Digital Commons Network^™

Molecular Investigation Of Minor Genomic Populations And Biological Exposures In Human Health, Brandon Ned Johnson

Dissertations and Theses

The study of genetics has contributed to countless discoveries related to human health and disease. However, the complexities of human biology reside not only in the genome but also in the contributions from environmental exposures, as measured via the classical twin design. To understand the influence of biological exposures, I implemented study designs to explore both the health associations and propagation of foreign genetic material. Microchimerism has been studied for association with several clinical conditions, and I further investigated if male microchimerism could elucidate the etiology of Mayer-Rokitansky-Küster-Hauser syndrome. Prevalence of male microchimerism in women with no history of pregnancy …

Regulation Of Erk3 By Kras Signalling And Its Role In The Growth Of Lung Adenocarcinoma (Luad) Cells, Shreya Akunapuram

Browse all Theses and Dissertations

Extracellular signal related kinase 3 (ERK3) is one of the atypical mitogen activated protein kinases (MAPK). It is expressed ubiquitously and plays a role in a variety of cellular processes, including cell growth and differentiation. ERK3’s role in promoting migration and invasion in various cancers has been well established. ERK3 is upregulated in non-small cell lung cancers (NSCLCs) and has been shown to promote NSCLC tumor growth and progression. However, the regulation of ERK3 in lung cancers remains largely unclear. A recent study indicates that ERK3 phosphorylation at S189, an indicator of ERK3 activity, is upregulated by KRAS in NSCLCs. …

Dissecting Interactions Across Gene Regulatory Layers In C. Elegans, Morgan Taylor

Biological Sciences Theses and Dissertations

The nematode Caenorhabditis elegans is a powerful tool for studying nervous system genetics. Though relatively simple compared to mammals, C. elegans boasts a remarkably well-conserved neuronal genome and proteome, and its utility in the characterization of neuronal genes has been well-established. However, gene expression is often controlled by complex interactions between multiple genes, and teasing apart the functions of individual genes within such networks remains a challenge. Dissecting these interaction networks is crucial in determining the multifaceted functions of important, conserved regulatory genes. Here we explore interactions between gene regulatory layers in the C. elegans nervous system, employing a synthetic …

Elucidating Mechanisms Of Biofluorescence And Bioluminescence In Marine Organisms, Andrew M. Guarnaccia

Dissertations, Theses, and Capstone Projects

Biofluorescence and bioluminescence are two methods of light emission that entail separate mechanisms of action but end at the same process: a colorful display that have tremendous ecological and behavioral benefits, whether it be used to communicate with conspecifics, camouflage into a multicolored background, attract unsuspecting prey, or alert others to a predator. In biofluorescence, higher-energy, shorter wavelength light is absorbed then re-emitted as lower-energy, longer-wavelength light. Bioluminescence on the other hand entails a chemical reaction in which a small molecule is oxidized by an enzyme, creating a high-energy intermediate that sheds the excess energy in the form of visible …

Assembly Of The Peripheral Arm Subunits Of Escherichia Coli Complex I And Analysis Of Clinical Mutations, Hind Alkhaldi

Biological Sciences Theses and Dissertations

Respiratory Complex I from E. coli is a proto-type of the mitochondrial enzyme, consisting of a 6-subunit peripheral arm (B-CD-E-F-G-I) and a 7-subunit membrane arm. When subunits E-F-G (N-module), were expressed alone they formed an active complex as determined by co-immunoprecipitation and native gel electrophoresis. When co-expressed with subunits B and CD, only a complex of E-F-G was found. When these five subunits were co-expressed with subunit I and two membrane subunits, A and H, a complex of B-CD-E-F-G-I was membrane-bound, constituting the N- and Q-modules. Assembly of Complex I was also followed by splitting the genes between two plasmids, …

Development And Analysis Of Next-Generation Polymeric And Bio-Ceramic Based Orthopedic Scaffolds By Advanced Manufacturing Techniques, Sudeep K. Gummadi

ETD Archive

Gliomas express mutant isocitrate dehydrogenases producing excessive amounts of D 2-hydroxyglutarate (D2HG) and releasing some of it into the environment. The immune surveillance is reduced as a result, however, the mechanisms behind lymphocyte suppression by the D2HG stereoisomer remain unknown. I incubated Jurkat T cells with D2HG at concentrations present within and surrounding gliomas, or its obverse L2HG stereoisomer, and quantified 2HG isomers within washed cells by TSPC derivatization with stable isotope-labeled D2HG and L2HG internal standards, HPLC separation, and mass spectrometry. D2HG was found in quiescent cells in double the amount of L2HG. External D2HG or L2HG increased the …

