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Novel Roles Of The Protein Tyrosine Phosphatase Shp2 In Non-Small Cell Lung Cancer, Valentina Schneeberger

USF Tampa Graduate Theses and Dissertations

The gene PTPN11 was identified in the early 1990s, and encodes the non-transmembrane protein tyrosine phosphatase SHP2. SHP2 is expressed ubiquitously in cells, and plays an important role in cancer. Unlike most phosphatases, SHP2 positively regulates several signaling pathways including the Ras/MAPK and Src signaling pathways and acts as a proto-oncogene. SHP2 is also a cancer essential gene in certain types of carcinomas, and promotes growth, survival, and epithelial to mesenchymal transformation. Gain of function (GOF) SHP2 mutations are known leukemic oncogenes, and have been identified to a smaller extent in solid tumors as well. Currently, the roles of SHP2 …

Defining The Sites Of Interaction Of The Fancd2, Fance, And Fancl Proteins, Joseph Mcclanaghan

Senior Honors Projects

Fanconi anemia (FA) is a rare genetic disease characterized by congenital defects, bone marrow failure and increased cancer susceptibility. FA is caused by mutations in any one of 16 genes. These genes encode for proteins that function in the FA-BRCA pathway to repair damaged DNA. Because of its important r­­­ole in DNA repair, this pathway is considered a major cellular tumor suppressor pathway, i.e. is critical for the prevention of cancer. Underscoring this fact, several of the FA genes - including BRCA2, BRIP1, PALB2, and RAD51C - are bona fide breast and ovarian cancer susceptibility genes.

My …

The Development Of A Comprehensive Antifungal Susceptibility Testing Assay For Vulvovaginal Candidiasis Therapy, David Hardaker

Graduate School of Biomedical Sciences Theses and Dissertations

Vulvovaginal candidiasis is the most common fungal infection of the female urogenital tract, commonly caused by Candida albicans and C. glabrata. However, treatment can be difficult when caused by non-albicans strains due to resistance to the oral antifungal drug fluconazole. The mechanism through which strains of Candida, particularly C. glabrata, develop resistance to different antifungal classes has not been completely characterized. The ergosterol biosynthesis pathway, an important component of the cell membrane, is the major target of antifungals such as fluconazole, an inhibitor of the ERG 11 gene. Current research shows that an upregulation of specific genes in C. glabrata …

Molecular Chaperone Tools For Use Against Neurodegenerative Diseases, Matthew Tinkham

Senior Honors Projects

A noted characteristic found in several neurodegenerative disorders, including Alzheimer’s Disease, Parkinson’s Disease, Huntington’s Disease and bovine spongiform encephalopathy, is the accumulation of amyloid plaques in the brain. Amyloid plaques contain deposits of fibrillar aggregates of misfolded proteins that disrupt normal functionality in neurons. Certain variants of these misfolded proteins are self-replicating; these self-replicating amyloids are termed prions (for infectious protein). We are interested in how protein misfolding contributes to amyloid formation and how molecular chaperone proteins can change the formation of amyloid deposits. Chaperone proteins function by catalyzing the proper folding of other proteins, the refolding of misfolded proteins, …

Biology And Control Of Rice False Smut Caused By Ustilaginoidea Virens (Teleomorph Villosiclava Virens), Andrew Clayton Jecmen

Graduate Theses and Dissertations

Rice false smut (FS), a disease caused by Ustilaginoidea virens (Cke.) Takahashi (1896), was first reported in northeastern Arkansas counties in 1997. The first objective of this research was to establish a collection of U. virens isolates from geographically diverse regions of Arkansas. Three U. virens isolates and chlamydospores from `Templeton' and `Clearfield-151' rice cultivars were used to determine the effects of temperature and pH on mycelial growth and germination. A nested-PCR protocol and histological methods were used to determine if U. virens infects and colonizes rice seedlings and spikelets on panicles. The sensitivity of three U. virens isolates was …

