Cell and Developmental Biology Commons^™

Uncovering Novel Small Regulatory Rna In Protostome, Sweta Khanal

Dissertations

Small RNAs play pivotal roles in post-transcriptional gene regulation across diverse phylum of protostomes. In this study, we investigate the functional significance of atypical miRNAs, mirtron miR-1017 in Drosophila. Through ectopic expression in neuronal cells, we demonstrate that miR-1017 extends lifespan by targeting its host transcript, acetylcholine receptor Dα2, and influencing its splicing. This novel trans-regulatory function suggests a mechanism for mirtron evolution, highlighting the interplay between splicing and post-transcriptional regulation. Additionally, we profile small RNA populations in the polychaete developmental model Capitella teleta, shedding light on the small RNA landscape in annelid worms. Our analysis reveals a rich …

Characterization And Genomic Analysis Of Two Escherichia Coli O157:H7 Bacteriophages Isolated From Pigeons, Mohamad I. Alolama

Dissertations

Enterohemorrhagic Escherichia coli, also known as EHEC, is a subset of Shiga toxin-producing E. coli (STEC), and it has recently been identified as one of the principal foodborne pathogens. E. coli O157:H7 is the most important serotype of STEC for its role in causing foodborne illnesses. E. coli O157:H7 could cause various gastroenteritis symptoms such as diarrhea, hemolytic uremic syndrome, hemorrhagic colitis, and thrombotic thrombocytopenic purpura and may cause death. Elimination of E. coli O157:H7 during food processing and storage is a possible solution. Bacteriophages have a significant impact on bacterial populations in nature due to their ability to …

Investigating Smooth Muscle Myosin Dynamics And Assembly In Physiology And Pathology, Maggie Bennett

Dissertations

Vascular smooth muscle cells (SMCs) surround many blood vessels throughout the body,where they dynamically alter vessel diameter to regulate blood pressure, provide structural integrity, and absorb shock on a beat-to-beat timescale. As smooth muscle function fails, profound vascular disease ensues, often with tragic results- even death. Smooth muscle myosin 2 (SM2) is the dominant motor protein that actuates contractility and allows SMCs to perform these vital functions. To function, SM2 monomers dynamically assemble into filaments, which upon SMC activation, associate with filamentous actin to drive contractility. Despite the critical contribution of SM2 to SMC function, foundational aspects of SM2 assembly …

Ferrocenium Salt Aided Substitution Reactions And Synthesis Of Glycosylated Curcumin Derivatives, Deva Saroja Talasila

Dissertations

Organic synthesis has been significantly advanced with the employment of transition metal complexes. The discovery of transition metal catalysts provided the synthetic community with powerful tools for accelerating reactions and making them more selective and efficient. Many chemical reactions do not happen without a catalyst.

Iron-based catalysts have several advantages for the chemical industry because it is a non-toxic and ecologically friendly metal. Our group previously found that ferrocenium cations with a 3+ oxidation state of iron-catalyzed propargylic substitution reactions at low temperatures. The sandwich structure of ferrocenes allows substituents to be introduced on the cyclopentadienyl rings, which allows for …

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 …

Elucidating The Role Of Kmt2d In Her2+ Breast Cancer, Emily Ma

Dissertations

Breast cancer is the second leading cause of cancer-related deaths among women in the US. Histone modifications are common epigenetic changes that have been implicated in several breast cancers, including HER2+ breast cancers. Current therapies, such as trastuzumab, block the HER2 receptor to reduce tumor growth. Unfortunately, HER2+ breast cancers have found ways to overcome these therapies, leading to tumor resistance, metastases, and poor survival outcomes. A better understanding of the underlying mechanisms contributing to HER2+ breast cancer resistance may lead to the discovery of novel therapeutic strategies to improve patient outcomes.One prognostic marker for poor survival in women with …

Dicer Represses Antiviral Innate Immunity Pathways In Mouse Embryonic Stem Cells, Chandan Gurung

Dissertations

Recent studies have demonstrated that embryonic stem cells (ESCs) are deficient in expressing type I interferons (IFN), the cytokines that play key roles in antiviral responses. However, the underlying molecular mechanisms and biological implications of this finding are poorly understood. In this study, I used a synthetic RNA-based assay that can simultaneously assess multiple forms of antiviral responses in ESCs. Dicer is an enzyme essential for RNA interference (RNAi), which is used as a major antiviral mechanism in invertebrates but not clear in vertebrates. RNAi activity is detected in wild-type ESCs but is abolished in Dicer knockout ESCs (D−/−ESCs) as …