Instability And Extrachromosomal Circular Dna Formation At Microsatellites And Unstable Dna Sequences, Matilyn M. Shanahan

Browse all Theses and Dissertations

We have previously documented our evidence of genetic instabilities at the (Pu/Py)78 and (ATTCT)47 sequences and our reasoning for identifying break-induced replication (BIR) as the mode of repair responsible for the mutations in the DNA flanking the unstable inserts. Now, as our lab investigates the protein mechanisms at play in the BIR pathway taking place at these sites, we are also expanding our knowledge of how this mechanism extends into the pathways responsible for forming extrachromosomal circular DNA (eccDNA) molecules. We have documented the phenomena posed as the driving factors for eccDNA formation in our systems containing (Pu/Py)78 and (ATTCT)47. …

Genomic Instability At A Polypurine/Polypyrimidine Repeat Sequence, Nathen S. Zavada

Browse all Theses and Dissertations

Microsatellite repeat sequences have been shown to induce replication stalling, fork collapse, double-strand breaks (DSBs), and possibly stimulate break-induced replication. In this study we use a dual-fluorescent HeLa model that is designed to monitor recombination at an ectopic site through use of flow cytometry and inverse PCR with a microsatellite in the lagging strand for DNA synthesis. To test the stability of the 78 bp polypurine/pyrimidine repeat from the PDK1 locus, we subjected cells to replication stress drugs designed to induce DSBs and measure break-induced replication (BIR). The study revealed that polypurine repeat cells undergo endogenous stress contributing to instability …

The Role Of Pi4kb In Cellular Localization Of Small Gtpases, Parisa Sadrpour

Browse all Theses and Dissertations

Constitutively active oncogenic mutant K-Ras is one the principal contributors to human cancers including 90% of pancreatic, 50% of colorectal and 32% of non-small cell lung cancers. However, except for K-Ras G12C oncogenic mutant, which only presents in about 13% of non-small cell lung cancer patients, there is no anti-K-Ras therapy for a considerable subset of K-Ras mutations in human tumors, reflecting challenges for targeting oncogenic K-Ras activity. K-Ras is a membrane-bound small GTPase; when active, it triggers multiple signaling pathways regulating a variety of key cellular functions such as cell growth, proliferation and survival. To initiate these signaling cascades, …

Reconstruction Of Gut Microbiome Via Intermittent Feeding, Kourtney Sprague

Browse all Theses and Dissertations

The benefits of intermittent fasting have been studied across many facets of health. It is known that physiologically fasting results in a metabolic switch from liver-derived glucose to adipose cell derived ketones to be used as energy and signaling molecules. Fasting down-regulates inflammation, increases expression of antioxidant defenses, and activates pathways for DNA repair and autophagy. While fasting or intermittent feeding effects on host physiology have been identified, the relationship between fasting and gut microbiome is not as well known. In times of gut rest, resident microbes undergo detoxication and motility, and in times of activity, gut microbes undergo DNA …

Molecular Mechanisms Of Aberrant Protein Glycosylation In Pancreatic Cancer Stemness And Metastasis, Frank Leon

Theses & Dissertations

A myriad of genetic and other abnormal changes underlies the aggressiveness and dissemination properties observed in pancreatic cancer (PC). Aberrant protein glycosylation is a commonly observed feature in PC. The modification of protein O-glycosylation is mediated by glycosyltransferases, which attach and sequentially elongate monosaccharides on Serine/Threonine (Ser/Thr) motifs. Aberrant glycosylation is recognized as an emerging hallmark of cancer where a disruption in normal glycosylation results in irregular O-glycans.

This dissertation research has investigated the consequences of aberrant protein glycosylation on stemness and enhancement of metastatic properties in pancreatic ductal adenocarcinoma (PDAC). Several publications have reported aberrant O-glycosylation increases in oncogenic …

Dynamic Mechanisms Of Multidrug Resistance In Human Cancers And Gram Negative Pathogens, Lauren Ammerman

Biological Sciences Theses and Dissertations

This dissertation focuses upon dynamic agents of multidrug resistance (MDR). We used a combination of in silico and in vitro techniques to investigate two membrane transporters that confer MDR – P-glycoprotein, which confers MDR in human cancers, and MtrD, which confers MDR in Neisseria gonorrhoeae. Inhibitors targeting both proteins have tremendous potential for use as co-therapeutics in the treatment of multidrug resistant cancers, or of multidrug resistant infections. However, previously identified inhibitors of P-gp have failed clinical trials due to off-target effects and associated toxicities. Furthermore, the molecular mechanism of antibiotic transport by MtrD is poorly understood, and this dearth …