Phthalates And Phthalate Alternatives: Effects On Proliferative And Estrogenic Target Genes In Ishikawa Cells, Ranjani Sundar '15, Ping Yin, Serdar E. Bulun

Student Publications & Research

Phthalates are used as plasticizers in many of the products found in medical, household, and industrial applications. Much research has not been completed on the effects of these phthalates as potential endocrine disrupting chemicals (EDCs). As these chemicals are ingested, the mechanism by which they affect the reproductive system is largely unknown. The purpose of this study was to observe how 2 phthalates, Di-n-butyl phthalate (DBP) and Diisononyl phthalate (DINP), and 2 phthalate alternatives, Dioctyl terephthalate (DOTP) and BHT (butylated hydroxytoluene)affect uterine cells in comparison to a vehicle treatment and 17β-Estradiol treatment. Changes in expression of mRNA were observed using …

The Effect Of Small Molecule 390 On Cxcr4 Receptors, Selam B. Zenebe-Gete '14, Shruti R. Topudurti '14, Shum Andrew, Richard J. Miller

Student Publications & Research

CXCR4 is the chemokine receptor which aids in chemotaxis of stem cells, such as those in the bone marrow or the brain. SDF-1 is the natural ligand for the CXCR4 receptor. Similarities between novel molecule 390 synthesized by the Miller Lab and SDF-1 make this novel small molecule a possible agonist of the CXCR4 receptor. To determine whether 390 is an agonist to the CXCR4 receptor, we transfected cells with CXCR4 and exposed them to no agonist [vehicle control], SDF-1, or varying concentrations of our agonist drug. Next, we took calcium images using the dye fura-2, which indicates changes in …

Modeling The Adaptive Immune Response To Mutation-Generated Antigens, Rory J. Geyer

University Scholar Projects

Somatic mutations may drive tumorigenesis or lead to new, immunogenic epitopes (neoantigens). The immune system is thought to represses neoplastic growths through the recognition of neoantigens presented only by tumor cells. To study mutations as well as the immune response to mutation-generated antigens, we have created a conditional knockin mouse line with a gene encoding, 5’ to 3’, yellow fluorescent protein (YFP), ovalbumin (which is processed to the immunologically recognizable peptide, SIINFEKL), and cyan fluorescent protein (CFP), or, YFP-ovalbumin-CFP. A frame shift mutation has been created at the 5’ end of the ovalbumin gene, hence YFP should always be expressed, …

Molecular Mechanisms For Regulating The Assembly Of The Mixed Lineage Leukemia-1 (Mll1) Core Complex, Michael Hu

Renée Crown University Honors Thesis Projects - All

Epigenetics is the study of heritable traits that are not caused by alterations to DNA. An example of such a mechanism is histone methylation, specifically histone H3 lysine 4 (H3K4). Histones are proteins that wrap and package DNA into nucleosomes. Modifications to a histone’s chemistry can up or down regulate specific gene expression programs through changes in nucleosome positioning. Human mixed lineage leukemia protein-1 (MLL1) is a member of the SET1 family of H3K4 methyltransferases. Previous studies in the Cosgrove lab have shown that an interaction between two protein sub-complexes, MLL1 and WRAD (WDR5, RbBP5, Ash2L, DPY30), leads to dimethylation. …

The Purification And Characterization Of The Drosophila Melanogaster Trithorax Protein And Its Implications In The Studies Of The Set Domain Family Of Proteins, Kelsey Monteith

Renée Crown University Honors Thesis Projects - All

Methylation at histone H3 lysine 4 (H3K4) is a post-translational modification often associated with transcriptional regulation through altering the structural state of chromatin. The human mixed lineage leukemia protein-1 protein (MLL1) is a known histone methyltransferase that catalyzes the transfer of methyl groups to H3K4. MLL1 works in a core complex with other essential components, proteins WDR5, RbBP5, Ash2L, DPY-30 (WRAD), which is required for H3K4 dimethylation. Trithorax (TRX) protein is the Drosophila melanogaster ortholog to human MLL1, and although structurally similar is unable to perform dimethylation when in complex with the human components. The goal of this study is …

