Molecular Biology Commons^™

The Yeast Glucose Sensing Receptor Is Stabilized By Interaction With Casein Kinases, Emma E. Mohler

Senior Honors Theses

Many metabolic diseases are rooted in the inability to process glucose or regulate its uptake. These processes can be explored using yeast as a model. Rgt2 is a glucose sensing receptor in yeast, and it detects glucose concentration outside the cell. This receptor is present on the plasma membrane in high glucose conditions but absent in low glucose. Western blot and a yeast two-hybrid assay were used to investigate the relationship between Rgt2 and the plasma membrane-tethered yeast casein kinases, Yck1 and Yck2. This research demonstrated that in high glucose, Rgt2 is stabilized on the plasma membrane by interaction with …

Investigating The Role Of Fun30, A Chromatin Remodeler, In Dna Repair, Mehwish Iqbal

Dissertations

The repair of DNA double-strand breaks (DSBs) is crucial for maintaining genome stability. DSB repair needs to take place within the complex organization of the chromatin, and this requires changes in the chromatin structure adjacent to DSB sites. These changes occur through covalent histone modifications that alter histone-DNA contacts as well as by the action of ATP-dependent chromatin remodelers. Many chromatin remodelers, including Fun30, are involved in DSB repair. Fun30 facilitates DNA end resection at DSB site during the homologous recombination repair pathway. Apart from its role in DNA repair, Fun30 promotes gene silencing at heterochromatic loci such as telomeres, …

Efforts To Increase The Ketamine-Like Activity Of The Rapid-Acting Antidepressant Ro-25-6981 (Mi-4) By Increasing Ampa Potentiation, Nathan Heger

Annual Research Symposium

The small molecule ketamine has generated much interest due to its rapid antidepressant effects. Despite having a rapid onset, ketamine has poor bioavailability, short duration of action, toxicities with long-term use, and a high potential for abuse. The molecule MI-4 (RO 25-6981) has also been shown to have both a rapid and sustained antidepressant effect. Most of the research into the mechanism of the rapid onset of MI-4 and ketamine has focused on their interaction with the NMDA receptor in addition to some monoamine transporters. Some recent publications have shown a significant role of AMPA receptors in the ketamine antidepressant …

Endogenous Cftr Expression In Human Epithelial Cell Lines, Zithlaly Amezquita

Undergraduate Research Conference

CFTR (Cystic Fibrosis Transmembrane-conductance Regulator) is a plasma membrane protein that functions as a chloride ion channel on many epithelial cells. There are over 1000 mutations that affect the function of this protein; however, the most common mutation is DF508. CFTR mutations target many secretory organs like pancreas and lungs causing the genetic disease cystic fibrosis (CF), common among Caucasians of northern European origin. Detection of endogenous CFTR expression is possible by a difficult and costly immunoprecipitation method but not by the most used western blotting method. Therefore, the goal of this project was to detect the endogenous CFTR expression …

Analyzing Inhibitors Of The Sars-Cov-2 Endoribonuclease Nsp15, Mary Staunton

Honors Scholar Theses

The global pandemic caused by the virus SARS-CoV-2 has devastated the world. A flurry of research into the structures and activities of the virus have identified several viable targets for drug therapy, including the endoribonuclease nsp15, also known as EndoU. EndoU has been shown to play a role in diminishing the host cell’s immune response to the virus by cleaving signaling dsRNA, specifically targeting uridine sequences. The development of the crystal structure nsp15 allowed our lab along with others to perform virtual screenings to identify inhibitors that might be able to dock at the active site and inhibit the activity …

The Role Of Eicosanoid Metabolism In Mammalian Wound Healing And Inflammation, Kenneth D. Maus