Purification And Functional Characterization Of The Iron-Responsive Transcription Factor Aft1 From C. Glabrata, Jade Ikahihifo-Bender

Senior Theses

Due to its unique ability to serve as both an electron donor and acceptor, iron is utilized as a co-factor for many biological processes, including electron transfer, oxygen binding, and vitamin synthesis. Iron is also a key factor during fungal infections as the human host and invading pathogens battle over limited iron pools. The primary iron-responsive transcription factor Aft1 in the opportunistic pathogenic yeast Candida glabrata responds to iron deficiency by activating expression of iron acquisition genes. However, the mechanisms for sensing intracellular iron levels and regulating Aft1 activity in response to iron are unknown. The C. glabrata iron regulation …

Quantitated Effects Of Nutritional Supplementation On Exercise Induced Sweat, Andrew Blake Austin Browder

Browse all Theses and Dissertations

Discovery studies have identified many metabolites contained in human sweat. However, quantitative analysis of the sweat metabolome content remains mostly unknown. Furthermore several attributes, including rate, have been defined to affect sweat metabolite content, while other effectors, like diet, remain unknown. This study works to quantitatively define the metabolite impact caused by nutritional supplementation. To better understand the effect diet plays, a LC-MS method was developed focusing on improving resolution and peak width. While the literature provided examples of how diet affected sweat metabolite concentrations, the long-term effects of diet have not been explored. The experiment described here attempts to …

Potential Drug Treatment For Duchenne Muscular Dystrophy Which Could Be Through Upregulation Of Lipin1, Rajsi Y. Thaker

Browse all Theses and Dissertations

Duchenne muscular dystrophy (DMD) is a genetic disorder leading to progressive muscle degeneration and weakness due to mutation in dystrophin gene, which is very important for maintaining muscle membrane integrity. Dystrophin is the largest gene in the human genome therefore more prone to mutation. There is currently no cure for DMD. Our lab recently found that Lipin1 deficient myofibers showed upregulation of necroptosis correlated with the loss of muscle membrane integrity. Our primary approach for ameliorating dystrophic phenotype in DMD is through reduction of necroptosis using drugs which can potentially upregulate Lipin1 expression. In this study, we identified two drugs …

A Proteomic Analysis Of Corydoras Sterbai Secretions And Tissues, Erik Powell Wictor

University of the Pacific Theses and Dissertations

Defensive mechanisms vary widely in the animal kingdom ranging from physical defenses like spines to chemical defenses such as toxins. Toxins in these secretions and tissues can fluctuate from enzymes to lipids to uncharacterized chemicals. Next generation -omics technology and mass spectrometry are extremely important in analyzing these samples because of their ability to distinguish minute amounts of toxic substance within a complicated sample. The goal of this experiment was to look at secretions and tissues from Corydoras sterbai. All samples in this study were proteolyzed using a mixture of Trypsin and Lys-C, fractionated, and run through nanoLC-MS/MS analysis using …

Alternative Splicing Of Mdm4 In Human Melanomas, Abdullah Salem S. Alatawi

Browse all Theses and Dissertations

Melanoma is a potentially lethal type of skin cancer and regarded to be the third most common type of skin cancer. Although melanoma is not as common as basal cell carcinoma (BCC) and squamous cell carcinoma (SCC), it is more likely to metastasize than BCC and SCC. Interestingly, the incidence of melanoma continues to go up (expected 2% in 2020), but the deaths continue to decrease (-5.3% in 2020) due to improvements in detection and treatment. The treatment of melanoma depends on several aspects but most importantly the tumor's stage and the location. In the early stages, melanoma can be …

Apoptosis And Necrosis Drive Muscle Fiber Loss In Lipin1 Deficient Skeletal Muscle, Sandhya Ramani Sattiraju

Browse all Theses and Dissertations

Mutations in lipin1 are suggested to be a common cause of massive rhabdomyolysis episodes in children, however, the molecular mechanism involved in the regulation of myofiber death by lipin1 is not known. In this study, we utilized the skeletal muscle from cell-type-specific lipin1 knockout (Lipin1Myf5cKO) mice to define cell death pathways involved in lipin1 deficient muscles. We observed a significant increase in centrally nucleated fibers and embryonic myosin heavy chain (EMyHC)-positive regenerating fibers in Lipin1Myf5cKO mice compared to wild-type (WT) mice, indicating an increased cycle of degeneration and regeneration in lipin1 deficient muscles. Lipin1 deficient muscles had significantly elevated pro-apoptotic …

