Life Sciences Commons^™

Role Of Integrin-Linked Kinase In Epidermal Integrity And Barrier Function, Samar Sayedyahossein

Electronic Thesis and Dissertation Repository

Integrin-linked kinase (ILK) is a β integrin adaptor protein that translates extracellular stimuli to intracellular signaling events. ILK plays a role in actin cytoskeleton dynamics and cell adhesion. The structure and function of the epidermis is highly dependent on cell-cell adhesion and cell-basement membrane interactions. The mechanisms whereby ILK contributes to epidermal integrity are poorly understood. Using a mouse model of epidermis-restricted Ilk gene inactivation, I observed that ILK loss causes abnormal morphology and presence of intra-epidermal and epidermal-dermal microblisters in embryos as early as E17.5. ILK-deficient epidermis is also characterized by abnormal localization or/and absence of adherens junctions, tight …

Reverse Genetics Approach To Examine Myosin Xi Functions In Pollen Tube Growth, Stephanie Lin Madison

Doctoral Dissertations

Pollen tube growth is an essential aspect of plant reproduction because it is the mechanism through which non-motile sperm cells are delivered to ovules thus allowing fertilization to occur. A pollen tube is a single cell that only grows at the tip, and this tip growth depends on actin filaments. Plants encode class VIII and class XI myosins as actin-based motor proteins, of which class XI myosins are required for cell expansion in vegetative tissues.

In Arabidopsis thaliana, 6 of 13 myosin XI genes are expressed in pollen: XIA, XIB, XIC, XID, XIE, and …

The Role Of Nag-1 In Tumorigenesis, Kyung-Won Min

Doctoral Dissertations

This dissertation explores the nature of a divergent member of the Transforming Growth Factor-β [beta] superfamily, the non-steroidal anti-inflammatory drugs activated gene (NAG-1), as it relates to its regulation and biological activity in cancer context. Our lab has extensively studied on the molecular mechanism by which phytochemicals and NSAIDs induce apoptosis correlation with NAG-1 expression in human colorectal cancer (CRC) cells. Significant data from in vitro studies suggest that NAG-1 has an anti-tumorigenic activity which elicits apoptosis in a cyclooxygenase (COX)-independent manner in CRC cells. Indeed, NAG-1 transgenic mice developed less aberrant polyp foci (APC) compared to those of control …

Understanding The Meiotic Roles Of Sisters Unbound In Drosophila Melanogaster, Badri Krishnan

Doctoral Dissertations

During meiosis, cohesin is required for sister chromatid cohesion and for formation of chromosome cores. Multiple processes including chromosome segregation, recombination and synaptonemal complex (SC) are dependent on cohesin. Cohesin complex consists of two SMC subunits- SMC1, SMC3 and two non-SMC subunits RAD21/REC8 in meiosis and SA. But in Drosophila, non-SMC subunits have not been shown to be required for cohesion. We have identified a gene sisters unbound, which along with previously identified ord and solo, form a group of three genes (sos) which do not have any sequence similarity to cohesins but performs functions …

Characterization Of Histidine Decarboxylase In Drosophila Using An Internal Flag Epitope, Maxwell Mianecki

Masters Theses

Histamine is a neurotransmitter in arthropods and is responsible for synaptic transmission in vision, mechanosensation, temperature sensing and sleep cycle in Drosophila. Histamine is synthesized by the enzyme histidine decarboxylase (HDC). While histamine is detectable within tissues using current immunofluorescent labeling techniques, immunological approaches have not been successful for HDC itself, with both direct antibodies and terminal epitope tags determined to be ineffective. In order to avoid loss of the epitope tag through putative N-­‐ and C-­‐terminal proteolytic cleavage, known to occur for HDC in other organisms, an internal epitope tag that does not disrupt enzyme function was utilized. A …

Mdm2-Mediated Degradation Of Sirt6 Phosphorylated By Akt1 Promotes Tumorigenesis And Trastuzumab Resistance In Breast Cancer, Umadevi Thirumurthi

Dissertations & Theses (Open Access)