Molecular Mechanism Of Action Of The Natural Polyphenolic Compound And The P300 Inhibitor “Carnosol” Against The Triple Negative Breast Cance, Halima Ali Mohammed Salem Alsamri

Dissertations

Carnosol, a naturally occurring Phyto polyphenol found in sage, oregano, and rosemary, has been extensively studied by our laboratory for its anticancer effects in various types of cancer. In human Triple-Negative Breast Cancer (TNBC), carnosol was shown to inhibit cellular viability, colony growth, induced cell cycle arrest, autophagy, and apoptosis. Nonetheless, very little is known about the molecular mechanism of action. In the current study, the ability of carnosol to inhibit metastasis and tumour growth was examined. Wound healing and invasion assays revealed that carnosol inhibited migration and invasion at non-cytotoxic concentrations of MDA-MB-231 cells. Also, carnosol was found to …

Human 5’-Tailed Mirtrons Are Processed By Rnasep, Mohammad Farid Zia

Dissertations

Approximately a thousand microRNAs (miRNAs) are documented from human cells. A third appear to transit non-canonical pathways that typically bypass processing by Drosha, the dedicated nuclear miRNA producing enzyme. The largest class of non-canonical miRNAs are mirtrons which eschew Drosha to mature through spliceosome activity. While mirtrons are found in several configurations, the vast majority of human mirtron species are 5’-tailed. For these mirtrons, a 3’ splice site defines the 3’ end of their hairpin precursor while a “tail” of variable length separates the 5’ base of the hairpin from the nearest splice site. How this tail is removed is …

Reconstituting The Cyanobacterial Circadian Clock In Vitro, Pyong Hwa Kim

Dissertations

Cyanobacteria are photosynthetic organisms that are known to be responsible for oxygenating Earth’s early atmosphere. Having evolved to ensure optimal survival in the periodic light/dark cycle on this planet, their genetic codes are packed with various tools, including a sophisticated biological timekeeping system. Among the cyanobacteria is Synechococcus elongatus PCC 7942, the simplest clock-harboring organism with a powerful genetic tool that enabled the identification of its intricate timekeeping mechanism. The three central oscillator proteins—KaiA, KaiB, and KaiC—drive the 24 h cyclic gene expression rhythm of cyanobacteria, and the "ticking" of the oscillator can be reconstituted inside a test tube just …

Identification And Characterization Of Novel Genes And Genetic Interactions That Influence Iba Metabolism, Vanessica Jawahir

Dissertations

Indole-3-butyric acid (IBA) is an endogenous storage auxin important for maintaining appropriate indole-3-acetic acid (IAA) levels that influences primary root elongation and lateral root development. IBA is metabolized into free IAA in the peroxisome in a multistep process similar to fatty acid β-oxidation. Although many components specific to IBA metabolism and peroxisome function have been identified, our understanding is incomplete. I sought to identify novel components of IBA metabolism or peroxisome function by conducting a forward genetic screen for Arabidopsis thaliana plants with enhanced resistance to IBA. I identified Long chain acyl-CoA synthetase 4 (LACS4) as a novel gene functioning …

The Role Of Homologous Cyclin B’S In The Cell Cycle And, In Their Absence, The Effect On Zebrafish Early Development, Tetiana Petrachkova

Dissertations

Regulation of cell division is essential for normal embryo development. The Cyclins and their Cyclin-dependent kinases are key regulators controlling this process. In this thesis, I examine the role of cyclin B1 and cyclin B2 in zebrafish development. It is thought that both Cyclins are necessary for a cell to progress past the G2/M checkpoint into mitosis. First, I show that zygotic Cyclin B1 is essential for normal cell cycle progression, but not for cells to enter mitosis. Lack of zygotic Cyclin B1 in the early arrest mutant specter, which carries a nonsense mutation in the cyclin B1 gene, …

The Histoplasma Capsulatum Ddr48 Gene Is Required For Survival Within Macrophages And Resistance To Oxidative Stress And Antifungal Drugs, Logan Blancett

