Biochemistry Commons^™

Regulation Of Gene Expression By Rna Binding Proteins And Micrornas, Kyle Cottrell

Arts & Sciences Electronic Theses and Dissertations

Regulation of gene expression is essential to life. Post-transcriptional regulation of gene expression is a complex process with many inputs that lead to changes in localization, translation and stability of mRNAs. The translation and stability of many mRNAs is regulated by cis-elements, such as mRNA-structure or codon optimality; and by trans-acting factors such as RBPs and miRNAs. Here I report on the complex interactions between RBPs, miRNAs and characteristics of their target mRNAs in respect to effects on translation and RNA stability.

Using a reporter based approach we studied modulation of microRNA-mediated repression by various mRNA characteristics. We observed the …

Binding Affinity And Specificity Of Sh2 Domain Interactions In Receptor Tyrosine Kinase Signaling Networks, Tom Ronan

McKelvey School of Engineering Theses & Dissertations

Receptor tyrosine kinase (RTK) signaling mechanisms play a central role in intracellular signaling and control development of multicellular organisms, cell growth, cell migration, and programmed cell death. Dysregulation of these signaling mechanisms results in defects of development and diseases such as cancer. Control of this network relies on the specificity and selectivity of Src Homology 2 (SH2) domain interactions with phosphorylated target peptides. In this work, we review and identify the limitations of current quantitative understanding of SH2 domain interactions, and identify severe limitations in accuracy and availability of SH2 domain interaction data. We propose a framework to address some …

Determination Of The Effects Of Different Maillard Reaction Products On The Taxonomic Composition Of The Gut Microbiota, Nesreen Hamdan Aljahdali

Graduate Theses and Dissertations

The Maillard Reaction (MR) is a non-enzymatic chemical reaction which results in linkage between the amino group of amino acids and the carbonyl group of reduced sugars. This reaction generates Maillard reaction products (MRPs) which are not present naturally in foods, and are responsible for a range of colors, odors, flavors, and other sensory properties. Conflicting reports of MRPs impacts on human health are probably due to the fact that bioconversion of these digestible molecules by the gut microbiota has been marginally taken into account. This study aimed to determine the effects of different MRPs on rodent’s gut microbiota through16S …

Insights Into The Therapeutic Potential Of Salt Inducible Kinase 1: A Novel Mechanism Of Metabolic Control, Randi Fitzgibbon

Dissertations & Theses (Open Access)

Salt inducible kinase 1 (SIK1) has been considered a stress-inducible kinase since it was first cloned in 1999. Continued efforts since this time have been dedicated to characterizing the structure and function of SIK1. Such research has laid the ground work for our understanding of SIK1 action and regulation in tissue and stimuli dependent manners. The fundamental findings of this dissertation continue in this tradition and include investigations of SIK1 regulatory mechanisms in skeletal muscle cells, the cellular and physiological effects of SIK1 loss of function in vitro and in vivo, and intracellular metabolic and mitochondrial regulation by this …

Dissecting Molecular Pathways That Ensure Proper Chromosome Segregation And Cell Division, Anna Ye

Doctoral Dissertations

Equal segregation of the genome is a prerequisite for cell survival. During cell division the duplicated DNA is compacted into chromosomes and a multi-protein macrostructure, known as the kinetochore (Kt), is assembled on each copy of compacted DNA. Simultaneously, the mitotic spindle, which is made up of microtubules (MTs), is built to facilitate the equal distribution of the chromosomes between the resulting daughter cells. Kinetochores mediate the interaction between the MTs and the chromosomes, properly positioning them for segregation. To ensure that the DNA is equally divided in every cell division, cells have built a surveillance system to detect any …

Regulated Proteolysis Of Dnaa Coordinates Cell Growth With Stress Signals In Caulobacter Crescentus, Jing Liu

Doctoral Dissertations

DNA replication is an essential process in all domains of life. Replication must be precisely regulated, especially at the step of initiation. In bacteria, the replication initiator DnaA is regulated by multiple post-translational regulations to ensure timely replication. Caulobacter crescentus has the most strict replication regulation that DNA only replicates once per cell cycle, and proteolysis of DnaA identified in this species is the only irreversible way to inhibit DnaA, suggesting it might be pivotal to restricting DNA replication. However, the responsible protease(s) and mechanism for its degradation remain unclear since its first discovery in 2005. In this thesis, I …