Peroxiredoxin 6 And Inflammation In Alzheimer's Disease, Jared Ferrell-Penniman

Biological Sciences Theses and Dissertations

Alzheimer’s disease (AD) is known for its debilitating symptoms and poor prognosis. However, despite intense research into neurodegenerative diseases, there are few therapies targeted at the underlying mechanisms of the disease. Oxidative stress (OS) and inflammation are cellular phenomena thought to be key to the progression of the disease. Critically, peroxiredoxin 6 (Prx6), an antioxidant protein with multiple functions, has been identified from mammalian studies as a potential regulator of both OS and inflammation that may have a specific effect on AD. This project seeks to elucidate the role of Prx6 in AD as well as the underlying mechanisms. Drosophila …

Characterization Of Epigenetic Plasticity And Chromatin Dynamics In Cancer Cell Models, Diana Lea Gerrard

Graduate College Dissertations and Theses

Cancer progression is driven by cumulative changes that promote and maintain the malignant phenotype. Epigenetic alterations are central to malignant transformation and to the development of therapy resistance. Changes in DNA methylation, histone acetylation and methylation, noncoding RNA expression and higher-order chromatin structures are epigenetic features of cancer, which are independent of changes in the DNA sequence. Despite the knowledge that these epigenetic alterations disrupt essential pathways that protect cells from uncontrolled growth, how these modifications collectively coordinate cancer gene expression programs remains poorly understood. In this dissertation, I utilize molecular and informatic approaches to define and characterize the genome-wide …

∆Np63Α Positively Regulates Erk3 Expression In Non-Melanoma Skin Cancer, Eid Salem Alshammari

Browse all Theses and Dissertations

Non-melanoma skin cancer (NMSC) is a group of skin cancer that includes basal cell carcinoma of the skin (BCC), squamous cell carcinoma of the skin (SCC), actinic keratoses, a precursor to SCC, and other rare cutaneous carcinomas. p63, a member of the p53 gene family, is an important regulator for epithelial tissue growth and development. ∆Np63α, a main isoform of p63, is highly expressed in NMSC and plays essential roles in NMSCs development. Extracellular signal-regulated kinase 3 (ERK3) is an atypical member of the MAPK family. It possesses a single phosphorylation site (serine 189) in its activation loop, which makes …

Lipin1 Regulates Skeletal Muscle Differentiation Through The Pkc/Hdac5/Mef2c:Myod -Mediated Pathway, Abdulrahman M. Jama

Browse all Theses and Dissertations

Our previous characterization of global lipin1-deficient (fld) mice demonstrated that lipin1 played a novel role in skeletal muscle (SM) regeneration. The clinical relevance of lipin1 has been observed in patients with lipin1 null mutations where they exhibited severe rhabdomyolysis with aggregated and dysfunctional mitochondria. Lipin1 is a key gene that plays an important role in lipid biosynthesis and metabolism. It has dual functions as it contains a phosphatase activity that converts phosphatidic acid (PA) to diacylglycerol (DAG), the penultimate step in triglycerides (TAG) biosynthesis as well as transcriptional co-activator function. In the cytosol and ER, lipin1 carries out its lipid …

Avicin Is A Potent Sphingomyelinase Inhibitor That Blocks K-Ras Plasma Membrane Interaction And Its Oncogenic Activity, Christian M. Garrido

Browse all Theses and Dissertations

Ras proteins are small GTPases that regulate cell growth, differentiation and apoptosis. There are three main isoforms: H-, N-, and K-Ras in mammalian cells, and they cycle between an active GTP- and inactive GDP-bound states. Constitutively active Ras mutations are found in ~15% of all human cancers. Of those, oncogenic K-Ras is found in ~98% of pancreatic, ~52% colorectal, and ~32% of lung cancers. In nearly 30 years since its discovery, there are no anti-K-Ras drugs currently available for clinical use. Since K-Ras must be localized to the plasma membrane (PM) for its full biological activity, targeting K-Ras PM interaction …

Phylogenetic Diversity Of The Bacterial Communities In Craft Beer, Lindsey Rodhouse