Dissertations

Histoplasma capsulatum(Hc)is a systemic, dimorphic fungal pathogen that affects upwards of 500,000 individuals in the United States annually. Hc grows as a multicellular mold at environmental temperatures; whereas, upon inhalation into a human or other mammalian host, it transforms into a unicellular, pathogenic yeast. The research presented in this dissertation is focused on characterizing the DNA damage-responsive gene HcDDR48. HcDDR48was originally isolated via a subtractive DNA library enriched for transcripts enriched in the mold-phase of Hcgrowth. Upon further analysis we found that HcDDR48is not just expressed in the mold morphotype, but both growth programs …

The Antimicrobial Activity And Cellular Targets Of Plant Derived Aldehydes And Degradable Pro-Antimicrobial Networks In Pseudomonas Aeruginosa, Yetunde Adewunmi

Dissertations

Essential oils (EOs) are plant-derived products that have been long exploited for their antimicrobial activities in medicine, agriculture, and food preservation. EOs represent a promising alternative to conventional antibiotics due to the broad-range antimicrobial activity, low toxicity to human commensal bacteria, and the capacity to kill microorganisms without promoting resistance. Despite the progress in the understanding of the biological activity of EOs, many aspects of their mode of action remain inconclusive. The overarching aim of this work was to address these gaps by studying molecular interactions between antimicrobial plant aldehydes and the opportunistic human pathogen Pseudomonas aeruginosa. We initiated …

Roles Of Phospholipases And Ribosomal S6 Kinase In Lipid Remodeling And Growth In Arabidopsis Response To Phosphate Deprivation, Yuan Su

Dissertations

Phosphate (Pi) is one of three macronutrients for plants, which is vital for plant growth and development. Understanding the mechanism by which plants respond and adapt to Pi deficiency not only unveils functions of genes and pathways involved, but also provides potential tools to manipulate crops to better stand Pi stress in low Pi-containing lands. One of the significant metabolic changes in plants under Pi starvation is the membrane lipid remodeling that converts Pi-containing lipids such as phospholipids to Pi-free lipids, such as glycolipids. To elucidate the metabolism and regulation of lipid remodeling, this dissertation characterizes the role of two …

Development Of Innate Immunity During In Vitro Differentiation Of Mouse Embryonic Stem Cells, William D'Angelo

Dissertations

Embryonic stem cells (ESCs) hold enormous promise for the goals of regenerative medicine and tissue engineering, and extraordinary progress has been made in defining conditions for differentiation to desired cell types. However, an often overlooked aspect of ESC biology is innate immunity, the ability of cells to detect and respond to pathogens and inflammatory cytokines. A number of recent studies by our lab and others have established that ESCs and other types of pluripotent cells from both mice and humans do not mount typical immune responses to viral or bacterial stimuli. There are also indications that various cell types differentiated …

Uncovering The Identity And Metabolism Of Bacterial Coa-Rna, Joseph R. Spangler

Dissertations

Coenzyme A is an indispensable molecule in all known life with roles in metabolism, gene regulation, and macromolecule synthesis. As CoA is derived from RNA itself, it’s incorporation into RNA by in vitro methods has proven useful in research probing the origin of life based on the RNA World theory. The discovery in contemporary bacteria of RNA modified with CoA, however, provided an unexpected twist to previously well-characterized bacterial systems. The identity of sequences associated with CoA-RNA has been elusive since their discovery in 2009 based on the difficulties in isolation while maintaining RNA quality. The aim of this study …

The Impact Of Altered T Cell Receptor—Peptide-Major Histocompatibility Complex Interactions On Antigen Recognition And T Cell Function, Timothy T. Spear

Dissertations

Adoptive cell transfer (ACT) using T cell receptor (TCR) gene-modified T cells is an exciting and rapidly evolving field. Numerous basic science and clinical studies have demonstrated various levels of feasibility, safety, and efficacy using TCR-engineered T cells to treat cancer and viral infections. Genomic instability of targeted diseases, however, requires effective and safe TCRs to cross-recognize mutated antigens while minimizing on- or off-target toxicities. Thus, improvements to T cell-based therapeutics mandate a broader understanding of the principles governing antigen recognition. This dissertation addresses critical biologic questions evaluating which parameters are most important in facilitating antigen recognition, and how alterations …

The Role Of Alveolar Macrophages In Pulmonary Inflammation After Intoxication And Burn Injury, Jill Ann Shults