Molecular Mechanisms Of C-Terminal Eps15 Homology Domain Containing (Ehd) Protein Function, Kriti Bahl

Theses & Dissertations

Endocytic trafficking is not only an essential process for the maintenance of cellular homeostasis but also plays a vital role in regulating diverse cellular processes such as signaling, migration and cell division. The C-terminal Eps 15 Homology Domain proteins (EHD1-4) play pivotal roles in regulating distinct steps of endocytic trafficking. Among the EHDs, EHD2 is disparate both in terms of sequence homology (70%) and its subcellular localization at the caveolae. The crystal structure of EHD2 has been solved and it contains an unstructured loop consisting of two proline-phenylalanine (PF) motifs: KPFRKLNPF. However, the other paralogs EHD1, …

The Structural And Functional Properties Of A Double Mutant Of Human Acidic Fibroblast Growth Factor (Hfgf-1), Arwa Mohammed Alghanmi

Graduate Theses and Dissertations

Human acidic Fibroblast Growth Factor 1 (FGF-1), a member of the FGF superfamily, is a potent mitogen and heparin-binding protein involved in a broad spectrum of biological processes, including angiogenesis, cell proliferation, and wound healing. Design of hFGF-1 with an increased thermal stability and an enhanced cell proliferation activity is highly desired for wound healing applications. Herein, we have designed the variant of FGF-1 by substituting two important amino residues in the heparin-binding pocket. The variant was overexpressed in Escherichia coli and was successfully purified to homogeneity using an affinity chromatographic procedure. Far-UV circular dichroism spectroscopic analysis showed that the …

Regulation Of Extracellular Signal-Regulated Kinase By Histone Deacetylase 6, Jheng-Yu Wu

USF Tampa Graduate Theses and Dissertations

Extracellular signal-regulated kinases 1/2 (ERK1/2) are important kinases regulating cell proliferation and cell migration, and have been established as therapeutic targets for cancer treatment. Previously, we found that ERK1 phosphorylates histone deacetylase 6 (HDAC6) to regulate its enzymatic activity. However, whether HDAC6 reciprocally modulates ERK1 activity is unknown. Here, we have discovered that ERK1/2 are acetylated proteins and shown that HDAC6 manipulates ERK1’s kinase activity via deacetylation. We demonstrated that both ERK1 and ERK2 interact with HDAC6 physically. We showed that the acetylation level of GST-ERK1/2 increased in a dose- and time-dependent manner upon treatment with a pan-HDAC inhibitor, Trichostatin …

The Key Question In Symbiotic Nitrogen Fixation: How Does Host Maintain A Bacterial Symbiont?, Onur Oztas

Doctoral Dissertations

The fact that plants cannot use nitrogen in the gaseous form makes them dependent on the levels of usable nitrogen forms in the soil. Legumes overcome nitrogen limitation by entering a symbiotic association with rhizobia, soil bacteria that convert atmospheric nitrogen into usable ammonia. In root nodules, bacteria are internalized by host plant cells inside an intracellular compartment called the symbiosome where they morphologically differentiate into nitrogen-fixing forms by symbiosome-secreted host proteins. In this project, I explained the host proteins required to maintain bacterial symbionts and described their delivery to the symbiosome. I showed that the SYNTAXIN 132 (SYP132) gene …

Mechanisms Of G Protein Regulation By Rgs Proteins And Small Molecule Inhibitors, Stanley Michinobu Kanai

Arts & Sciences Electronic Theses and Dissertations

G protein coupled receptors transduce diverse extracellular signals like hormones, neurotransmitters, and photons to specific cellular responses through heterotrimeric G proteins. G proteins activate numerous effectors and signal transduction pathways, and therefore the regulation of G proteins is crucial for faithful propagation of specific cellular and physiological responses. A better understanding of the mechanisms that regulate G proteins should provide new insight into signaling pathways that govern healthy and disease states, and also provide opportunities for discovery of novel therapeutic targets.Regulator of G protein signaling (RGS) proteins are crucial regulators of G proteins, for they control amplitude and duration of …

Phopsphorylation And Ubiquitin Modification At Dna Damage Sites In Response To Double-Strand Breaks, Atanu Paul

Dissertations & Theses (Open Access)