Mechanisms Of Action And Co-Optive Evolution For Hypervariable Courtship Pheromones In Plethodontid Salamanders., Wilburn Beau Wilburn

Electronic Theses and Dissertations

Pheromones are an important type of chemical cue used by most animals to convey information between individuals. For more than 100 million years, male plethodontid salamanders have utilized a system of non-volatile, proteinaceous pheromones to regulate female mating behavior and receptivity. One of these pheromone components, Plethodontid Modulating Factor (PMF), is a hypervariable protein related to the three-finger protein (TFP) superfamily. Previous studies revealed that PMF persists as a rapidly evolving multi-isoform mixture. However, many characteristics of PMF as a pheromone remained undetermined, including gene structure and transcriptional regulation, translational regulation, protein structure, evolutionary mechanisms, and the isoform effects on …

Inflammation-Resolving Lipid Mediators Promote Revascularization To Enhance Wound Healing., Michael Jingyuan Zhang

Electronic Theses and Dissertations

Wound healing is a highly concerted cellular process that begins with inflammation and proceeds to resolution to revascularize the site of injury. Although inflammation is essential to revascularization during wound healing, it is now recognized that resolution is an active process that is equally important. Other investigations have implicated a beneficial effect of resolving inflammation and promoting resolution in the remission of inflammatory pathologies. Recently investigations have yielded a novel class of ?-3 fatty acid derived lipid mediators, biosynthesized by leukocytes, which are capable of resolving inflammation and promoting resolution. We therefore hypothesized that these leukocyte-derived pro-resolving lipid mediators can …

Epigenetic Modifications Associated With Aortic Remodeling In Hyperhomocysteinemia., Nithya Narayanan

Electronic Theses and Dissertations

Background: Hyperhomocysteinemia (HHcy) is prevalent in hypertensive patients and is an independent risk factor for aortic pathologies. HHcy is known to cause an imbalance between matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) leading to accumulation of collagen in the aorta resulting in stiffness and development of hypertension. Although the exact mechanism of extracellular matrix remodeling (ECM) is unclear, emerging evidence implicates epigenetic regulation involving DNA methylation. The purpose of the study was to investigate whether inhibition of DNA methylation reduces high blood pressure by regulating aortic ECM remodeling in HHcy. Methods: In the first set of experiments, we …

Myocyte-Specific Overexpression Of Stromal Cell-Derived Factor 1 Facilitates Cardiac Regeneration And Improves Myocardial Function After Infarction In Mice., Detlef Ernst-Rudolf Obal

Electronic Theses and Dissertations

Background: Interruption of cardiac stromal cell-derived factor 1 (SDF1)-CXCR4 axis by chronic AMD3100 administration increased myocardial injury after permanent coronary artery ligation, demonstrating the important role of this chemokine in cardiac regeneration. Hypothesis: Cardiomyocyte-specific conditional overexpression of SDF prevents heart failure after permanent coronary ligation and facilitates cardiac regeneration. Methods and Results: Tetracycline-controlled, a-myosine heavy chain promoter directed overexpression of cardiac SDF resulted in a significant increase of SDF expression (SDF: 8.1 ng / g protein) compared with littermate WT mice (0.02 ng / g protein) four weeks after doxycycline withdrawal. SDF overexpression increased AKT and casein kinase 1 levels …

Augmentation Of Ras-Induced Cell Transformation : A New Role For Mir-200a In Malignancy., Lindsey Erin Becker

Electronic Theses and Dissertations

Cancer is a multistep disease that begins with malignant cell transformation and frequently culminates in metastasis and death. MicroRNAs (miRNAs) are small regulatory 21-25-nt RNA molecules and are frequently deregulated in cancer. The majority of miRNAs are estimated to be co-expressed with neighboring miRNAs as clusters. Many miRNA clusters coordinately regulate multiple members of cellular signaling pathways or protein interaction networks. miR-200a is a member of the miR-200 family, which are known to be strong inhibitors of the epithelial to mesenchymal transition. As such, the tumor suppressive role of miR-200a in oncogenesis has been well studied; however, recent studies have …