Dissertations

A positive blood alcohol concentration is detected in nearly half of burn patients admitted to the emergency room. The combined insult of being intoxicated at the time of burn injury results in more clinical complications, in comparison to non-intoxicated burn patients. Severe burn, with or without inhalation injury, is a common predisposing factor for the development of acute respiratory distress syndrome (ARDS). Exacerbated pulmonary inflammation and a net result of insufficient gas exchange underlie the large percentage of burn fatalities due to pulmonary complications. Previous studies in our laboratory indicate a drastic elevation in pulmonary inflammation in a mouse model …

The Gut-Liver Axis Regulates Pulmonary Inflammation After Intoxication And Burn Injury, Michael M. Chen

Dissertations

The increasing prevalence of binge drinking and its high association with traumatic injury warrant further investigation into the mechanisms underlying the worsened clinical outcomes associated with this combined injury. Existing literature suggests the liver is at the center of the postburn systemic response with crosstalk between the intestinal microbiome and the liver playing an important role after injury. There is a paucity in the literature, however, regarding the role of the gut-liver axis in the common clinical scenario of burns preceded by alcohol intoxication. Previous studies in our laboratory demonstrated intoxication potentiates postburn hepatic damage and IL-6 production in a …

Antiviral Responses In Mouse Embryonic Stem Cells: Differential Development Of Cellular Mechanisms In Type I Interferon Production And Response, Ruoxing Wang

Dissertations

Embryonic stem cells (ESCs) have been recognized as a promising cell source for regenerative medicine. Intensive research over the past decade has led to the possibility that ESC-derived cells will be used for the treatment of human diseases. However, increasing evidence indicates that ESC-derived cells generated by the current differentiation methods are not fully functional. It is recently recognized that ESC-derived cells lack innate immunity to a wide range of infectious agents and inflammatory cytokines. When used in patients, ESC-derived cells would be placed in wounded sites that are exposed to various pathogens and inflammatory cytokines; therefore, their viability and …

Effects Of Acute Alcohol Exposure On Post Burn Intestinal Immunity: Role Of Il-23, Juan L. Rendon

Dissertations

More than one million burn injuries are reported yearly within the United States. These injuries result in approximately 500,000 emergency room visits and 40,000 hospitalizations annually. Greater than 50% of these injuries occur under the influence of alcohol/ethanol (EtOH) intoxication. Burn victims who sustain injury under the influence of EtOH exhibit significantly higher rates of morbidity and mortality than patients without EtOH exposure at the time of injury. Several lines of evidence suggest that gut pathogens and/or their products may play a role in the development of sepsis and multiple organ failure reported in burn and trauma patients. In line …

Effect Of Advanced Age On The Innate Immune Response To Cutaneous Wound Infection, Aleah Lin Brubaker

Dissertations

An estimated 25 billion in US health care expenditure is spent on care of chronic, non-healing wounds. The failure to effectively heal wounds is often compounded by co-morbidities, such as diabetes or obesity. Another major patient population afflicted with chronic wounds are the elderly. Advanced age is associated with a decline in immunologic function that contributes to a poor response to vaccination, infection and tissue injury resulting in prolonged hospital stays and age-related morbidity and mortality. Specifically, clinical observations and laboratory studies have suggested an age-related decline in cutaneous wound healing, marked by protracted wound closure, wound dehiscence and chronic …

Effects Of Binge Alcohol Exposure On Canonical Wnt Signaling During Fracture Repair, Kristen Leigh Lauing

Dissertations

Excessive alcohol consumption is associated with increased fracture risk, delayed bone healing and fracture non-union. Binge alcohol consumption has occurred in 25-40% of all orthopaedic trauma cases. The canonical Wnt pathway, through tight regulation of stabilized beta-catenin, plays an essential and pivotal role in the formation of new bone and cartilage to initiate bone repair. We sought to determine the molecular mechanisms behind alcohol-related fracture complications by testing the hypothesis that binge alcohol exposure deregulates canonical Wnt signaling in the fracture callus, leading to impaired healing.

