Molecular Biology Commons^™

Immunomodulatory Effects Of Resolvin D2 In A Model Of Infection, Prem Yugandhar Kadiyam Sundarasivarao

Graduate School of Biomedical Sciences Theses and Dissertations

Dysregulated hyperinflammatory host immune response to underlying bacterial infections is a characteristic of sepsis. In sepsis, bacteria often trigger abnormal hyperinflammatory responses which can cause multiple organ failure and if sustained can lead to an immunosuppressive phase where the host is susceptible to secondary infections caused by opportunistic bacteria like Pseudomonas aeruginosa (P. aeruginosa). In our studies, we used a 2-hit model of cecal ligation and puncture (CLP) followed by P. aeruginosa secondary lung infection to investigate cellular and molecular mechanisms in the beneficial action of resolvin D2 (RvD2). Resolvins of the D-series are a group of fatty acids known …

Review Of Biomedical Applications Of Cardiovascular Tissue Engineering, Natalie M. Howard

Honors College Theses

Tissue engineering can be defined as processes that aim to generate three-dimensional functional tissues in vitrothat have been favorably altered according to the structural, biochemical, electrophysiological, and biomechanical properties of the desired tissue before implantation into the human body. In relation to cardiac tissues, these properties would include the ability to conduct action potentials, withstand systolic pressure, permit sufficient O₂ and CO₂penetration, sufficient vascularization to supply nutrients for cellular activity, surface topology that enables cellular communication, and more. As heart diseases and instances of myocardial infarction continue to rise worldwide, there is an increasing need for …

E-Cadherin Force Transmission And Stiffness Sensing, Mazen Mezher

Mechanical & Aerospace Engineering Theses & Dissertations

E-cadherin is the chief mediator of cell-cell adhesion between epithelial cells and is a known mechanosensor. Force transmission and stiffness sensing are two crucial aspects of E-cadherin mechanobiology.

E-cadherin has an extracellular adhesive region, a transmembrane region and an intracellular region that binds to adhesion-associated proteins. Here, we assessed how different factors affect the level of force transmission (i) from inside the cell such as adhesion-associated proteins, (ii) on the cell membrane, such as growth factor receptors and (iii) outside the cell, such as different binding partners in adhesion. To study the level of force transmission inside the cell, we …

Combination Therapy With Trastuzumab And Niraparib: Quantifying Early Proliferative Alterations In Her2+ Breast Cancer Models, Ameer Mansur, Patrick N. Song, Yun Lu, Andrew C. Burns, Luke Sligh, Eddy S. Yang, Anna G. Sorace

Markey Cancer Center Faculty Publications

HER2–targeted treatments have improved survival rates in HER2+ breast cancer patients, yet poor responsiveness remains a major clinical obstacle. Recently, HER2+ breast cancer cells, both resistant and responsive to HER2–targeted therapies, have demonstrated sensitivity to poly–(ADP– ribose) polymerase (PARP) inhibition, independent of DNA repair deficiencies. This study seeks to describe biological factors that precede cell viability changes in response to the combination of trastuzumab and PARP inhibition. Treatment response was evaluated in HER2+ and HER2– breast cancer cells. Further, we evaluated the utility of 3′–Deoxy–3′–[18F]–fluorothymidine positron emission tomography ([18F]FLT–PET) imaging for early response assessment in a HER2+ patient derived xenograft …

Comprehensive Long- And Short- Read Sequencing Approaches To Screen The Mutational Landscape After Therapeutic Crispr-Editing For Duchenne Muscular Dystrophy, Mary Jia, Made Harumi Padmaswari, Landon Burcham, Shilpi Agrawal, Alexis Ivy, Abbey Bryan, Christopher E. Nelson

