Microbiology Commons^™

C-Jun N-Terminal Kinases Regulate Adenovirus-Mediated Autophagy And Antigen Presentation, Sarah R. Klein

Dissertations and Theses (Open Access)

Targeted immunotherapy with recombinant, oncolytic adenoviruses is under investigation for the treatment of cancer. Evidence indicates adenoviruses induce autophagy that is required for oncolysis, but the molecular regulation of autophagy in infected cells remains under investigation. Our data suggested the canonical pathway regulating starvation-induced autophagy was not implemented in adenovirus-induced autophagy; however, adenovirus infection triggered phosphorylation of c-Jun N-terminal kinases (JNK) that was essential for autophagy. Adenoviral replication within the host cell elicited JNK pathway activation leading to B cell lymphoma-2 (Bcl-2) phosphorylation. JNK-dependent Bcl-2 phosphorylation stimulated the dissociation of Bcl-2/beclin 1 heterodimers, enabling beclin 1 to initiate autophagy. Moreover, …

Development And Characterization Of An In Vitro Four-Species Anaerobic Dental Biofilm Model, Fernando Andrade

Dissertations and Theses (Open Access)

Dental caries is the most common chronic disease worldwide. It is characterized by the demineralization of tooth enamel caused by acid produced by cariogenic dental bacteria growing on tooth surfaces, termed bacterial biofilms. Cariogenesis is a complex biological process that is influence by multiple factors and is not attributed to a sole causative agent. Instead, caries is associated with multispecies microbial biofilm communities composed of some bacterial species that directly influence the development of a caries lesion and other species that are seemingly benign but must contribute to the community in an uncharacterized way. Clinical analysis of dental caries and …

Interaction Of Bacillus Anthracis Exosporium Protein Bcla With Complement Factor H And Spore Persistence In The Lung, Sarah A. Jenkins

Dissertations and Theses (Open Access)

Anthrax outbreaks in the United States and Europe and its potential use as a bioweapon have made Bacillus anthracis an interest of study. Anthrax infections are caused by the entry of B. anthracis spores into the host via the respiratory system, the gastrointestinal tract, cuts or wounds in the skin, and injection. Among these four forms, inhalational anthrax has the highest lethality rate and persistence of spores in the lungs of animals following pulmonary exposure has been noted for decades. However, details or mechanisms of spore persistence were not known. In this study, we investigated spore persistence in a mouse …

Molecular And Genomic Based Insights Into The Evolution Of Enterococcus Faecium From Commensal To Hospital-Adapted Pathogen, Jessica R. Galloway-Pena

Dissertations and Theses (Open Access)

The basis for the recent transition of Enterococcus faecium from a primarily commensal organism to one of the leading causes of hospital-acquired infections in the United States is not yet understood. To address this, the first part of my project assessed isolates from early outbreaks in the USA and South America using sequence analysis, colony hybridizations, and minimal inhibitory concentrations (MICs) which showed clinical isolates possess virulence and antibiotic resistance determinants that are less abundant or lacking in community isolates. I also revealed that the level of ampicillin resistance increased over time in clinical strains. By sequencing the pbp5 gene, …

Biochemical Characterization Of Binding Partners Of Two Hsp70 Co-Chaperones In Saccharomyces Cerevisiae, Jacob Verghese

Dissertations and Theses (Open Access)

Cells are exposed to a variety of environmental and physiological changes including temperature, pH and nutrient availability. These changes cause stress to cells, which results in protein misfolding and altered cellular protein homeostasis. How proteins fold into their three-dimensional functional structure is a fundamental biological process with important relevance to human health. Misfolded and aggregated proteins are linked to multiple neurodegenerative diseases, cardiovascular disease and cystic fibrosis. To combat proteotoxic stress, cells deploy an array of molecular chaperones that assist in the repair or removal of misfolded proteins.