To test this, C57BL/6 or beta-catenin/TCF-reporter male mice were exposed to intraperitoneal alcohol or …

The Tumor Suppressor Pkcδ Is Critical For Uv-Induced G2/M Checkpoint Activation And Apoptosis, Edward Lewis Lagory

Dissertations

Protein kinase C delta (PKCδ) is an essential component of the intrinsic apoptotic program. Following DNA damage, such as exposure to UV radiation, PKCδ is cleaved in a caspase-dependent manner, generating a constitutively active catalytic fragment (PKCδ-cat) which is necessary and sufficient for keratinocyte (KC) apoptosis. We found that in addition to inducing apoptosis, expression of PKCδ-cat caused a pronounced G2/M cell cycle arrest in both primary human KCs and immortalized HaCaT cells. Consistent with a G2/M arrest, PKCδ-cat induced phosphorylation of Cdk1 (Tyr15), a critical event in the G2/M checkpoint. Treatment with the ATM/ATR inhibitor caffeine was unable to …

Notch Signaling Is Important In The Survival, Proliferation, And Self-Renewal Of The Putative Breast Cancer Stem Cell Population, Peter Grudzien

Dissertations

Numerous studies have identified stem-like cells, termed cancer stem cells (CSCs), in breast tumors and established cell lines. It has been hypothesized that CSCs are responsible for breast cancer formation, progression and recurrence; therefore, a deeper understanding of the signaling pathways regulating CSC survival will benefit development of novel therapeutic strategies. Notch signaling, which is dysregulated in breast cancer and has been implicated in mammary stem cell self-renewal, and can be effectively blocked by gamma-secretase inhibitors (GSIs). While GSIs are currently in clinical trials for breast cancer, it is not fully understood how these compounds will affect CSCs or if …

Characterization Of The Ap-1 And Nf-Kappa B Transcription Factors In The U-87 Mg Astrocytoma Cell Line, Denise L. Smith

Dissertations

Growth factors, signaling molecules and transcription factors are frequently mutated in cancer and platelet-derived growth factor (PDGF) has been shown to be over-expressed in most gliomas. In the astrocytoma cell line U-87 MG, the over-expression of PDGF has been shown to lead to over-expression of transcription factors activator-protein-1 (AP-1) and nuclear factor-kappa B (NF-κB), and it is believed that the individual subunits of AP-1 (c-Jun, JunB, JunD, c-Fos, FosB, Fra-1, and Fra-2) and NF-κB (c-Rel, p50, p52, p65, and RelB) play different roles in proliferation, survival and differentiation in the U-87 MG cell line.

To assess these different roles, transient …

Mapk Survival Signaling In Melanoma, Matthew W. Vanbrocklin

Dissertations

Extracellular signals activate mitogen-activated protein kinase (MAPK) cascades potentiating biological activities such as cell proliferation, differentiation, and survival. Constitutive activation of MAPK signaling pathways is implicated in the development and progression of many human cancers, including melanoma. Mutually exclusive activating mutations in NRAS or BRAF are found in ∼85% of all melanomas resulting in constitutive activation of the MAPK pathway (RAS-BRAF-MEK-ERK-RSK). We have previously demonstrated that inhibition of this pathway with small molecule MEK inhibitors selectively induces apoptosis in human melanoma cells both in vitro and in vivo, but not in normal melanocytes. These results support the notion that the …

The Effects Of Aging And Activity On Glial Cell Line-Derived Neurotrophic Factor Expression In Skeletal Muscle, Nathan Peplinski

Dissertations

Glial cell line-derived neurotrophic factor (GDNF) is an extracellular signaling protein that is produced by skeletal muscle and is important for the motor neurons that control muscle movement. GDNF has been shown to keep neurons alive under conditions that they would otherwise not persist. In skeletal muscle, GDNF has been shown to be one of the most potent neurotrophic factors that influence motor neuron survival. While the role of GDNF has been well studied during early development, not much is known about what happens to GDNF expression in the adult and with advanced aging. Previous results from our lab have …

Characterization Of Molecular Events Following Heavy Metal Cadmium Exposure: A Cellular Regulatory Symphony In Response To Early Cadmium Insult, Chin-Ju Jean Hsiao

Dissertations

Cadmium, a non-essential toxic heavy metal, has been classified as a human carcinogen and one of the most hazardous substances in our environment. Since it can circulate within the food chain, has a long biological half-life and can bio-accumulate, it has the potential to induce detrimental health effects linked to toxicity, cancer and chronic diseases.

In recent years progress has been made to dissect and understand the molecular mechanisms underlying the effects of cadmium exposure. However, few studies have focused on investigating potential protective and/or defense mechanisms naturally applied by cells upon initial exposure, prior to any damage or toxic …