Sirtuin6 (SIRT6) is one of the members of the Sirtuin family and functions as a longevity assurance gene by promoting genomic stability. It also regulates various cancer-associated pathways and was recently established as a bonafide tumor suppressor in colon cancer. This suggests that SIRT6 is an attractive target for pharmacological activation in cancer treatment, and hence, identification of potential regulators of SIRT6 would be an important and critical contribution towards cancer treatment. Here, we show that AKT1 phosphorylates SIRT6 at Ser³³⁸ and induces MDM2-SIRT6 interaction, priming SIRT6 for degradation via the MDM2-dependent ubiquitin-proteasome pathway. Blocking SIRT6 Ser³³⁸ phosphorylation …

The Role Of Stem Cells In Adipose Tissue Remodeling., Candice Reshay Holden 1988-

Electronic Theses and Dissertations

The work presented in this dissertation outlines the role of stem cells in the remodeling of adipose tissue under conditions of nutrient excess. Obesity-induced variations in adipose tissue stem cell distribution were uncovered by analysis of the stromal vascular fraction isolated from mice fed a high fat diet for several weeks. Bone marrow cell transplantation was used to determine the derivation of progenitor cells found in adipose tissue; and supplementation of depleted progenitor cell populations achieved via direct cell transplantation, helped to determine the contribution of these progenitor cells to the remodeling process. The dissertation is presented in five chapters …

Arsenic Targets Neural Plate Border Specifier Cells In P19 Cells, Christopher Mccoy

All Theses

Epidemiological studies have shown that arsenic exposure during early embryogenesis can cause reduced weight gain and neurological deficits later on in life. In addition, in vitro and in vivo studies have indicated that arsenic suppresses neurogenesis and myogenesis. The exact mechanism of how arsenic causes these undesired developmental outcomes is poorly understood, however both skeletal muscle and sensory neuron development require the Wnt/β-catenin signaling pathway to initiate the specific differentiation of precursor cells. We were interested in determining the target cell population of arsenic and its metabolites. Arsenic's metabolites were of interest because they have been shown to be more …

Investigation Of Nuclear And Cytoplasmic Functions Of The Dlipin Protein Of Drosophila Melanogaster, Qiuyu Chen

Graduate Theses and Dissertations

Lipin family proteins are highly conserved proteins present in species ranging from mammals to yeast. Lipin 1, the first Lipin gene identified in fatty liver dystrophy (fld) mutant mice, encodes the bifunctional protein Lipin 1, which can serve as an Mg2+-dependent phosphatidic acid phosphatase (PAP) and transcriptional co-regulator. dLipin, the single Lipin ortholog of Drosophila melanogaster, is required in triglyceride synthesis and fat body development. To study the transcriptional co-regulator activity of dLipin, nuclear receptors were screened to find receptors that interact with dLipin. The genetic interaction data indicated that Drosophila hepatic nuclear receptor 4 (HNF4) was a promising candidate …

Novel Posttranslational Modification In Lkb1 Activation And Function, Szu-Wei Lee

Dissertations & Theses (Open Access)

Cancer cells display dramatic alterations in cellular metabolism to meet their needs of increased growth and proliferation. In the last decade, cancer research has brought these pathways into focus, and one emerging issue that has come to attention is that many oncogenes and tumor-suppressors are intimately linked to metabolic regulation (Jones and Thompson, 2009). One of the key tumor-suppressors involved in metabolism is Liver Kinase B1 (LKB1). LKB1 is the major upstream kinase of the evolutionarily conserved metabolic sensor—AMP-activated protein kinase (AMPK). Activation of the LKB1/AMPK pathway provides a survival advantage for cells under energy stress. LKB1 forms a heterotrimeric …

Lkb1 Deficient Non-Small Cell Lung Cancer Cells Are Vulnerable To Energy Stress Induced By Atp Depletion, Chao Yang

Dissertations & Theses (Open Access)

Lung cancer is the second most frequent cancer in United States, which represents about 13.5% of new cancer cases every year. It accounts for about 27.2% of all cancer related deaths, which is more than the sum of deaths caused by prancretic, breast and colorectal. On average, only about 16% of lung cancer patients survive beyond 5 years. LKB1 is the third most mutated gene in lung cancer. It has been shown that LKB1 is mutated in at least 15% to 30% of NSCLC. Tumor with LKB1 mutation is associated with poor differentiation, high metastasis and worse response to chemotherapy. …

P120-Catenin Regulates Rest And Corest, And Modulates Mouse Embryonic Stem Cell Differentiation, Moonsup Lee

Dissertations & Theses (Open Access)