USF Tampa Graduate Theses and Dissertations

Inflammatory wounds, both chronic and acute, lead to increased morbidity and mortality rates, especially in the elderly population. The annual healthcare cost for chronic wound care alone is over $39B in the US and the demographic of susceptible patients is steadily increasing due to an aging population and lifestyle-related diseases (e.g., hyperlipidemia, obesity, and type 2 diabetes). In fact, many chronic wounds currently have a worse 5-year outlook than certain types of cancers. This shows the need for expediting the wound healing process in such a way that compresses inflammatory signaling and encourages wound resolution without sacrificing pathogen removal and …

Screening For Binding Partners And Protein-Protein Interactions Of A Fungal Transcription Factor- Xdr1, Nishadi Punsara Gallala Gamage

Masters Theses

Clarireedia spp. (formerly Sclerotinia homoeocarpaF.T. Bennett) is the causal agent dollar spot, the most economically important turfgrass disease impacting golf courses in North America. The most effective strategy for dollar spot control is repeated application of multiple classes of fungicides. However, reliance on chemical application has led to resistance to four classes of fungicides as well as multidrug resistance (MDR). Fungi are known to detoxify xenobiotics, like fungicides, through transcriptional regulation of three detoxification phases: modification, conjugation and secretion. Little is known, however, of the protein-protein interactions that facilitate these pathways. Following next-generation RNA sequencing of Clarireedia spp., a …

Characterizing Mechanical Regulation Of Bone Metastatic Breast Cancer Cells, Boyuan Liu

Doctoral Dissertations

Breast cancer most frequently metastasizes to the skeleton. Bone metastatic cancer is incurable and induces wide-spread bone osteolysis, resulting in significant patient morbidity and mortality. Mechanical stimuli in the skeleton are an important microenvironmental parameter that modulates tumor formation, osteolysis, and tumor cell-bone cell signaling, but which mechanical signals are the most beneficial and the corresponding molecular mechanisms are unknown. This work focused on bone matrix deformation and interstitial fluid flow based on their well-known roles in bone remodeling and in primary breast cancer. The goal of our research was to establish a platform that could define the relationship between …

Delineating The Exocytic Fusion Machinery Required For Mast Cell Exocytosis, Pratikshya Adhikari

Dissertations

Mast cells undergo exocytosis to release a wide array of inflammatory mediators by utilizing membrane fusion proteins-SNAREs (soluble-N-ethyl-maleimide sensitive factor attachment protein receptor) along with essential regulatory Munc18 and Munc13 proteins. Accumulating evidence in mast cell biology suggests the existence of distinct pools of mast cell mediators. However, the precise mechanism underlying the release of each mast cell mediator is not clear. To determine whether different exocytic machineries are required for differential mediator release, I used reconstitution to investigate the differential role of Munc18s in fusion machinery regulation. Munc18a and Munc18c stimulated VAMP2 and VAMP3-mediated lipid mixing, whereas Munc18b only …

Role Of Bmi1 In Acute Lung Injury, María Helena Hernández-Cuervo

USF Tampa Graduate Theses and Dissertations

Acute Lung Injury (ALI) is a set of signs and symptoms that lead to acute hypoxemic respiratory failure characterized by bilateral pulmonary infiltrates not attributed to cardiogenic origin. It is caused by a massive innate immune response, with the migration of white blood cells (neutrophils and macrophages principally) and a cytokine storm, followed by alterations in mitochondrial function, increase in reactive oxygen species production, and oxidative stress that in turn induces more mitochondrial damage. Several studies have shown that mitochondrial alterations are key events in the mechanism of ALI and reducing mitochondrial dysfunction could be a possible target in the …

Potential Regulation Of Breast Cancer Invasion By Thymidine Kinase 1, Eliza E. Bitter, Rachel M. Morris, Toni O. Mortimer, Kai Barlow, Abigail Schekall, Michelle H. Townsend, Jonathan Skidmore, Brett E. Pickett, Kim L. O'Neill