Biomedical Engineering Undergraduate Honors Theses

Duchenne muscular dystrophy (DMD) is an X-linked muscle wasting disease caused by mutations in the Dmd gene resulting in non-functional dystrophin. Adeno-associated virus (AAV)-mediated delivery of classical CRISPR systems resulting in double-stranded breaks (DSBs) has yielded promise as a curative DMD therapeutic. However, the consequences of AAV-CRISPR require more thorough characterization. PCR-enriched short-read amplicon sequencing at cut sites is the most common method of sequencing CRISPR editing, but PCR bias results in unreliable quantification of on-target edits. We analyze orthogonal long- and short-read sequencing approaches to screen the outcomes of CRISPR technologies at the DNA and RNA-level for DMD. We …

Methyltransferase, Glucose Adaptation, And Import Complex In Trypanosoma Brucei, Emily Knight

All Dissertations

Trypanosoma brucei is a kinetoplastid parasite responsible for human African trypanosomiasis (HAT) and nagana, a livestock wasting disease, which both endemic to sub-Saharan Africa. Unique to kinetoplastids are the specialized peroxisomes, named glycosomes, which compartmentalize the first several steps of glycolysis and gluconeogenesis, nucleotide sugar biosynthesis, and many other metabolic processes. Kinetoplastids are unique in that they have a single mitochondrion. In this work, I present the first study into SET domain proteins in any kinetoplastid parasites. We have characterized a predicted SET domain protein, TbSETD3, that localizes to the mitochondrion and a depletion of the protein results in growth …

Minimal Carbon Requirements For Potential Colonizers Of Other Planets, Benjamin Tan

Biological Sciences Undergraduate Honors Theses

The NASA Office of Planetary Protection regulates the safe scientific exploration of other planets. Specifically, the office enacts rules to discourage interplanetary mission practices that would lead to the contamination of Earth-originating microbial life on other planets. Interplanetary contamination jeopardizes the potential to obtain reliable scientific evidence for extraterrestrial life. In coordination with this office, the biosignatures of potential colonizers of other celestials bodies are studied. Several organisms of Earth qualify as potential colonizers of other planets.

This experiment focused on the environment of Mars in particular. Two organisms were tested: Desulfovibrio arcticus and Desulfotalea psychrophila. Both are psychrotolerant or …

Spatial Metabolomics Reveals Glycogen As An Actionable Target For Pulmonary Fibrosis, Lindsey R. Conroy, Harrison A. Clarke, Derek B. Allison, Samuel Santos Valenca, Qi Sun, Tara R. Hawkinson, Lyndsay E. A. Young, Juanita E. Ferreira, Autumn V. Hammonds, Jaclyn B. Dunne, Robert J. Mcdonald, Kimberly J. Absher, Brittany Dong, Ronald C. Bruntz, Kia H. Markussen, Jelena A. Juras, Warren J. Alilain, Jinze Liu, Matthew S. Gentry, Peggi M. Angel, Christopher M. Waters, Ramon C. Sun

Saha Cardiovascular Research Center Faculty Publications

Matrix assisted laser desorption/ionization imaging has greatly improved our understanding of spatial biology, however a robust bioinformatic pipeline for data analysis is lacking. Here, we demonstrate the application of high- dimensionality reduction/spatial clustering and histopathological annotation of matrix assisted laser desorption/ionization imaging datasets to assess tissue metabolic heterogeneity in human lung diseases. Using metabolic features identified from this pipeline, we hypothesize that metabolic channeling between glycogen and N-linked glycans is a critical metabolic process favoring pulmonary fibrosis progression. To test our hypothesis, we induced pulmonary fibrosis in two different mouse models with lysosomal glycogen utilization deficiency. Both mouse models displayed …

Modeling Accuracy Matters: Aligning Molecular Dynamics With 2d Nmr Derived Noe Restraints, Milan Patel

Honors Scholar Theses

Among structural biology techniques, Nuclear Magnetic Resonance (NMR) provides a holistic view of structure that is close to protein structure in situ. Namely, NMR imaging allows for the solution state of the protein to be observed, derived from Nuclear Overhauser Effect restraints (NOEs). NOEs are a distance range in which hydrogen pairs are observed to stay within range of, and therefore experimental data which computational models can be compared against. To that end, we investigated the effects of adding the NOE restraints as distance restraints in Molecular Dynamics (MD) simulations on the 24 residue HP24stab derived villin headpiece subdomain to …