The canonical-Wnt pathway and beta-catenin have been extensively studied to determine their contributions to stem cell biology, but less is known about p120-catenin in the nuclear compartment. P120 is developmentally required as a consequence of its biochemical and functional interactions with cadherins, small-GTPases and transcriptional regulators. We report here that p120-catenin binds to and negatively regulates REST and CoREST, that others have indicated form a repressive complex having diverse key roles in developmental and pathologic gene regulation. We thus provide the first evidence for a direct upstream modulator of REST/CoREST function. Using mouse embryonic stem cells (mESCs), mammalian cell lines, …

The Identification And Characterization Of A Group Of Er Tpr-Containing Adapter Proteins, Johan C. Sunryd

Doctoral Dissertations

The endoplasmic reticulum (ER) is an organelle responsible for a variety of functions in all eukaryotic cells. Some of these functions are localized to specific regions in the ER, such as the ribosome-studded rough ER sheets or the ribosome-free smooth ER sheets. The smooth ER sheets have the ability to form ER tubules, which extend throughout the cell and make contact with other organelles. In order to accommodate these localized functional regions, a certain degree of heterogeneity and compartmentalization into sub-domains exists within the ER. Since the ER membrane and lumen are contiguous, the compartmentalization of the ER into sub-domain …

Characterization Of Ca2+ Influx Pathway(S) During Mouse Oocyte Maturation, Banyoon Cheon

Doctoral Dissertations

Ca²⁺ signaling induced at fertilization, also known as oscillations, is essential in mammalian eggs to initiate early embryonic development. The generation of the oscillations relies on optimization of Ca²⁺ toolkit components during oocyte maturation. In this dissertation, we intend to deepen our understanding of how this differentiation of the Ca²⁺ toolkit, especially those components associated with Ca²⁺ influx, is achieved during maturation, and how it contributes to the filling of the ER Ca²⁺ store during the maturation and fertilization. We first identified the expression and characterized the function of the components of the Store Operated …

Neural Stem Cells As A Model To Study Huntington’S Disease, Rawan Bakhsh

Dissertations, Masters Theses, Capstones, and Culminating Projects

Huntington’s disease (HD) is a heritable neurodegenerative disorder that affects muscle coordination and diminishes cognitive abilities, by affecting the medium spiny neurons in the brain. In HD patients, neurons are damaged and destroyed because of the toxicity of the mutant Huntington protein (mHtt). The mechanism of how mHtt protein affects the neurons is unknown. In this study we explored the effects of mHtt expression by looking at changes in huntingtin localization, changes in the expression and co-localization of related proteins and differences in cell morphology. We examine how this expression affects the cytoskeletal structures using neural stem cells Q7 (wild …

Genetic Analysis Of Ton2 Dependent Processes In Microtubule Organization In Arabidopsis Thaliana, Samantha Nicole Atkinson

Theses and Dissertations

Microtubules carry out many functions within the cell. They're used during mitosis and meiosis to move chromosomes to opposite sides of the cell so that cell division can occur. They're also used as structural support for the cell and are involved in establishing the cell shape. Microtubules are also able to reorient from a longitudinal orientation to a transverse orientation in response to gibberellic acid and auxin (two hormones that are involved in cell elongation). The mechanism for how this reorientation occurs is unknown. The TONNEAU2 (TON2) protein is necessary for proper microtubule organization. We looked at how TON2 affects …

Modulation Of Host Phosphatidylinositol Phosphates By Salmonella Effector Protein Sopb, Heather L. Piscatelli

Open Access Dissertations

Salmonella spp. are gram negative bacteria capable of infecting a number of eukaryotic hosts. In humans, Salmonella infection can range anywhere from acute gastroenteritis to typhoid fever which can oftentimes be fatal.Salmonella are facultative intracellular pathogens that have acquired the ability to enter non-phagocytic cells such as those lining the intestinal epithelium. Uptake into epithelial cells is mediated by the Salmonellapathogenicity island 1 (SPI1) encoded type III secretion system (T3SS), a needle-like complex composed of over 20 proteins that translocates effector proteins directly into the host cell cytosol. Salmonella possess a second type III secretion system encoded on …

Associated Behavioral, Genetic, And Gene Expression Variation With Alternative Life History Tactics In Salmonid Fishes, Ashley Chin-Baarstad