Library/Life Sciences Undergraduate Poster Competition 2022

Breast cancer is the most common cancer in women and is largely treatable within the early stages of the disease. However, patient mortality drastically declines as the tumor begins to invade other tissues and metastasize, making aggressive phenotypes especially problematic to treat. Such treatment typically requires an aggressive and decisive multidisciplinary approach. The recent expansion of immunotherapy as a viable treatment option has greatly improved treatment outcomes, especially with aggressive breast cancer phenotypes. Thymidine kinase 1 (TK1) is a DNA salvage pathway enzyme that is highly expressed during S phase and involved in cell cycle repair. Past studies indicated that …

Exendin‐4 Enhances Osteogenic Differentiation Of Adipose Tissue Mesenchymal Stem Cells Through The Receptor Activator Of Nuclear Factor‐Kappa B And Osteoprotegerin Signaling Pathway, Sarah A. Habib, Mohamed M. Kamal, Shohda A. El-Maraghy, Mahmoud A. Senousy

Pharmacy

The capability of mesenchymal stem cells (MSCs) to repair bone damage and defects has long been investigated. The receptor activator of nuclear factorkappa B (RANK), its ligand (RANKL) and the decoy receptor osteoprotegerin (OPG) axis is crucial to keep the equilibrium between osteoblastic and osteoclastic activity. Exendin‐4 utilization increased bone formation and enhanced bone integrity. This study aimed to investigate the mentioned axis and determine the effect of exendin‐4 upon adipose mesenchymal stem cells (Ad‐MSCs) osteogenic differentiation. Ad‐MSCs were isolated from rat epididymal fat, followed by characterization and then differentiation into osteocytes both in the presence or absence of exendin‐4. …

The Role Of Cd5 In Cd4+ T Cell Metabolism, Joshua Bennett, Kiara Whitley, Claudia Freitas Phd, Christopher Haynie, Carlos Moreno, Scott Weber

Library/Life Sciences Undergraduate Poster Competition 2022

T cells serve a key role in the immunological response in the adaptive system. Different receptors present on the T cell have certain roles and are able to either inhibit or stimulate signaling which consequently regulates the function and metabolism of the cell. CD5 is an inhibitory co receptor expressed on the surface of T cells known to regulate thymocyte selection and TCR signaling . Our goal is to better understand the effects of CD5 regulation in metabolism. Using metabolic flux assays, we found that CD5KO naïve T cells have increased glycolysis and mitochondrial respiration in comparison to wild type …

Salicylic Acid And N-Hydroxypipecolic Acid At The Fulcrum Of The Plant Immunity-Growth Equilibrium, Alyssa Shields, Vanessa Shivnauth, Christian Danve M. Castroverde

Biology Faculty Publications

Salicylic acid (SA) and N-hydroxypipecolic acid (NHP) are two central plant immune signals involved in both resistance at local sites of pathogen infection (basal resistance) and at distal uninfected sites after primary infection (systemic acquired resistance). Major discoveries and advances have led to deeper understanding of their biosynthesis and signaling during plant defense responses. In addition to their well-defined roles in immunity, recent research is emerging on their direct mechanistic impacts on plant growth and development. In this review, we will first provide an overview of how SA and NHP regulate local and systemic immune responses in plants. We …

Effect Of Rehmannia Glutinosa Libosch Extract On Proliferation And Cardiogenic Pre-Differentiation Of Human Mesenchymal Stem Cells, Huu Dat Nguyen, Len Ho Thi, Xuan Bach Ho, Van Anh Cao, Duy Minh Le Hoang

BioMedicine

Background: Vietnamese medicine tried and tested certain bioactive compounds from plants to increase the rate of tissue immunomodulation, regeneration, and differentiation. Although there are many research papers discovered about phytochemicals of Rehmannia glutinosa Libosch and differentiation induction potential of some substances purified from this herbal, it finds difficult to seek research that investigated the effect of Rehmannia glutinosa Libosch extract on proliferation and cardiogenic differentiation of mesenchymal stem cells, even though it has commonly been used for a long time because of its function as a restorative and as a critical role in cardiovascular treatment in traditional.