Determination Of Pore Size Distribution In Capillary-Channeled Polymer (C-Cp) Fiber Stationary Phases By Inverse Size-Exclusion Chromatography (Isec) And The Study Of The Role Of Interstitial Fraction On C-Cp Fibers On Protein Binding Capacity, Zhengxin Wang

All Theses

ABSTRACT High performance liquid chromatography (HPLC), first used in the 1960's, is a rapidly evolving analytical technique, widely employed for identification, separation, and purification in biotechnology and pharmaceutical industries. The development of the stationary phases has played an important role in improving this technique. Each stationary phase will have its own disadvantages. Polysaccharide-based stationary phases such as cross-linked dextran cannot tolerate high pressures and linear velocities; silica stationary phases are rigid enough but slow mass transfer in the pores on the surface causes another problem; with the introduction of non-porous and small bead packing materials, the low surface area and …

The Role Of Stress-Activated Protein Kinases In Parasites, Tamara Kreiss

Theses, Dissertations and Culminating Projects

Lymphatic filariasis and leishmaniasis are neglected tropical diseases that are caused by nematode and protozoal parasites. These diseases cause disfiguration, leaving their host socially marked, and in some cases cause more severe disease that can lead to death. These infections, which tend to persist for long periods of time, also lead to bacterial and fungal co-infections, which further exacerbate the disease. Currently there are insufficient treatment options. Current therapies are often too expensive, have toxicity associated with them and are subjected to growing resistance amongst parasite populations. In this thesis I investigated two potential drug targets. The first, is the …

Synthesis And Characterization Of Co(Ii) And Cu(Ii) Pyrophosphate Coordination Complexes For Treatment Of Pathogenic Agents., Leann Jade Miles

Renée Crown University Honors Thesis Projects - All

Candidiasis, the yeast infection of the mouth, genitals, and blood can be fatal in individuals who are immunocompromised. In humans, Candidiasis is most commonly caused by the yeast, Candida albicans (C. albicans), but can also be caused by other Candida species .^[1]

C. albicans demonstrates marked resistance towards Cu(II) but more sensitivity towards Co(II) compounds, especially in mutants that lack the CaCRP1 and CaCUP1 gene that encodes for the copper-transporting P-type ATPase and Metallothionein proteins, respectively.^[2] Here, I will describe the effects of Co(II) and Cu(II) metal based complexes that incorporate a bridging pyrophosphate …

Microgenomic Approaches To Identify Clinically Relevant Gene Signatures That Discriminate Histologic Types Of Breast Carcinomas., Sean Thomas Butterbaugh

College of Arts & Sciences Senior Theses

Background: Breast cancer presents itself in a variety of histologic types, and the two most common types are invasive ductal carcinoma (IDC) and invasive lobular carcinoma (ILC). Based on comparative genomic hybridization (CGH) analyses, ILC is more closely related to low grade IDC than it is to intermediate and high grade IDC. Results from the BIG 1-98 trial demonstrate that post-menopausal women who are affected with estrogen receptor positive (ER+) ILC or luminal B (high grade) IDC experience a greater magnitude of benefit when they are treated with the aromatase inhibitor (AI) letrozole compared to treatment with the antiestrogen tamoxifen. …

The Anti-Obesogenic Effects Of Nitric Oxide., Brian Edward Sansbury 1985-

Electronic Theses and Dissertations

Obesity is a strong risk factor for developing type 2 diabetes and cardiovascular disease and has quickly reached epidemic proportions with few tangible and safe treatment options. While it is generally accepted that the primary cause of obesity is energy imbalance, i.e., more calories are consumed than are utilized, understanding how caloric balance is regulated has proven a challenge. Molecular processes and pathways that directly regulate energy metabolism represent promising targets for therapy. In particular, nitric oxide (NO) is emerging as a central regulator of energy metabolism and body composition. NO bioavailability is decreased in animal models of obesity and …