Open Access Dissertations

Individual differences in behavior can have potential fitness consequences and often reflect underlying genetic variation. My research focuses on three objectives related to individual level variation: 1) evaluating the innate behavioral variation within and between individuals, families, and progeny of different life-history types across time; 2) testing for differences in gene expression within the brain associated with this behavioral variation; and 3) using genetic polymorphisms to test for associations with ecotype, as well as population structure, in polymorphic populations. First, we evaluated the variation in a suite of ecologically relevant behaviors across time in juvenile progeny produced from crosses within …

Intranuclear Strain Measured By Iterative Warping In Cells Under Mechanical And Osmotic Stress, Jonathan T Henderson

Open Access Dissertations

The nucleus is a membrane bound organelle and regulation center for gene expression in the cell. Mechanical forces transfer to the nucleus directly and indirectly through specific cellular cytoskeletal structures and pathways. There is increasing evidence that the transferred forces to the nucleus orchestrate gene expression activity. Methods to characterize nuclear mechanics typically study isolated cells or cells embedded in 3D gel matrices. Often report only aspect ratio and volume changes, measures that oversimplify the inherent complexity of internal strain patterns. This presents technical challenges to simultaneously observe small scale nuclear mechanics and gene expression levels inside the nuclei of …

A Rhog/Ilk/Elmo2/Rac1 Pathways Modulates Microtubule Stability, Bradley C. Jackson

Electronic Thesis and Dissertation Repository

Integrin-linked kinase (ILK) is a widely expressed scaffold protein important for cell adhesion, migration, and phagocytosis. ILK and its interacting partner, engulfment and cell motility protein 2 (ELMO2), may also modulate microtubule dynamics. Here, I show that sustained microtubule growth and acetylated tubulin levels are reduced in the absence of ILK and, conversely, that exogenous ILK exerts a stabilizing effect on microtubules. Further, exogenous ELMO2, ras homology growth related protein (RHOG) and ras-related C3 botulinum toxin substrate 1 (RAC1) stabilize microtubules in an ILK-dependent manner. Potential downstream effectors of the effect of ILK on microtubule stability include glycogen synthase kinase- …

Role And Regulation Of Snon/Skil And Plscr1 Located At 3q26.2 And 3q23, Respectively, In Ovarian Cancer Pathophysiology, Madhav Karthik Kodigepalli

USF Tampa Graduate Theses and Dissertations

Ovarian cancer is one of the most common causes of gynecological cancer related deaths in women. In 2014, the estimated number of deaths due to ovarian cancer is 14,270 with occurrence of over 22, 240 new cases (National Cancer Institute, http://seer.cancer.gov/statfacts/html/ovary.html). Despite improvement in treatment strategies, the 5-year survival rate is still below 50% mainly due to chemoresistance and relapse. Amplification of chromosomal region 3q26 is a common characteristic in various epithelial cancers including ovarian cancer. This region harbors various oncogenes including the TGFβ signaling mediators EVI1 and SnoN/SkiL, PKCι and PIK3CA amplified at 3q26.2 and 3q26.3, respectively, in ovarian …

Transcriptional Regulation Of Sinorhizobium Meliloti Cell Cycle-Related Genes In The Δcbra Mutant And Root Nodules Of Medicago Sativa, Corey S. Hazekamp

Graduate Masters Theses

Sinorhizobium meliloti is a Gram-negative alphaproteobacterium and nitrogen-fixing symbiont, which undergoes a novel cell cycle modification during its' host-microbe interaction. I intend to monitor the transcriptional regulation of cell cycle-related genes during free-loving growth, in addition to monitoring their expression during symbiosis. Using genes known to be regulated by CtrA in C. crescentus or predicted to be regulated by CtrA in S. meliloti, I aim to show how certain cell cycle genes are regulated in S. meliloti. In C. crescentus, CtrA acts as a transcription factor that is active when phosphorylated and inactive when not phosphorylated. In …

Functional Analysis Of Moss Class Viii Myosin And Its Role In Plant Cell Division, Shu-Zon Wu

Doctoral Dissertations

The moss Physcomitrella patens is a great model system for studying plant gene function using reverse genetic approaches. It undergoes efficient gene-targeting by homologous recombination, allowing the generation of specific gene knockout and tagging a gene at its endogenous locus. Additionally, RNAi is quite effective in P. patens, providing an effective tool for rapid gene silencing and phenotypic characterization. Taking advantage of these features, this dissertation described the establishment of a system to perform an unbiased gene-by-gene RNAi assay to screen for tip growth phenotypes in P. patens. A small set of RNAi constructs were tested, within them …