Targeting Metabolic Alterations Associated With Smooth Muscle Α-Actin Pathogenic Variant Attenuates Moyamoya-Like Cerebrovascular Disease, Anita Kaw

Dissertations and Theses (Open Access)

Heterozygous pathogenic variants in ACTA2, encoding smooth muscle α-actin (α-SMA), predispose to thoracic aortic aneurysms and dissections. De novo missense variants disrupting ACTA2 arginine 179 (p.Arg179) cause a multisystemic disease termed smooth muscle dysfunction syndrome (SMDS), which is characterized by early onset thoracic aortic disease and moyamoya disease-like (MMD) cerebrovascular disease. The MMD-like cerebrovascular disease in SMDS patients is marked by bilateral steno-occlusive lesions in the distal internal carotid arteries (ICAs) and their branches. To study the molecular mechanisms that underlie the ACTA2 p.Arg179 variants, a smooth muscle-specific Cre-lox knock-in mouse model of the heterozygous Acta2 R179C variant, termed …

Regulation And Function Of Zeb1 Acetylation In Lung Adenocarcinoma Progression And Metastasis, Mabel Perez-Oquendo

Dissertations and Theses (Open Access)

Lung cancer metastasis is leading the causes of cancer-related mortality in the United States and worldwide. Epithelial-to-mesenchymal transition (EMT) is a model for metastasis that results in loss of specialized epithelial cell contacts and acquisition of mesenchymal invasive capacity. Zinc finger E-box-binding homeobox 1 (ZEB1) recognizes and binds to E-boxes of epithelial gene promoters to repress its transcription. ZEB1 has inconsistent molecular weights, which have been attributed to post-translational modifications (PTMs). In the presented dissertation, I specifically addressed the gap in the molecular mechanisms by which PTMs of ZEB1 regulate its ability to induce EMT and how its activity might …

Neuronal Or Intestinal Knockdown Of C. Elegans Nadk-1 Decreases Oxygen Consumption And Reactive Oxygen Species, Jake Regan

Electronic Theses and Dissertations

Reactive oxygen species (ROS) such as H2O2 can damage cellular components and are formed as a byproduct of mitochondrial oxidative metabolism. Studies using the nematode C. elegans have found that increasing ROS during development or early adulthood can extend lifespan, while increasing ROS during later adulthood normally decreases lifespan. NADPH provides the reducing power for several cellular antioxidants and is synthesized in a two-step reaction from NAD+ with the first step being catalyzed by NAD kinase (NADK). In this study, the effects of knocking down C. elegans cytoplasmic NADK, nadk-1 globally or in a neuron or intestine-specific manner starting from …

Cyclic-Di-Gmp Regulates Salmonella Typhimurium Infection Of Epithelial Cells And Macrophages, Abdulafiz Musa

Electronic Theses and Dissertations

Regulation of the bacterial second messenger cyclic-di-GMP in Salmonella Typhimurium allows it to delicately alter phenotypes to optimize invasion and survive intracellularly in epithelial cells and macrophages to become virulent and cause infection. Cyclic-di-GMP concentration is regulated by the presence of external stimuli, sensory diguanylate cyclases (DGCs) and phosphodiesterases (PDEs), and cyclic-di-GMP binding effectors. Previous studies established that maintenance of low cyclic-di-GMP concentrations is required for survival in macrophages, and that deletion of 3 active PDEs reduces this survival. Here I showed that these 3 PDEs also influenced the infection of epithelial cells. Further studies re-established the decreased survival in …

New Dna Repair And Demethylation Functions In Uracil Dna Glycosylase Superfamily, Chenyan Chang

All Dissertations

Uracil-DNA glycosylase (UDG) superfamily, which consists of several groups of enzymes that recognize the damaged DNA bases and initiate the base excision repair (BER) pathway, is most important in dealing with DNA deamination and other base modifications. Thymine DNA glycosylase (TDG), which belongs to family 2 in the UDG superfamily, is able to specifically recognize and cleave the 5-methylcytosine (mC) oxidative derivatives including 5-formylcytosine (fC), 5-carboxylcytosine (caC), 5-hydromethyluracil (hmU) caused by active demethylation or DNA damage. My dissertation work is mainly focused on the fC and caC glycosylase activity within UDG superfamily. Chapter 1 is a general introduction to the …

