technologies To Study Infertility, Nicholas Rui Yuan Ho

Arts & Sciences Electronic Theses and Dissertations

An estimated one in eight couples in the United States are diagnosed with infertility. There is a significant genetic contribution to infertility, with estimates of heritability ranging from 0.2 to 0.5. We know surprisingly little about the genetic causes, with only slightly more than a hundred genes known to cause human infertility. I have been translating recent advances in genomics to study infertility in a more efficient manner, in order to improve our knowledge of the genetic causes. By using high throughput genomics and proteomics datasets from other groups, I was able to feed that into a machine learning algorithm …

Understanding The Interactions Of Developing Thymocytes And Antigen Presenting Cells In The Thymic Medulla, Justin Shaun Arnold Perry

Arts & Sciences Electronic Theses and Dissertations

Various thymic APC subsets have been invoked in deletional tolerance and Treg cell induction, but previous studies either assessed total T cell numbers or used TCR transgenic lines, obscuring roles that individual thymic APC subsets might serve for particular antigen-specific T cell populations. Utilizing T cell receptor sequencing, we found that medullary thymic epithelial cells (mTECs) and bone marrow-derived (BM) APCs delete or select unique conventional and Treg cell TCR repertoires, demonstrating distinct roles for these APCs. We show that BM APCs and mTECs each contribute to Aire-dependent T cell tolerance development, albeit through either cooperative or autologous antigen presentation …