Hsp70, an evolutionarily conserved molecular chaperone, promotes protein folding and helps maintain …

Regulation Of Alternative Carbon Metabolism In Candida Albicans, Arely Y. Gonzalez

Dissertations and Theses (Open Access)

Candida albicans is the most important fungal pathogen of humans. Transcript profiling studies show that upon phagocytosis by macrophages, C. albicans undergoes a massive metabolic reorganization activating genes involved in alternative carbon metabolism, including the glyoxylate cycle, β-oxidation and gluconeogenesis. Mutations in key enzymes such as ICL1 (glyoxylate cycle) and FOX2 (fatty acid β-oxidation) revealed that alternative carbon metabolic pathways are required for full virulence in C. albicans. These studies indicate C. albicans uses non-preferred carbon sources allowing its adaptation to microenvironments were nutrients are scarce. It has become apparent that the regulatory networks required for regulation of alternative …

The Role Of The Arched Helicases In Exosome-Mediated Function, A. Alejandra Klauer

Dissertations and Theses (Open Access)

RNA processing and degradation are two important functions that control gene expression and promote RNA fidelity in the cell. A major ribonuclease complex, called the exosome, is involved in both of these processes. The exosome is composed of ten essential proteins with only one catalytically active subunit, called Rrp44. While the same ten essential subunits make up both the nuclear and cytoplasmic exosome, there are nuclear and cytoplasmic exosome cofactors that promote specific exosome functions in each of the cell compartments. To date, it is unclear how the exosome distinguishes between RNA substrates. We hypothesize that compartment specific cofactors may …

Regulation Of The Heat Shock Response By Thiol-Reactive Compounds In The Yeast Saccharomyces Cerevisiae, Yanyu Wang

Dissertations and Theses (Open Access)

Cells govern their activities and modulate their interactions with the environment to achieve homeostasis. The heat shock response (HSR) is one of the most well studied fundamental cellular responses to environmental and physiological challenges, resulting in rapid synthesis of heat shock proteins (HSPs), which serve to protect cellular constituents from the deleterious effects of stress. In addition to its role in cytoprotection, the HSR also influences lifespan and is associated with a variety of human diseases including cancer, aging and neurodegenerative disorders. In most eukaryotes, the HSR is primarily mediated by the highly conserved transcription factor HSF1, which recognizes target …

Systems Biology Approaches To Probe Gene Regulation In Bacteria, Diogo F. Troggian Veiga

Dissertations and Theses (Open Access)

Mechanisms that allow pathogens to colonize the host are not the product of isolated genes, but instead emerge from the concerted operation of regulatory networks. Therefore, identifying components and the systemic behavior of networks is necessary to a better understanding of gene regulation and pathogenesis. To this end, I have developed systems biology approaches to study transcriptional and post-transcriptional gene regulation in bacteria, with an emphasis in the human pathogen Mycobacterium tuberculosis (Mtb).

First, I developed a network response method to identify parts of the Mtb global transcriptional regulatory network utilized by the pathogen to counteract phagosomal stresses …

Regulation Of To Xin Synthesis By Clostridium Difficile, Charles Darkoh

Dissertations and Theses (Open Access)

Clostridium difficile is the leading definable cause of nosocomial diarrhea worldwide due to its virulence, multi-drug resistance, spore-forming ability, and environmental persistence. The incidence of C. difficile infection (CDI) has been increasing exponentially in the last decade. Virulent strains of C. difficile produce either toxin A and/or toxin B, which are essential for the pathogenesis of this bacterium. Current methods for diagnosing CDI are mostly qualitative tests that detect the bacterium, the toxins, or the toxin genes. These methods do not differentiate virulent C. difficile strains that produce active toxins from non-virulent strains that do not produce toxins or produce …

Functional Characterization Of Atxa, The Bacillus Anthracis Virulence Regulator, Troy G. Hammerstrom

Dissertations and Theses (Open Access)

Coordinated expression of virulence genes in Bacillus anthracis occurs via a multi-faceted signal transduction pathway that is dependent upon the AtxA protein. Intricate control of atxA gene transcription and AtxA protein function have become apparent from studies of AtxA-induced synthesis of the anthrax toxin proteins and the poly-D-glutamic acid capsule, two factors with important roles in B. anthracis pathogenesis. The amino-terminal region of the AtxA protein contains winged-helix (WH) and helix-turn-helix (HTH) motifs, structural features associated with DNA-binding. Using filter binding assays, I determined that AtxA interacted non-specifically at a low nanomolar affinity with a target promoter (Plef) …