Genomes of all organisms are continuously damaged by numerous exogenous and endogenous sources leading to different kinds of DNA lesions, which if not repaired efficiently may trigger wide-scale genomic instability, a hallmark of cancer development. To overcome this, cells have evolved a sophisticated sensory network called the DNA damage response (DDR) comprised of a large number of distinct protein complexes categorized as sensor, mediator, transducer and effector proteins that amplify the DNA damage signal and activate cell cycle checkpoint to initiate DNA repair or trigger apoptosis where the defect is beyond repair. This intricate signaling pathway is tightly regulated by …

Understanding The Mechanism Of Genomic Instability During Replicative Aging In Budding Yeast, Sangita Pal

Dissertations & Theses (Open Access)

Aging brings a gradual decline in molecular fidelity and biological functionality, resulting in age related phenotypes and diseases. Despite continued efforts to uncover the conserved aging pathways among eukaryotes, exact molecular causes of aging are still poorly understood. One of the most important hallmarks of aging is increased genomic instability. However, there remains much ambiguity as to the cause. I am studying the replicative life span (RLS) of the genetically tractable model organism Saccharomyces cerevisiae, or budding yeast using the innovative “mother enrichment program” as the method to isolate unparalleled numbers of aged yeast cells to investigate the molecular changes …

The Role Of Adenosine Signaling In Mature Erythrocytes And Erythroid Progenitors, Hong Liu

Dissertations & Theses (Open Access)

Adenosine is a ubiquitous nucleoside in almost all the cells throughout our bodies. It is highly induced particularly under hypoxia or energy depletion conditions. Adenosine functions as a critical ligand, after binding to membrane-associated adenosine receptors, adenosine initiates a downstream signaling cascade and subsequently contributes to functions of nervous system, immune response, vascular function and even metabolism.

Hypoxia is a condition with limited O₂ availability in the whole body or a region of the body. It is a major consequence of many respiratory and cardiovascular diseases, as well as for people living and working at high altitudes or other …

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 …

Mechanisms Of Ikbke Activation In Cancer, Sridevi Challa

USF Tampa Graduate Theses and Dissertations

Cancer is the second leading cause of death in the USA and it is expected to surpass heart diseases making it important to understand the underlying mechanisms of cancer. The efforts to target single signaling molecule showed little success in increasing the patient survival and it can be due to increased compensation for cell survival by alternative pathway activations. Hence comprehensive understanding of the alternative signaling pathways may help us treat cancer better. Chronic inflammation is attributed to increased risk of cancer and emerging studies show the growing importance of both canonical and non-canonical IκB kinases such as IKKα, IKKβ, …

Alpha-Synuclein: Insight Into The Hallmark Of Parkinson's Disease As A Target For Quantitative Molecular Diagnostics And Therapeutics, Baggio A. Evangelista

Honors Undergraduate Theses

Parkinson’s disease (PD) is the second-most common neurodegenerative disease after Alzheimer’s disease. With 500,000 individuals currently living with Parkinson’s and nearly 60,000 new cases diagnosed each year, this disease causes significant financial burden on the healthcare system - amassing to annual expenditures totaling 200 billion dollars; predicted to increase through 2050. The disease phenotype is characterized by a combination of a resting tremor, bradykinesia, muscular rigidity, and depression due to dopaminergic neuronal death in the midbrain. The cause of the neurotoxicity has been largely discussed, with strong evidence suggesting that the protein, alpha-Synuclein, is a key factor. Under native conditions, …

Micro-Spectroscopy Of Bio-Assemblies At The Single Cell Level, Jeslin Kera

Honors Undergraduate Theses

In this thesis, we investigate biological molecules on a micron scale in the ultraviolet spectral region through the non-destructive confocal absorption microscopy. The setup involves a combination of confocal microscope with a UV light excitation beam to measure the optical absorption spectra with spatial resolution of 1.4 μm in the lateral and 3.6 μm in the axial direction. Confocal absorption microscopy has the benefits of requiring no labels and only low light intensity for excitation while providing a strong signal from the contrast generated by the attenuation of propagating light due to absorption. This enables spatially resolved measurements of single …

Examining The Role Of Grp And Lik1 In Wall Associated Kinase (Wak) Perception Of Pectin In The Plant Cell Wall, Jack Ryan Mitchell

