Molecular Biology Commons^™

Creation Of A Dual-Fluorescence Reporter System To Assay Mirna Regulation Of Apobec3a, Tolulope D. Ajayi

All Graduate Theses, Dissertations, and Other Capstone Projects

MicroRNAs have an important role in regulating gene expression, including regulating the transcripts of the APOBEC3 family of cytidine deaminases. The APOBEC3 family is part of the immune defense against viral infection but has also been implicated in cancer progression. The hypothesis for this thesis is that specific miRNAs could be identified that regulate APOBEC3A (A3A) expression. I used molecular cloning and confocal microscopy to evaluate the miRNA-mediated silencing of A3A. A commercially available vector, psiCHECK-2, can be used to assay microRNA (miRNA) activity on specific targets. Molecular cloning was performed to modify the psiCHECK-2 Dual luciferase system (Renilla and …

Investigating The Role Of Fnci-1 In The Development And Function Of The C. Elegans Nervous System, Meagan Collins

Open Access Master's Theses

Fanconi Anemia (FA) is a rare human genetic disease caused by mutation of any of twenty-three genes on the FA pathway, which functions in the repair of DNA damage and the maintenance of genome stability. FA is clinically characterized by congenital abnormalities and a predisposition to aplastic anemia and malignancies. Recently, progressive neurological symptoms and abnormal neurological findings on imaging have been noted in FA patients, including cerebral and cerebellar lesions, calcification, necrosis, gliosis, and associated acute and chronic neurological symptoms; this constellation of symptoms has been termed Fanconi Anemia-Associated Neurological Syndrome (FANS).

To elucidate the molecular etiology of FANS, …

Investigating Regulation And The Regulome Of The Nucleic Acid Binding Protein Spovg Within Borrelia Burgdorferi, Timothy C. Saylor

Theses and Dissertations--Medical Sciences

Prior studies indicated Borrelia burgdorferi SpoVG protein was regulated, SpoVG binds specifically to a number of nucleic acids, and SpoVG binds to the non-variable section of the major virulence factor vlsE. Although a direct transcriptional activator or inhibitor was not elucidated, it is shown that SpoVG is regulated based on cell culture density independent of the growth rate of B. burgdorferi, indicating that SpoVG regulation is outside the scope of the alternative sigma factor RpoS. Using a dCas9 CRISPRi construct to target SpoVG resulted in protein expression trending towards upregulation of the BosR-RpoS regulon, and a marked decrease …

Exploring The Interconnections Between Mitochondrial Carrier Proteins Involved In Energy Transfer: The Case Of Uncouplig Proteins 2 And 4, Adenine Nucleotide Translocase, And Phosphate Translocase, Marzieh Tabefam

Theses and Dissertations (Comprehensive)

Mitochondrial carrier (MC) proteins play an essential role in cellular energy metabolism by mediating the transport of solutes and protons across the mitochondrial inner membrane (MIM). This thesis explores the structural and functional characteristics of three groups of MC proteins—Uncoupling Proteins (UCPs), Adenine Nucleotide Translocase (ANT), and Phosphate Translocase (P_iT)—to elucidate their individual and interrelated cooperative roles in mitochondrial proton transport and energy regulation.

The study develops and applies novel methodologies for the overexpression, purification, and reconstitution of MC proteins into model lipid bilayers mimicking the native mitochondrial inner membrane environment. These methods ensure proteins maintain their native-like …

Reversible Degradation Of Peptidoglycan By Lytic Transglycosylases, Aaron Devereaux

Theses and Dissertations (Comprehensive)

Antimicrobial resistance continues to be a burden on the global healthcare system with an estimated cost of billions of dollars and millions of deaths each year. Recently, there has been little to no development on new antibiotics to treat bacterial infection, and any that are developed become resisted to within a few years. Both Gram-positive and Gram-negative bacteria contain a mesh-like layer called peptidoglycan (PG) that surrounds their cells providing strength, cell shape, and protection from their environments. This layer is a polymer of N-acetylmuramic acid (MurNAc) and N-acetylglucosamine (GlcNAc) connected via a b-1,4-glycosidic bond, with each strand being cross-linked …

Structural Insights Into Peptidoglycan O-Acetylation In Gram-Negative Bacteria, Tyler Malloch

Theses and Dissertations (Comprehensive)

The Centres for Disease Control has acknowledged that we are entering a post-antibiotic era, where increasing numbers of bacterial pathogens are becoming resistant to known antibiotics. One of the most successfully targeted features of bacteria is also the main component of the cell wall, peptidoglycan (PG). PG is a heteropolymer of alternating sugar residues, namely N-acetylmuramic acid (MurNAc) and N-acetylglucosamine (GlcNAc) which are joined together by b-(1,4)-glycosidic bonds and crosslinked via stem peptide chains. PG is a target, readily lysed by exogenous and endogenous autolysins, such as lysozyme and lytic transglycosylase, respectively. This event leads to cell death …