Results: Our …

A Tale Of Two-Hybrids: Investigating The Interactomes Of Cullin-Associated Proteins, Elyse Reitter

Dissertations and Theses

Cul3 is the major component of an E3 ligase in human cells. Cul3 was initially identified in the Singer lab as a protein that binds and degrades cyclin E, and subsequent studies have shown it to be part of a complex that is involved in a multitude of biological functions. The nature of this complex, its constituents, its regulation, and its dynamics is just beginning to be understood. The research presented here utilizes a series of two hybrid screens to identify families of interactomes with Cul3 at the center. This information will complement other work in the lab in which …

Illuminating Transfer Rna Variants As Genetic Modifiers In Models Of Human Disease, Jeremy T. Lant

Electronic Thesis and Dissertation Repository

Transfer RNAs (tRNAs) physically link the genetic code to an amino acid sequence, by recruiting amino acids to three-nucleotide codons in messenger RNAs. To ensure that the genetic code is translated as intended, tRNAs must be accurately aminoacylated and faithfully recognize codons in the ribosome during protein synthesis. Given the critical function of tRNAs, it has often been assumed that mutations in human tRNA genes would be either lethal to cells or not significantly impair tRNA function. My goal was to rigorously test this assumption in mammalian cell models, prompted by the recent discovery of unprecedented variation in human tRNA …

Reversible Lysine Fatty Acylation Of An Anchoring Protein Mediates Adipocyte Adrenergic Signaling., Rushita A Bagchi, Emma L Robinson, Tianjing Hu, Ji Cao, Jun Young Hong, Charles A Tharp, Hanan Qasim, Kathleen M Gavin, Julie Pires Da Silva, Jennifer L Major, Bradley K Mcconnell, Edward Seto, Hening Lin, Timothy A Mckinsey

Biochemistry and Molecular Medicine Faculty Publications

No abstract provided.

Whole-Genome Sequencing And Rna-Seq Reveal Differences In Genetic Mechanism For Flowering Response Between Weedy Rice And Cultivated Rice, Richard S. Garcia, Sapphire Coronejo, Jonathan Concepcion, Prasanta K. Subudhi

Faculty Publications

Flowering is a key agronomic trait that influences adaptation and productivity. Previous studies have indicated the genetic complexity associated with the flowering response in a photoinsensitive weedy rice accession PSRR-1 despite the presence of a photosensitive allele of a key flowering gene Hd1. In this study, we used whole-genome and RNA sequencing data from both cultivated and weedy rice to add further insights. The de novo assembly of unaligned sequences predicted 225 genes, in which 45 were specific to PSRR-1, including two genes associated with flowering. Comparison of the variants in PSRR-1 with the 3K rice genome (RG) dataset identified …

Docking And Molecular Dynamic Of Microalgae Compounds As Potential Inhibitors Of Beta-Lactamase, Roberto Pestana-Nobles, Yani Aranguren-Diaz, Elwi Machado-Sierra, Juvenal Yosa, Nataly J. Galan-Freyle,, Laura X. Sepulveda-Montano, Daniel G. Kuroda, Leonardo C. Pacheco-Londono

Faculty Publications

Bacterial resistance is responsible for a wide variety of health problems, both in children and adults. The persistence of symptoms and infections are mainly treated with beta-lactam antibiotics. The increasing resistance to those antibiotics by bacterial pathogens generated the emergence of extended-spectrum beta-lactamases (ESBLs), an actual public health problem. This is due to rapid mutations of bacteria when exposed to antibiotics. In this case, beta-lactamases are enzymes used by bacteria to hydrolyze the beta-lactam rings present in the antibiotics. Therefore, it was necessary to explore novel molecules as potential beta-lactamases inhibitors to find antibacterial compounds against infection caused by ESBLs. …

Scientific Development Of An Integrated Workflow For Latent Print, Questioned Document, And Dna Processing Of Paper Evidence, Ashley Morgan