Graduate Theses and Dissertations

The craft beer industry is increasing in popularity in the United States. The craft brewing process typically does not use a pasteurization step, therefore the boiling process is the primary critical control step. Any microorganisms introduced after boiling, or those that are not killed during boiling, are likely to participate in fermentation and persist in the final product. Previous culture-based studies have isolated bacteria and yeast from craft beers at specific time points, but little research has been done on the process as a whole. The objectives of this research are to (1) track bacteria development throughout the brewing process …

The Function Of Renalase, Brett Allen Beaupre

Theses and Dissertations

Renalase was originally reported to be an enzyme secreted into the blood by the kidney to lower blood pressure and slow heart rate. Despite multiple reports claiming to confirm this activity in vivo there has been considerable discord in regards to the reaction catalyzed by renalase. The structural topology of renalase resembles that of known flavoprotein oxidases, monooxygenases and demethylases, but the conserved active site residues are unique to renalase. It has been reported that the catalytic function of renalase is to oxidize circulating catecholamines, however in vitro studies have failed to demonstrate a catalytic activity in the presence of …

Novel Insight Into The Role Of Lxrα In Metabolic Regulation Via Dna Binding As A Heterodimer With Pparα And As A Homodimer, Andrea M. Klingler

Browse all Theses and Dissertations

Liver X receptor a (LXRa) plays a critical role in the maintenance of energy homeostasis within a cell through tight transcriptional regulation of genes involved in metabolism of lipids, glucose, and cholesterol. Although LXRa has been established to function as a heterodimer with the retinoid X receptor a (RXRa), recent studies have determined that LXRa also interacts directly with peroxisome proliferator-activated receptor a (PPARa). However, little is known regarding the functionality of this heterodimer, if any exists at all. This study determined that a heterodimer of PPARa and LXRa is capable of binding to candidate response elements in vitro with …

A Bioluminescence Sensor Of Nlrp3 Inflammasome Activation, Michael Alexander Winek

Master's Theses

The innate immune system is many organisms first line of defense against pathogenic insult or tissue damage. This defense strategy is intent on restoring homeostasis upon perturbation. Upon activation of the innate immune system in humans, an oligomeric protein complex termed the “Inflammasome” forms in myeloid cells. The canonical output of activation of any subset of inflammasome is Caspase-1-mediated secretion of pro-inflammatory cytokines IL1β and IL18. Chronic or uncontrolled inflammasome activation is at the core of myriad economically burdening diseases. In many of these diseases, endogenous factors chronically engage the innate immune system. To study these diseases in in vivo, …

Emetine As An Anti-Cancer Therapeutic In Bladder Cancer, Valerie Davidson

Master's Theses

Bladder cancer is a serious health concern among the older population, as it is responsible for thousands of deaths annually in the United States. Patients that are diagnosed with muscle-invasive disease have a 5-year survival rate of only 20 percent. Additionally, muscle-invasive disease has a high metastatic potential; half of all patients develop metastatic disease within 3 years. Patients with muscle-invasive disease are presented with few treatment options aside from surgery. The current standard of care is a chemotherapeutic combination therapy of cisplatin and gemcitabine. This therapy is highly toxic, and due to the high instance of co-morbidities in these …

Y- Box Protein 2 (Ybx2) Is A Major Mrna Specific Regulator Of Translation In Spermatogenesis And The Translational Regulation Of The Sperm Mitochondria Associated Cysteine Rich Protein (Smcp) Mrna, Tamjid A. Chowdhury

Graduate Doctoral Dissertations

Spermatogenesis is the process by which diploid stem cells differentiate into haploid male gametes, spermatozoa. As haploid cells differentiate into spermatozoa, they undergo profound changes in morphology and physiology, including total reorganization of chromatin in the nucleus which results in the inability to synthesize new mRNAs about one week before the differentiation of sperm is complete. Consequently, many mRNAs, such as the protamine 1 (Prm1) mRNA and sperm mitochondria-associated cysteine rich-protein (Smcp) mRNA, are transcribed in early haploid spermatogenic cells and stored as translationally inactive messenger ribonucleoprotein particles (free-mRNPs) for several days to a week before the mRNA is translated …

Elucidating The Role Of Dopamine In The Avoidance Response Of Caenorhabditis Elegans Utilizing Microfluidics, Patricia Frances Turturro

Theses Digitization Project

This study looks at dopamine modulation of ASH neurons, which are part of the avoidance circuit of C. elegans, and their response to water soluble stimuli. Because nematodes such as Caenorhabditis elegans utilize dopamine at the molecular and cellular level in much the same way as mammals, they offer researchers a unique opportunity to study the mechanisms of dopamine signaling.