Acetate Metabolism In The Fungal Pathogen Cryptococcus Neoformans, Oly Ahmed

All Dissertations

Cryptococcus neoformans is an environmental basidiomycetous fungus with a worldwide distribution and a wide range of habitats. Inhalation of the desiccated yeasts or spores of C. neoformans often leads to opportunistic pulmonary infections in immunocompromised individuals, and in severe cases causes lethal meningitis following hematogenous dissemination. During infection, depending on the tissue and disease state, the invading fungi experience a range of nutrient microenvironments within the host body. As a result, rapid metabolic adaptations geared towards efficient utilization of carbon sources alternative to glucose become one of the prime determinants of survival and growth for the pathogen. Incidentally, cryptococcal infection …

Effects Of Cations And Dipicolinic Acid On B. Anthracis Spore Physiology And Cytotoxicity, Chandler P. Hassan

UNLV Theses, Dissertations, Professional Papers, and Capstones

Inhalation of Bacillus anthracis spores leads to the most severe form of anthrax. Following phagocytosis by alveolar macrophages, spores germinate inside the phagolysosome. The mechanism used by the newly germinated B. anthracis cells to survive within macrophages is not completely understood.B. anthracis spores contain large concentrations of calcium in complex with dipicolinic acid (Ca-DPA). Upon germination, the cell excretes the large depot of Ca-DPA. DPA is an amphipathic molecule that could act as a buffer and modulate phagolysosome acidification. Intracellular calcium overload can disrupt signaling pathways required for normal macrophage function and trigger cell death. We hypothesized the release of …

Long-Range Regulation By The Shigella Virulence Gene Regulator, Virb, Cody Cris

UNLV Theses, Dissertations, Professional Papers, and Capstones

Nucleoid-structuring proteins (NSPs) in bacterial cells can exert gene regulatory effects over great distances on DNA. This contrasts with the traditional view of bacterial transcriptional regulation, where transcription factors bind cis-acting regulatory sites proximal (<250 bp) to promoters to influence gene expression. In Shigella species, the histone-like nucleoid-structuring protein (H-NS) transcriptionally silences many genes on the large (~220 kb) virulence plasmid. During host colonization, however, silencing by H-NS is alleviated by the anti-silencer protein, VirB, which remodels the H-NS:DNA complex. This transcriptional anti-silencing is mediated by VirB:DNA interactions and the modulation of DNA supercoiling, which lead to the up-regulation of virulence genes. VirB is a central regulator of Shigella virulence, but the mechanistic basis for transcriptional regulation is incompletely characterized. Here, I focus on the potential for VirB to regulate transcription from remotely located DNA binding sites. Employing complementary in vivo approaches, I assessed the spatial stringency of the VirB binding site and elucidated how varying cellular VirB levels impacts regulation over different distances. I also evaluated how a molecular roadblock on DNA influences remote regulation by VirB. Additionally, I developed control tools to validate the ability of VirB to spread along DNA, an activity of VirB that putatively drives transcriptional regulation. These data are consistent with transcriptional anti-silencing being driven by VirB spreading along DNA. Overall, these findings highlight how VirB is not a conventional transcriptional regulator and broaden our perspective on transcriptional control in bacteria. Furthermore, this work contributes to our understanding of long-range gene regulatory effects, which are frequently overlooked and poorly characterized in bacteria.

Gonads Without Glp-1: Silencing Glp-1 In The Male Somatic Gonad In Caenorhabditis Elegans, Matthew Titus

Undergraduate Theses

In C. elegans, the gene glp-1 encodes for a Notch receptor called GLP-1, one of two found in C. elegans’ genome. The gene has been previously implicated in the development of the hermaphroditic germline as well as playing a role in the mitosis/meiosis decision. Genetic screening has further identified it as potentially playing a role in the development of the male somatic gonad, making it an ideal candidate for a reverse genetic. We did this by silencing glp-1 and observing if any alterations to the gonad’s phenotype occur.