Dissertations, Theses, and Capstone Projects

Touch paper evidence could be the source of probative human DNA but recovery is challenging and forensic laboratories instead prioritize processing by the Latent Print and Questioned Document disciplines. Recent advances in DNA collection methods and the increased sensitivity of STR typing kits have improved success rates for DNA testing of paper evidence; but prior to implementing DNA collection, laboratories have to decide in which order to examine paper for the different types of forensic evidence. This thesis developed and tested a multi-discipline workflow for processing paper evidence by DNA, Latent Prints and Questioned Documents experts. Preliminary sampling studies indicated …

Quantitative Imaging Of Tensile Forces At Cell-Cell Junction With Dna-Based Probes, Puspam Keshri

Doctoral Dissertations

Mechanical forces are an integral part in biology, they regulate several cellular properties, such as morphology, proliferation, migration. These forces are also involved in receptor signaling and the differentiation of different cell types. Different proteins and biomolecules such as cadherin, integrin, notch proteins are essential elements of these processes. Measuring these intercellular forces are challenging considering the minimal intensity (piconewton-level) of these molecular forces. In our lab, we have developed a membrane DNA tension probe (MDTP) that uses a DNA hairpin module to sense tensile forces and has a lipid anchor to modify onto live-cell membranes. The programmability of DNA …

Extracellular Secretion Of A Cutinase With Polyester-Degrading Potential By E. Coli Using A Novel Signal Peptide From Amycolatopsis Mediterranei, Yeqi Tan, Gemma K. Kinsella, Gary T. Henehan, Barry J. Ryan

Articles

Recent studies in this laboratory showed that an extracellular cutinase from A. mediterranei (AmCut) was able to degrade the plastics polycaprolactone and polybutylene succinate. Such plastics can be slow to degrade in soils due to a lack of efficient polyester degrading organisms. AmCut also showed potential for the biocatalytic synthesis of esters by reverse hydrolysis.

The gene for AmCut has an upstream leader sequence whose transcript is not present in the purified enzyme. In this study, we show using predictive modelling, that this sequence codes for an N-terminal signal peptide that directs transmembrane expression via the Sec secretion pathway.

E. …

Paddling Along The Voltage Gated Sodium Channel Galaxy With Sea Anemone Toxins: Structural Studies Of The Interaction Between The Paddle Motif From Nav1.5div And Sea Anemone Toxin, Adel K. Hussein

Dissertations, Theses, and Capstone Projects

Voltage gated sodium channels (VGSC) are membrane proteins that serve an important function in the central nervous system (CNS), peripheral nervous system (PNS), and cardiac muscles amongst others. The main function of VGSC is in the propagation of electrical signals by depolarizing excitable cells. Nine mammalian VGSC subtypes have been characterized, Na_V1.1 – Na_V1.9, that are expressed in a tissue specific manner, each with unique gating properties. Numerous diseases have been linked to defects in VGSC including epilepsy, mental retardation, long QT syndrome, and Brugada disease. Furthermore, these channels are one of the primary targets of …

Repurposing Metformin And Antifolates For The Treatment Of Hepatocellular Carcinoma, Sherouk Mohamed Tawfik

Theses and Dissertations

Hepatocellular carcinoma (HCC), one of the most prevalent types of cancers worldwide, continues to maintain high levels of resistance to standard therapy. As clinical data revealed poor response rates, the need for developing new methods has increased to improve the overall wellbeing of patients with HCC. Due to its safety, wide availability and previously reported anti-cancer effects, metformin (MET) serves to be a possible therapeutic agent when combined with other well-known anti-cancer agents. The aim of this study was to investigate the potential anti-cancer effects of MET, an anti-diabetic agent, when combined with two antifolate drugs: trimethoprim (TMP) or methotrexate …

Conformational Flexibility And Local Frustration In The Functional States Of The Sars-Cov-2 Spike B.1.1.7 And B.1.351 Variants: Mutation-Induced Allosteric Modulation Mechanism Of Functional Dynamics And Protein Stability, Gennady M. Verkhivker