Characterizing The Targeting Of Toc159 Receptors To The Chloroplast Outer Envelope Membrane And Their Role In The Assembly Of Distinct Toc Complexes, Michael Fish

Theses and Dissertations (Comprehensive)

Global agriculture and food security face challenges from climate change, population growth, and other human-driven pressures. Enhancing our understanding of plant biology, particularly chloroplast function, is critical for improving crop yield and stress resilience. As the sites of photosynthesis and key regulators of plant responses to environmental cues, chloroplasts depend on the precise import of thousands of nuclear-encoded proteins to maintain their biogenesis and function. While much is known about the core translocon components in the chloroplast outer envelope membrane that form the TOC complex, the mechanisms underlying their targeting, assembly, and regulation remain incompletely understood.

Plastids, including chloroplasts, evolved …

Discovery Of Photosynthetic Oxic N2-Fixation In Cyanobacteria Using Wet Lab And Machine Learning Approaches, James A. Young Iii

Electronic Theses and Dissertations

No abstract provided.

Characterization Of The Erv41-Erv46 Complex As A Retrograde Receptor For Misfolded Secretory Proteins, John Andrew Fuesler

Dartmouth College Ph.D Dissertations

The Endoplasmic Reticulum (ER), the site of secretory protein biosynthesis

provides a favorable environment to promote polypeptide folding, protein

oligomerization and export. Although the ER contains many chaperones and other

factors that assist in protein folding, this process is error prone. Eukaryotes have

evolved protein quality control checkpoints to maintain homeostasis in the secretory

pathway, however these processes are imperfect and may result in proteotoxicity

when toxic levels or aggregates of proteins arise. Mutations in proteins involved in

trafficking in the early secretory pathway are closely associated with disease states.

Therefore, it is of great interest to further understand how …

Molecular Interactions Between The Shellfish Pathogen Vibrio Coralliilyticus Re22 And The Probiont Phaeobacter Inhibens S4sm, Jessica Coppersmith

Open Access Dissertations

Oyster larvae aquaculture is an important industry, both economically and environmentally, helping to create sustainable food systems by supplying seed oysters to farms, and also to restoration projects aimed at building coastal resilience and improving water quality by rejuvenating wild populations. However, oyster larvae are highly susceptible to rapid mortality events, crashes, which can seriously impact the industry. One source of these crashes in hatchery systems is vibriosis, caused by pathogenic species of Vibrio bacteria that are ubiquitous in temperate coastal waters, and with rising surface water temperatures their prevalence and virulence are increasing. To prevent these larval crashes from …

Optimization Of L-Citrulline Production In Corynebacterium Glutamicum, And The Effect Of The Human Small Intestine Environment, Simran S. Ghulldu

Honors Undergraduate Theses

L-citrulline is a crucial amino acid involved in nitric oxide production, which supports cardiovascular health, physical performance, and intestinal well-being. Nitric oxide deficiency is linked to conditions like hypertension, highlighting the need for efficient L-citrulline production. This study attempts to enhance L-citrulline synthesis in Corynebacterium glutamicum, a bacterium widely used in industrial amino acid production, as well as test a The hypothesis is that overexpressing carAB, which encodes carbamoyl phosphate synthetase, will increase carbamoyl phosphate levels and boost L-citrulline production. Two C. glutamicum strains were used: a wild type and the CRBG strain, which has mutations that promote …

Microbial And Plant Biodiversity In Appalachian Agroecosystems And Their Implications For Soil Health, Kinsey Reed Close

Graduate Theses, Dissertations, and Problem Reports (ETD)

The sustainability of agricultural landscapes depends on understanding the complex interactions between plants, soil, and microbes. In Appalachia, a region shaped by complex terrain and long histories of human and ecological interaction, small family farms produce hay, livestock, and row crops. These agroecosystems, though often treated as “artificial” by many ecologists, have the potential to support both food production and biodiversity conservation if managed with an understanding of ecological complexity. This dissertation examines how plant and microbial diversity interact with soil and management to influence nutrient cycling, soil health, and long-term resilience in Appalachian agricultural systems. A literature review revealed …

Using Lateral Flow Assay To Assess The Prevalence Of Trachemys Hybridization, Lillian Groves

Honors Program Theses

The current study focuses on Trachemys species, which is a group of freshwater turtles found throughout the Greater Antilles. The role of studying an isolated island, such as the Dominican Republic, is significant because it serves as a way to better understand how the combined effects of species migration, mass tourism, and climate change can greatly impact isolated species. Of particular significance is how these forces interact in complex ways to reshape habitats and threaten biodiversity. Additionally, the introduction of a highly invasive species, Trachemys scripta elegans (T. scripta), in this environment pressures native species by acting as a source …

Probing The 10-23 Dnazyme Catalytic Core: Insights From Structural And Mechanistic Studies, Evan Rhyse Cramer

Graduate Theses, Dissertations, and Problem Reports (ETD)