Control Of The Master Virulence Regulatory Gene Atxa In Bacillus Anthracis, Jennifer L. Dale

Dissertations and Theses (Open Access)

Transcription of the Bacillus anthracis structural genes for the anthrax toxin proteins and biosynthetic operon for capsule are positively regulated by AtxA, a transcription regulator with unique properties. Consistent with the role of atxA in virulence factor expression, a B. anthracis atxA-null mutant is avirulent in a murine model for anthrax. In batch culture, multiple signals impact atxA transcript levels, and the timing and steady state level of atxA expression is critical for optimal toxin and capsule synthesis. Despite the apparent complex control of atxA transcription, only one trans-acting protein, the transition state regulator AbrB, has been demonstrated …

Upregulation Of Reactive Oxygen Species During The Retrovirus Life Cycle And Their Roles In A Mutant Of Moloney Murine Leukemia Virus, Ts1-Mediated Neurodegeneration, Soo Jin Kim

Dissertations and Theses (Open Access)

Viral invasion of the central nervous system (CNS) and development of neurological symptoms is a characteristic of many retroviruses. The mechanism by which retrovirus infection causes neurological dysfunction has yet to be fully elucidated. Given the complexity of the retrovirus-mediated neuropathogenesis, studies using small animal models are extremely valuable. Our laboratory has used a mutant moloney murine leukemia retrovirus, ts1-mediated neurodegneration. We hypothesize that astrocytes play an important role in ts1-induced neurodegeneration since they are retroviral reservoirs and supporting cells for neurons. It has been shown that ts1 is able to infect astrocytes in vivo and in …

Novel Mechanisms Of Antigen Processing That Enhance Bcg Vaccine Efficacy, Christopher R. Singh

Dissertations and Theses (Open Access)

Mycobacterium tuberculosis, the causative agent of tuberculosis, is the most lethal single infectious agent afflicting man today causing 2 million deaths per year. The World Health Organization recommends a vaccine as the best option to prevent this disease. The current vaccine, BCG, has a variable efficacy and does not protect adults. It is known that BCG vaccine becomes sequestered in special phagosome compartments of macrophages that do not fuse with lysosomes. Since lysosome fusion is necessary for peptide production and T cell priming leading to protective T_H1 immunity, we hypothesized that vaccine efficacy is reduced and occurs perhaps …

Mechanisms Of Adenovirus-Mediated Autophagy, Erin White

Dissertations and Theses (Open Access)

A patient diagnosed with a glioma, generally, has an average of 14 months year to live after implementation of conventional therapies such as surgery, chemotherapy, and radiation. Glioblastomas are highly lethal because of their aggressive nature and resistance to conventional therapies and apoptosis. Thus other avenues of cell death urgently need to be explored. Autophagy, which is also known as programmed cell death type II, has recently been identified as an alternative mechanism to kill apoptosis- resistant cancer cells. Traditionally, researchers have studied how cells undergo autophagy during viral infection as an immune response mechanism, but recently researchers have discovered …

Mechanisms Of Adenovirus-Mediated Autophagy, Erin White

Dissertations and Theses (Open Access)

A patient diagnosed with a glioma, generally, has an average of 14 months year to live after implementation of conventional therapies such as surgery, chemotherapy, and radiation. Glioblastomas are highly lethal because of their aggressive nature and resistance to conventional therapies and apoptosis. Thus other avenues of cell death urgently need to be explored. Autophagy, which is also known as programmed cell death type II, has recently been identified as an alternative mechanism to kill apoptosis- resistant cancer cells. Traditionally, researchers have studied how cells undergo autophagy during viral infection as an immune response mechanism, but recently researchers have discovered …

Eluding Antibiotic Resistance: Capitalizing On Antimicrobial Peptides Interaction With The Lipid Bilayer, Danielle M. Mcgrath

Dissertations and Theses (Open Access)

It is widely accepted that the emergence of drug-resistant pathogens is the result of the overuse and misuse of antibiotics. Infectious Disease Society of America, Center for Disease Control and World Health Organization continue to view, with concern, the lack of antibiotics in development, especially those against Gram-negative bacteria.