Mathematics, Physics, and Computer Science Faculty Articles and Research

Structural and functional studies of the SARS-CoV-2 spike proteins have recently determined distinct functional states of the B.1.1.7 and B.1.351 spike variants, providing a molecular framework for understanding the mechanisms that link the effect of mutations with the enhanced virus infectivity and transmissibility. A detailed dynamic and energetic analysis of these variants was undertaken in the present work to quantify the effects of different mutations on functional conformational changes and stability of the SARS-CoV-2 spike protein. We employed the efficient and accurate coarse-grained (CG) simulations of multiple functional states of the D614G mutant, B.1.1.7 and B.1.351 spike variants to characterize …

Glycocalyx Mechanotransduction Mechanisms Are Involved In Renal Cancer Metastasis, Heriberto Moran, Limary M. Cancel, Peigen Huang, Sylvie Roberge, Tuoye Xu, John M. Tarbell, Lance L. Munn

Publications and Research

Mammalian cells, including cancer cells, are covered by a surface layer containing cell bound proteoglycans, glycoproteins, associated glycosaminoglycans and bound proteins that is commonly referred to as the glycocalyx. Solid tumors also have a dynamic fluid microenvironment with elevated interstitial flow. In the present work we further investigate the hypothesis that interstitial flow is sensed by the tumor glycocalyx leading to activation of cell motility and metastasis. Using a highly metastatic renal carcinoma cell line (SN12L1) and its low metastatic counterpart (SN12C) we demonstrate in vitro that the small molecule Suberoylanilide Hydroxamic Acid (SAHA) inhibits the heparan sulfate synthesis enzyme …

Plasma Induced Reactive Oxygen Species-Dependent Cytotoxicity In Glioblastoma 3d Tumourspheres, Janith Wanigasekara, Carlos Barcia, Patrick J. Cullen, Brijesh Tiwari, James F. Curtin

Articles

The aim of this study was to determine the effects of a pin‐to‐plate cold atmospheric plasma (CAP) on U‐251 MG three‐dimensional (3D) glioblastoma spheroids under different conditions. 3D tumorspheres showed higher resistance to the CAP treatment compared to 2D monolayer cells. A single CAP treatment was able to induce cytotoxicity, while multiple CAP treatments augmented this effect. CAP was also able to induce cytotoxicity throughout the tumoursphere, and we identified that reactive oxygen species(ROS) plays a major role, while H2O2plays a partial role in CAP‐induced cytotoxicity in tumour-spheres. We conclude that ROS‐dependent cytotoxicity is induced uniformly throughout glioblastoma and epidermoid …

Plants, Pipettes And Pcr, Elizabeth Vierling

Science and Engineering Saturday Seminars

Plants are amazing organisms that provide us with food, building materials, the pleasure of gardens, as well as providing the foundation of critical world ecosystems. Although they may look like they are just stuck in one place and doing not more than growing, they have many, many complex ways in which they respond to the environment. The goal of this STEM Ed session will be to discuss ways that plants can respond to the environment with hands on exercises and exploration of possible classroom activities. Participants will engage in state-of-the art methods of testing plant DNA composition using the polymerase …

Establishing A Biochemical System For The Purification And Atpase Activity Of Gst-Dbp5, Sarah R. Utley, Rachel E. Rigsby Phd, Rebecca L. Adams Phd

Science University Research Symposium (SURS)

The export of mRNA out of the nucleus is a crucial step for eukaryotic gene expression. The export of mRNA transcripts is aided by Mex67, which allows export through the nuclear pore complex doorways in the nuclear envelope. Once out of the nucleus, a protein known as Dbp5, bound to ATP, Gle1, and Nup42 aids in the directionality of mRNA export by helping remove Mex67 from the mRNA strand. Following interaction with RNA, Dbp5 then hydrolyzes ATP so that it unbinds the mRNA, allowing for enzyme recycling. Previous efforts worked towards the purification of Dbp5, but the attempts were unsuccessful …