Honors Projects

Wall associated kinases (WAKs) are cell membrane bound receptor kinases that bind pectin and pectin fragments (OGs).The binding of WAKs to pectin sends a growth signal required for cell elongation and plant development. WAKs bind OGs with higher affinity than native pectin and instead activate a stress response. Glycine rich proteins (GRPs) are secreted cell wall proteins of unknown function. Seven GRPs with 65% sequence similarity are coded on a 90kb locus of Arabidopsis chromosome 2. GRP3 and WAK1 have been shown to bind in vitro, but single null mutations have no discernible phenotype, suggesting that the GRPs are redundant. …

Studies Of Sumoylation In Regulating Mif Stability And Rangap1 Nucleo-Cytoplasmic Shuttling In Controlling Its Sumo Modification, Progga Sen

Wayne State University Dissertations

SUMOylation is an essential post-translational modification that regulates a variety of critical cellular pathways ranging from nuclear transport to protein stability. Accumulating lines of evidence have shown that a perturbation of the SUMOylation pathway is associated with human diseases, especially various types of cancer. Our recent proteomic studies revealed a drastic increase in levels of SUMO2/3 modification on the proinflammatory cytokine MIF in the metastatic breast cancer cell line compared to the non-metastatic control cell line. Interestingly, the increase in levels of both MIF and global SUMO-2/3 modification in the metastatic cells are positively correlated to that of unmodified MIF …

The Role Of N-Myc Downstream Regulated Gene 1 In Breast Cancer Lipid Metabolism, Christopher James Sevinsky

Legacy Theses & Dissertations (2009 - 2024)

Altered carbohydrate and lipid metabolism are increasingly well characterized hallmarks of aggressive breast cancers. While aerobic glycolysis, or “the Warburg effect”, is a well-established metabolic adaptation exploited by tumor cells, the understanding of unique aspects of cancer lipid metabolism lags behind. This is especially true regarding the coordination of complex lipid synthesis and trafficking pathways, which remains poorly understood. N-Myc Downstream Regulated Gene1 (NDRG1) is overexpressed in many solid tumors, but its function is unclear. The importance of NDRG1 is best exemplified by the effect of null mutations on human physiology: inactivating mutations give rise to the severe autosomal recessive …

Changes In Threonyl-Trna Synthetase Expression And Secretion In Response To Endoplasmic Reticulum Stress By Monensin In Ovarian Cancer Cells, Jared Louis Hammer

Graduate College Dissertations and Theses

Aminoacyl-tRNA synthetases (ARS) are a family of enzymes that catalyze the charging of amino acids to their cognate tRNA in an aminoacylation reaction. Many members of this family have been found to have secondary functions independent of their primary aminoacylation function. Threonyl-tRNA synthetase (TARS), the ARS responsible for charging tRNA with threonine, is secreted from endothelial cells in response to both vascular endothelial growth factor (VEGF) and tumor necrosis factor-α (TNF-α), and stimulates angiogenesis and cell migration. Here we show a novel experimental approach for studying TARS secretion, and for observing the role of intracellular TARS in the endoplasmic reticulum …

Functional And Mechanistic Consequences Of Dual Oxidase 1 Suppression In Lung Cancer, Andrew Charles Little

Graduate College Dissertations and Theses

The NADPH oxidase homolog, dual oxidase 1 (DUOX1), is an H2O2 producing transmembrane enzyme highly expressed in the airway epithelium. DUOX1-dependent redox signaling has been characterized to regulate many homeostatic processes in the lung epithelium, such as host defense, wound healing, and type II immune responses. Intriguingly, DUOX1 has been found to be suppressed in many epithelial cancers, including lung cancer, by hypermethylation of its promoter. Epigenetic silencing of DUOX1 in cancer is paradoxical to the understanding that tumors harbor elevated levels of reactive oxygen species (ROS), suggesting that DUOX1 may be a tumor suppressor.

Since DUOX1 loss occurs in …

Myo2 Motor Function In The Contractile Ring And The Regulation Of Fission Yeast Cytokinesis, Luther Woodrow Pollard

Graduate College Dissertations and Theses

Animals, fungi, and amoebas require an actomyosin contractile ring at the division site to perform cytokinesis. The contractile ring initiates and guides the invagination of the plasma membrane as it forms new barriers between the nuclei at the cell equator. Defects in the contractile ring can result in misdirected, delayed, or premature cytokinesis, which leads to abnormal chromosome numbers. Aneuploidies resulting from failed cytokinesis sometimes lead to aggressive forms of cancer. This dissertation was motivated by the goal of better understanding the properties of the contractile ring and how it drives cytokinesis.

Actomyosin is initially recruited to the cell equator …