Normally this could be done by performing a gene knockout. …

Role Of Cdx4 And Sp5l In Zebrafish Development, Wesley Tsai

Honors Theses

The Caudal Type Homeobox transcription factors cdx are a family of genes found in vertebrates that regulates body regionalization and anterior-posterior patterning. They are also responsible for regulating axial elongation, but the mechanisms behind this behavior are not known. Previous studies in mouse embryonic stem cells have shown that the cdx genes are necessary for upregulating the gene sp5 which may be linked to axial elongation. Sp5 is a zinc-finger transcription factor belonging to the specificity protein (sp) family. Our group has used in-situ hybridization experiments on zebrafish embryos to show that sp5-like (sp5l) is transcribed within tailbud tissues that …

Bryophytes Of Goochland County, Virginia, Mikayla Quinn

Honors Theses

Bryophytes are non-vascular land plants that include mosses, liverworts, and hornworts. Although easier to overlook because of their smaller size, bryophytes are a fundamental part of the ecosystem. As such, maintaining record of their biodiversity is important. Yet, records of bryophyte species in Goochland County, VA were low compared to more thoroughly documented counties such as Prince Edward County. This study expands the documentation of bryophyte flora and presents a checklist of bryophyte species found Virginia’s Goochland County from 2020-2023. Fieldwork conducted at public and privately-owned properties throughout the county between January 2020 and March 2023 yielded 702 specimens that …

Potential Use Of The Gel Extract Of Butterfly Pea Flower As Topical Therapy To Prevent Photodamage By Downregulating Tnf-Α And Caspase-3 Expression Levels In Uvb-Exposed Rats, Elvana Cahyani, Agung Putra, Prasetyowati Subchan

Makara Journal of Health Research

Background: Prolonged exposure to UVB radiation causes DNA damage in skin cells by raising the levels of reactive oxygen species, resulting in the production of inflammatory factors and skin issues. Plant extracts are frequently used to counteract photodamage due to their antioxidant properties. One example is the floral extract of the butterfly pea plant, which contains flavonoid antioxidants. However, the effect of the extract on inflammatory factors is unknown. This study investigated how tumor necrosis factor-alpha (TNF-α) and caspase-3 expression changed when a butterfly pea flower extract gel was applied topically to UVB-exposed animals.

Methods: Experimental and control …

Investigating The Helicase Activity Of Methylated Vs Unmethylated Ded1, Hannah Lukow

Honors Theses

Ded1 is an RNA helicase protein of the DEAD-box subfamily in eukaryotic organisms (Sharma & Jankowsky, 2014) which can act as an activator or initiation factor, during translation (Hilliker et al., 2011). Ded1 has several functions in yeast including assembly of translational initiation factors, scanning the mRNA for the start codon, and unwinding any double stranded segments of mRNA with its helicase ability. Ded1 was discovered to be methylated at four arginine sites in vivo (Low et al., 2013), with a fifth methylation site being discovered recently (Low et al., 2020), however the purpose of such post-translational modifications is still …

The Role Of Cerium(Iii) In Bacterial Growth And The Microbial Transformation Of Aromatic Hydrocarbons, Shruti Sathish

Honors Theses

Biofilms are communities of surface-attached bacterial cells encased in an exopolymeric matrix. In this state, they are more resistant to antimicrobial treatment and can have adverse effects in medical, agricultural, and industrial settings. Whereas, as biocatalysts, biofilms from nonpathogenic bacteria enhance their performance and stability in catalysis. Unfortunately, there are several challenges when using bacteria in organic transformations due to their complex cellular chemistry. Trivalent lanthanide metals were discovered to serve regulatory roles in some bacterial catalytic processes, including those of Pseudomonas putida KT2440 (P. putida), a non-infectious Gram-negative bacterium. The main goal of our research is to use cerium(III) …