Antimicrobial peptides (AMPs) have been proposed as an alternative to antibiotics due to their selective activity against microbes and minor ability to induce resistance. For example, the Food and Drug Administration approved Daptomycin (DAP) in 2003 for treatment of severe skin infections caused by susceptible Gram-positive organisms. Currently, there are 12 …

Role Of Vitamin-D3 And Retinoic Acid In A Human Thp-1 Macrophage Model Of Mycobacterium Tuberculosis Infection, Jaymie L. Estrella

Dissertations and Theses (Open Access)

Mycobacterium tuberculosis (Mtb) replicates within the human macrophages and we investigated the activating effects of retinoic acid (RA) and vitamin D3 (VD) on macrophages in relation to the viability of Mtb. A combination of these vitamins (RAVD) enhanced the receptors on THP-1 macrophage (Mannose receptor and DC-SIGN) that increased mycobacterial uptake but inhibited the
subsequent intracellular growth of Mtb by inducing reactive oxygen species (ROS) and autophagy. RAVD also enhanced antigen presenting and homing receptors in THPs that suggested an activated phenotype for THPs following RAVD treatment. RAVD mediated activation was also associated with a marked phenotypic change in Mtb …

Multiple Posttranscriptional Regulatory Features Control Expression Of Ethanolamine Utilization Genes In Enterococcus Faecalis, Kristina A. Fox

Dissertations and Theses (Open Access)

Enterococcus faecalis is a Gram-positive bacterium that lives as a commensal organism in the mammalian gastrointestinal tract, but can behave as an opportunistic pathogen.  Our lab discovered that mutation of the eutK gene attenuates virulence of E. faecalis in the C. elegans model host.  eutK is part of the ethanolamine metabolic pathway which was previously unknown in E. faecalis.  I discovered the presence of two unique posttranscriptional regulatory features that control expression of eut locus genes.  The first feature I found is an AdoCBL riboswitch, a cis-acting RNA regulatory element that acts as a positive regulator of gene …

The Immune Inhibitor A1 Protease Of Bacillus Anthracis, Kathryn Pflughoeft

Dissertations and Theses (Open Access)

Bacillus anthracis, an organism ubiquitous in the soil and the causative agent of anthrax, utilizes multiple mechanisms to regulate secreted factors; one example is the activity of secreted proteases. One of the most abundant proteins in the culture supernates of B. anthracis is the Immune Inhibitor A1 (InhA1) protease. Here, I demonstrate that InhA1 modulates the abundance of approximately half of the proteins secreted into the culture supernates, including substrates that are known to contribute to the ability of the organism to cause virulence. For example, InhA1 cleaves the anthrax toxin proteins, PA, LF, and EF. InhA1 also targets …

Stress-Induced Targeting Of Molecular Chaperones In The Yeast Saccharomyces Cerevisiae, Hugo Tapia

Dissertations and Theses (Open Access)

The eukaryotic stress response is an essential mechanism that helps protect cells from a variety of environmental stresses. Cell death can result if cells are not able to properly adapt and protect themselves against adverse stress conditions. Failure to properly deal with stress has implications in human diseases including neurodegenerative disorders and distinct cancers, emphasizing the importance of understanding the eukaryotic stress response in detail. As part of this response, expression of a battery of heat shock proteins (HSP) is induced, which act as molecular chaperones to assist in the repair or triage of unfolded proteins. The 90-kDa HSP (Hsp90) …

Characterization Of The Agrobacterium Tumefaciens Virb2 Pilin Of The Virb/D4 Type Iv Secretion System, Jennifer Kerr

Dissertations and Theses (Open Access)

The Agrobacterium tumefaciens VirB/D4 type IV secretion system (T4SS) delivers oncogenic T-DNA and effector proteins to susceptible plant cells. This leads to the formation of tumors termed Crown Galls. The VirB/D4 T4SS is comprised of 12 subunits (VirB1 to VirB11 and VirD4), which assemble to form two structures, a secretion channel spanning the cell envelope and a T-pilus extending from the cell surface. In A. tumefaciens, the VirB2 pilin subunit is required for assembly of the secretion channel and is the main subunit of the T-pilus. The focus of this thesis is to define key reactions associated with the T4SS …