The 10-23 DNAzyme is a catalytically active DNA oligonucleotide capable of cleaving RNA at purine-pyrimidine junctions in the presence of divalent metal ions, most notably magnesium. Despite its promise as a programmable RNA knockdown tool, its clinical and biotechnical utility has been hampered by limited structural characterization, low catalytic efficiency under physiological conditions, and poor understanding of its in-cell activity. This dissertation presents a comprehensive framework for the structural and functional analysis of DNAzymes using the 10-23 DNAzyme as a model system. In the absence of high-resolution structures, we first employed detailed in vitro biochemical assays to map essential residues …

Caspases Protect Against Hyperglycemia Induced Necroptosis-Like Cell Death In Brain Endothelial Cells, Aleena S. Mathew

Theses and Dissertations

Acute diabetes characterized by high spikes in blood glucose levels in association with severe hyperglycemia can lead to an increased risk of ischemic stroke in the brain. To better understand high blood glucose levels, hyperglycemia, in the brain we explored its effects on endothelial cells. Endothelial cells line the inner walls of blood vessels and are directly impacted in hyperglycemia. Brain endothelial cells are a major component of the blood brain barrier (BBB) which tightly regulates the brain’s microenvironment. The type of cell death such as apoptosis, caspase dependent, or necroptosis, caspase independent, can exacerbate diabetes associated complications. There is …

Characterization Of Dietary And Hormonal Influences On Renal Sodium Transporters And Channels In Rodent Models, Evelyn Chan

Scripps Senior Theses

Within kidneys, a diverse array of renal epithelial cells works to maintain electrolyte and water balance to regulate blood pressure and pH. These cells contain varying types of sodium transporters that adjust sodium reabsorption according to the body’s needs. Previous studies have observed sex-specific differences in renal sodium transporter regulation, which are thought to optimize reproduction in females. In this study, we performed high resolution microscopy and detailed characterization of cell type-specific expression and localization of major sodium transporters and channels in rodents in response to dietary and hormonal influences. Specifically, we examined the impact of dietary salt intake and …

The Role Of Fruiting Bodies In Protecting A Community Of Myxococcus Xanthus Spores From The Environment, Dave Lall

Wayne State University Dissertations

Environmental microorganisms have evolved a variety of strategies to survive fluctuations in environmental conditions, including the production of biofilms and differentiation into spores. Myxococcus xanthus are ubiquitous soil bacteria that produce starvation-induced multicellular fruiting bodies filled with environmentally resistant spores (a specialized biofilm). Isolated spores have been shown to be more resistant than vegetative cells to heat, ultraviolet radiation, and desiccation. The evolutionary advantage of producing spores inside of fruiting bodies is not clear. Here, we examine a hypothesis that the fruiting body provides additional protection from environmental insults. We developed a high-throughput method to compare the recovery (outgrowth) of …

Examining The Biotic And Abiotic Degradation Of Simple Phenols, Madeline Gruenberg

Browse all Theses and Dissertations

Plant secondary metabolites are a diverse class of compounds that encompass small phenols to large polymeric structures such as tannins. Smaller compounds such as phenolic acids can serve as substrates for soil microbes and enzymes. We monitored the degradation of three small phenols pyrogallol, gallic acid, and benzoic acid in biotic and abiotic conditions. They were chosen to observe the impacts of specific functional groups on both degradation processes and act as model compounds to mimic larger tannin reactivity. Both abiotic and biotic degradation of pyrogallol resulted in the formation of a stable quinone, while abiotic degradation of gallic acid …

The Role Of Fbxl16 In Regulating Erk3 Stability And Cell Migration, Seth Charles Murdock

Browse all Theses and Dissertations

Extracellular signal-regulated kinase 3 (ERK3) is a member of the atypical mitogen-activated protein kinase (MAPK) family. While ERK3’s role in promoting cell migration and invasion in multiple types of cancer has been well-documented, its activation and regulation remain poorly understood. Unlike other typical MAPK proteins, whose activation is regulated via the phosphorylation of an upstream kinase, ERK3 is regulated through its abundance, as it is constitutively phosphorylated when present in the cell. While USP20 has been established as a deubiquitinating protein that stabilizes ERK3 by preventing its degradation, both TRIM21 and FBXW7 have been identified as E3 ligases that target …

Investigating Mono-Driver Versus Multi-Driver Oncogenesis Via Alterations To Cell Signaling Pathways, Abygail Chapdelaine

Open Access Dissertations

The latest edition of Global Cancer Statistics released by the American Cancer Society reported approximately 20 million newly diagnosed cancer cases worldwide in 2022 with 9.7 million deaths. This ranks cancer as the second leading cause of death worldwide. For the majority of cancer patients, the standard treatment approach is chemotherapy and/or radiation therapy, and while this has been successful in some patients, a lack of specificity to cancer cells causes severe side-effects when these agents damage healthy cells. Despite extensive efforts that have gone into developing targeted cancer therapeutics that block functions specific to cancer, they have not been …