Testing Of Indazole Inhibitors Of Kasa, A Vital Enzyme Of M. Tuberculosis, Karissa Highlander

Departmental Honors & Graduate Capstone Projects

Tuberculosis is a disease that affects the lungs caused by Mycobacterium tuberculosis (M. tuberculosis). Although drug treatment options exist, increased rates of antibiotic resistant strains have become more prevalent in recent years, driving a need for new treatment approaches. KasA, a β-ketoacyl synthase, has been found to synthesize parts of the cell wall and been identified as an attractive drug target. Previous medicinal chemistry research has been completed to synthesize six effective competitive inhibitors of KasA that would potentially block the enzyme from binding the substrate, preventing elongation of the backbone and creation of the mycolic fatty acids that …

Functional Domains Of The Hsv-1 Ul37 Protein In Virus Infectivity And Retrograde Transport In Neurons, Carolyn Clark

LSU Doctoral Dissertations

Herpes simplex virus type 1 is large alphaherpesvirus with 20 envelope proteins and 23 tegument proteins. Virus entry is via fusion of the viral envelope with cellular membranes mediated by a complex of viral glycoproteins. Glycoprotein B (gB) is the only fusogenic glycoprotein, while other viral glycoproteins and membrane proteins including gD, gH, gL, gK and UL20 membrane protein modulate gB’s fusogenicity. Previously, Our laboratory developed the HSV-1 (VC2) vaccines strain with partial deletions in the amino termini of gK (ΔgK(31–68)) and the UL20 membrane protein (ΔUL20(4–22)). VC2 enters epithelial cells only via endocytosis followed by release of the capsid …

A Potential Role Of Gsk-3beta In The Development Of Pain Post-Spinal Cord Injury, Emily Ernst

Undergraduate Theses

Chronic neuropathic pain is a very common consequence following spinal cord injury (SCI) and poses significant clinical challenge. Current treatments are largely ineffective and as a result, patients are often left with debilitating pain at and below the level of the spinal cord injury. After spinal injury, a cascade of events occurs within the nervous system. One important aspect of this cascade for investigation is the anatomical changes that occur post-SCI. There is evidence of maladaptive sprouting/growth in sensory nerve fibers that is thought to play a role in the development and amplification of pain signals. Sprouting of primary afferents …

Novel Plant Peroxidase Investigation For Use In Various Biochemical And Practical Applications, Sally Hess

Liberty University Research Week

Undergraduate

Basic

Characterization Of A Sas3 Homolog In The Human Pathogenic Yeast Cryptococcus Neoformans, Lillianne Bartone

Liberty University Research Week

Undergraduate

Basic

Characterization Of Select Lysine Mutations Of The Cystine/Glutamate Transporter, System XC-, Anna Koppin, Claire Buck, Amanda Gibson

22nd Annual Celebration of Undergraduate Research and Creative Activity (2023)

System xc- is a membrane transport system that plays a critical role in mitigating oxidative stress. As such, its regulation is critical for proper brain functioning. Recent work in our lab has shown that System xc- activity increases immediately during an oxidative insult by undergoing a change in localization to the plasma membrane, but we have yet to identify the specific mechanism for the redistribution of the transporter. Previous studies have demonstrated that post-translational modifications of proteins can lead to differential protein distribution within cells. Therefore, in this study, we sought to determine if post-translational modification (PTM) of the transporter …

Does Phosphorylation On Serine 26 Of System Xc- Lead To Changes In Cell Surface Expression?, Katherine Lane

22nd Annual Celebration of Undergraduate Research and Creative Activity (2023)

System x_c^- is involved in transporting cystine into cells and glutamate out of cells, and ultimately in production of the antioxidant glutathione. Antioxidants are important in protecting cells from oxidative stress which can occur when waste products like H₂O₂ build up in the cell. Previous studies have shown that a critical regulator mTORC regulates system xc- by phosphorylating serine 26 (S26) on the N-terminus of the cystine/glutamate antiporter xCT, leading to a reduction in transport activity. The specific objective of this study is to determine the mechanism by which phosphorylation of S26 affects activity. To …