Redox Active Motifs In Selenoproteins, Fei Li, Yuliya Pepelyayeva, Elias S. J. Arner, Craig A. Bayse, Sharon Rozovsky

Chemistry & Biochemistry Faculty Publications

Selenoproteins use the rare amino acid selenocysteine (Sec) to act as the first line of defense against oxidants, which are linked to aging, cancer, and neurodegenerative diseases. Many selenoproteins are oxidoreductases in which the reactive Sec is connected to a neighboring Cys and able to form a ring. These Sec-containing redox motifs govern much of the reactivity of selenoproteins. To study their fundamental properties, we have used Se-77 NMR spectroscopy in concert with theoretical calculations to determine the conformational preferences and mobility of representative motifs. This use of Se-77 as a probe enables the direct recording of the properties of …

Quantum Mechanical Study Of Weak Molecular Interactions, Upendra Adhikari

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Molecular interactions play key role in the existence of biomolecules like proteins and nucleic acids, and various materials. Molecular interactions are weak forces that hold different molecules or different fragments of the same molecule together, and are often referred to as noncovalent interactions. Due to the complexity in biomolecules, these interactions are still poorly understood. This dissertation presents results from quantum mechanical simulations of various types of noncovalent interactions, which are extremely important for the structure and functions of biomolecules and materials.

A new sort of noncovalent interaction is identified. This new sort of interaction originates from a direct interaction …

Control Of Cellular Motility By Neuropilin-Mediated Physical Interactions, Xiaobo Li, Matthew W. Parker, Craig W. Vander Kooi

Molecular and Cellular Biochemistry Faculty Publications

The neuropilin (Nrp) family consists of multifunctional cell surface receptors with critical roles in a number of different cell and tissue types. A core aspect of Nrp function is in ligand-dependent cellular migration, where it controls the multistep process of cellular motility through integration of ligand binding and receptor signaling. At a molecular level, the role of Nrp in migration is intimately connected to the control of adhesive interactions and cytoskeletal reorganization. Here, we review the physiological role of Nrp in cellular adhesion and motility in the cardiovascular and nervous systems. We also discuss the emerging pathological role of Nrp …

The Use Of A Small Molecule To Improve The Thermostability Of Dna Junctions, Arik Shams

Honors Theses

The short-term goal of this research project is to employ small molecules as a means to stabilize four-way DNA junctions (4WJs) composed of natural DNA and chimeric nucleic acids. The long-term goal of the project is utilizing the 4WJs as extracellular therapeutic inhibitors of DNA binding proteins [i.e. Histones and High Mobility Group Protein B (HMGB1b)]. A number of studies have shown that classical intracellular DNA-binding proteins have a variety of deleterious side-effects when present in the extracellular milieu. In order to develop a successful 4WJ therapeutic, we are focused on using modified nucleic acids to enhance the stability of …

Cloning, Expression And Interaction Studies Of The Potential Rubisco Activase Cbbq, Salma A. Dawoud

Honors Theses

Carboxysomes are polyhedral microcompartments found in all cyanobacteria and in many chemoautotrophs. Within their shell they contain the enzyme ribulose-1, 5-bisphosphate carboxylase/oxygenase (RubisCO), which fixes CO₂. Downstream of the carboxysome operon there is another gene cluster containing several genes that may enhance carboxysome function. Two of these genes, cbbQ and cbbO, encode potential RubisCO activases. Using recombinant CbbQ and CbbO protein, and RubisCO isolated from carboxysomes the interaction between these proteins was studied. The CbbO and CbbQ proteins were both His tagged, allowing them to be purified with Ni²⁺-NTA column chromatography. Each of these tagged …