Agrobacterium Virb10 Contributions To Type Iv Substrate Secretion, T-Pilus Biogenesis, And Outer Membrane Pore Formation, Isaac Garza

Dissertations and Theses (Open Access)

The VirB/D4 type IV secretion system (T4SS) of Agrobacterium tumefaciens functions to transfer substrates to infected plant cells through assembly of a translocation channel and a surface structure termed a T-pilus. This thesis is focused on identifying contributions of VirB10 to substrate transfer and T-pilus formation through a mutational analysis. VirB10 is a bitopic protein with several domains, including a: (i) cytoplasmic N-terminus, (ii) single transmembrane (TM) α-helix, (iii) proline-rich region (PRR), and (iv) large C-terminal modified β-barrel. I introduced cysteine insertion and substitution mutations throughout the length of VirB10 in order to: (i) test a predicted transmembrane topology, (ii) …

The Domains Of The Catalytic Subunit Of The Eukaryotic Rna Degrading Exosome, Rrp44p, Have Distinct Functions, Daneen Schaeffer

Dissertations and Theses (Open Access)

The exosome is a 3’ to 5’ exoribonuclease complex that consists of ten essential subunits. In the cytoplasm, the exosome degrades mRNA in a general mRNA turnover pathway and in several mRNA surveillance pathways. In the nucleus, the exosome processes RNA precursors to form small, stable, mature RNA species, including rRNA, snRNA, and snoRNA. In addition to processing these RNAs, the nuclear exosome is also involved in degrading aberrantly processed forms of these RNAs, and others, including mRNA.

The 3’ to 5’ exoribonuclease activity of the exosome is contributed by the RNB domain of the only catalytically active subunit, Rrp44p, …

A Metagenomic Study Of The Tick Midgut, Daniel T. Yuan

Dissertations and Theses (Open Access)

A Metagenomic Study of the Tick Midgut
Daniel Yuan, B.S.
Supervisory Professor : Steven J. Norris, Ph.D.
Southern tick–associated rash illness (STARI) or Master’s disease is a Lyme-like illness that occurs following bites by Amblyomma americanum, the lone-star tick. Clinical symptoms include a bull’s eye rash similar to the erythema migrans lesions of Lyme disease, as well as fever and joint pains. Lyme disease is caused by Borrelia burgdorferi and related spirochetes. However, B. burgdorferi has not been detected in STARI patients, or in ticks in the South Central U.S. The causative agent of STARI has not been identified, …

Host-Pathogen Interactions Of Secreted And Surface Staphylococcus Aureus Factors, Vanessa Vazquez

Dissertations and Theses (Open Access)

Staphylococcus aureus is an opportunistic bacterial pathogen that can infect humans and other species. It utilizes an arsenal of virulence factors to cause disease, including secreted and cell wall anchored factors. Secreted toxins attack host cells, and pore-forming toxins destroy target cells by causing cell lysis. S. aureus uses cell-surface adhesins to attach to host molecules thereby facilitating host colonization. The Microbial Surface Components Recognizing Adhesive Matrix Molecules (MSCRAMMs) are a family of cell-wall anchored proteins that target molecules like fibronectin and fibrinogen. The Serine-aspartate repeat (Sdr) proteins are a subset of staphylococcal MSCRAMMs that share similar domain organization. Interestingly, …

Molecular Basis Of Corynebacterium Diphtheriae Virulence And Infection In The Caenorhabditis Elegans Model Host, Melissa M. Broadway

Dissertations and Theses (Open Access)

Corynebacterium diphtheriae is the causative agent of cutaneous and pharyngeal diphtheria in humans. While lethality is certainly caused by diphtheria toxin, corynebacterial colonization may primarily require proteinaceous fibers called pili, which mediate adherence to specific tissues. The type strain of C. diphtheriae possesses three distinct pilus structures, namely the SpaA, SpaD, and SpaH-type pili, which are encoded by three distinct pilus gene clusters. The pilus is assembled onto the bacterial peptidoglycan by a specific transpeptidase enzyme called sortase. Although the SpaA pili are shown to be specific for pharyngeal cells in vitro, little is known about functions of the three …