Metal Stopping Reagents Facilitate Discontinuous Activity Assays Of The De Novo Purine Biosynthesis Enzyme Pure, Kelly L. Sullivan, Loredana C. Huma, Elwood Mullins, Michael E. Johnson, T. Joseph Kappock

Department of Biochemistry Faculty Publications

The conversion of 5-aminoimidazole ribonucleotide (AIR) to 4-carboxy-AIR (CAIR) represents an unusual divergence in purine biosynthesis: microbes and nonmetazoan eukaryotes use class I PurEs while animals use class II PurEs. Class I PurEs are therefore a potential antimicrobial target; however, no enzyme activity assay is suitable for high throughput screening (HTS). Here we report a simple chemical quench that fixes the PurE substrate/product ratio for 24 h, as assessed by the Bratton-Marshall assay (BMA) for diazotizable amines. The ZnSO₄ stopping reagent is proposed to chelate CAIR, enabling delayed analysis of this acid-labile product by BMA or other HTS methods

Numerical Assessment Of Sequence Conservation In Flu-Virus Hemagglutinin, Scott S. Norton

Honors Scholar Theses

The flu virus was investigated to find a common recognition domain to which an antibody against human-infected viruses can bind. If such a target site is structurally and electrostaticly conserved or invariant, only a single antibody would be required to attack the virus in all cases. The sequence of one of the viral surface proteins contains 24 amino acids that do not vary through mutation. However, these amino acids are neither contiguous in sequence or in space, and the ones that are associated with each other are not readily accessible to an antibody. They do provide a first impression of …

Characterization Of Udp-Arabinopyranose Mutase Genes In The Arabidopsis Cell Wall Mutant Mur5, Christopher A. Hart

Honors Scholar Theses

The genome of Arabidopsis thaliana contains several coding regions for UDP-arabinopyranose mutases (UAMs) that are also known as reversibly glycosylated polypeptides (RGPs). The mur5 cell wall mutant of Arabidopsis shows a 30% decrease in cell wall arabinose content, and a missense mutation in the Reversibly Glycosylated Polypeptide 2 gene was recently proposed to cause this mutant phenotype. Through a traditional complementation analysis, mur5 and a T-DNA insertion mutant in the RGP2 gene were shown not to complement each other, indicating that the two genes are mutant alleles of the same locus. The mur5 SNP located in RGP2 caused a more …

Modeling The Adaptive Immune Response To Mutation-Generated Antigens, Rory J. Geyer

Honors Scholar Theses

Somatic mutations may drive tumorigenesis or lead to new, immunogenic epitopes (neoantigens). The immune system is thought to represses neoplastic growths through the recognition of neoantigens presented only by tumor cells. To study mutations as well as the immune response to mutation-generated antigens, we have created a conditional knockin mouse line with a gene encoding, 5’ to 3’, yellow fluorescent protein (YFP), ovalbumin (which is processed to the immunologically recognizable peptide, SIINFEKL), and cyan fluorescent protein (CFP), or, YFP-ovalbumin-CFP. A frame shift mutation has been created at the 5’ end of the ovalbumin gene, hence YFP should always be expressed, …

Functional Analysis Of Cytosolic Hsp70 Nucleotide Exchange Factor Networks In Yeast, Jennifer Lynn Abrams

Dissertations and Theses (Open Access)

The Hsp70 class of molecular chaperones play critical roles in protein homeostasis via an ATP-dependent folding cycle. Cytosolic Hsp70s in the budding yeast Saccharomyces cerevisiae, Ssa and Ssb, interact with up to three distinct nucleotide exchange factors (NEFs) homologous to human counterparts; Sse1/Sse2/HSP110, Fes1/HspBP1, and Snl1/Bag1. In an effort to understand the differential functional contributions of the cytosolic NEFs to protein homeostasis (“proteostasis”), I carried out comparative genetic, biochemical and cell biological analyses. For these studies, I developed protocols to monitor protein disaggregation and reactivation in a near real-time coupled assay that revealed the importance of aggregate dynamics in the …