The Roles Of Myosin Xi And Rop In Moss Tip Growth, Graham M. Burkart

Doctoral Dissertations

Because of the large number of myosin XI and ROP genes found in many angiosperms, it has been difficult to determine their precise role with respect to tip growth. In contrast, there are only two myosin XI genes in four ROP genes in the moss Physcomitrella patens. To determine their role in tip growth using a loss-of-function approach, I used RNA interference (RNAi) and found that both of these proteins are essential for tip growth. Consistent with a role in tip growth, I show that a functional, full-length fusion of mEGFP to myosin XI accumulates at a subcortical, apical …

The Role Of The Novel Lupus Antigen, Acheron, In Moderating Life And Death Decisions, Ankur Sheel

Masters Theses

Programmed cell death (PCD) is a major regulatory mechanism employed during development and homeostasis. The term PCD was coined to describe the death of the intersegmental muscles (ISMs) of moths at the end of metamorphosis. The timing of ISM death in the Tobacco Hawkmoth, Manduca sexta, is regulated by a fall in the titer of the steroid molting hormone 20-hydroxyecdysone (20E) late on day 17of pupal-adult development. This triggers the release of the peptide hormone, Eclosion Hormone (EH), which mediates its effects via the secondary messenger cGMP. It has been previously demonstrated that ISM death requires de novo gene expression. …

Rapid Method Of Processing Sperm For Nucleic Acid Extraction In Clinical Research, Matthew K. De Gannes

Masters Theses

Background: Sperm contain highly compact nuclei, inhibiting DNA extraction using traditional techniques. Current methods extracting sperm DNA involve lengthy lysis and no means of stabilizing DNA, hindering clinical research.

Objective: We sought to optimize an efficient method of extracting high quality human sperm DNA.

Methods: Sperm from three volunteers were isolated using PureCeption. We tested 1) proteinase K with DNA/RNA Shield, 2) DTT and TCEP as reducing agents, 3) QIAshredder homogenization, and 4) stability of sperm DNA fresh (baseline) or after 4 weeks of storage at 4^OC in DNA/RNA Shield using modified Quick-gDNA MiniPrep. DNA was PCR amplified …

Karst Estuaries: A Newly Described Ecosystem Governed By Aquifer Hydrology, Damian M. Menning

USF Tampa Graduate Theses and Dissertations

The overall goal of this dissertation is to define the hydrological, geochemical, and biological characteristics of a Karst Estuary. These types of estuaries represent a unique ecosystem created by freshwater inputs from direct flow through karst conduits and/or diffuse flow through a karst matrix. In order to determine the characteristics of a Karst Estuary we monitored short-term tidal fluctuations, long-term rainfall patterns, aquifer levels, spring discharge, multiple geochemical parameters, microbial communities in the water column and sediment, and macrofaunal communities in the sediment along a transect from a submarine spring through the Gulf of Mexico. Four sites were selected along …

Elucidating The Signalling Pathway Of Mer Tyrosine Kinase Receptor In Efferocytosis, Ekenedelichukwu Azu

Electronic Thesis and Dissertation Repository

Efferocytosis is the clearance of apoptotic cells and is necessary for homeostasis. Mer Tyrosine Kinase (MerTK) is a crucial efferocytic receptor whose loss is associated with chronic inflammatory diseases and autoimmunity. While previous studies have shown that MerTK mediates efferocytosis through a unique mechanism that requires integrins, MerTK signalling pathway remains unknown. Given this unusual internalization mechanism, I hypothesized that MerTK signals and engages integrins through a novel signalling pathway different from that used by other phagocytic receptors. Therefore, this study aimed to identify the signalling pathways activated by MerTK, utilizing conventional cell biology and pharmacological approaches.

I found that …

Serotonergic Signaling Pathways That Suppress Amyloid Beta In Mouse Models Of Ad, Jonathan Robert Fisher

All Theses and Dissertations (ETDs)

A diagnosis of Alzheimer's disease is one of the most devastating things one can hear. This terrible disease robs people of their ability to remember cherished events as their brains become riddled with beta amyloid plaques. Alzheimer's is especially terrifying because there currently are no effective treatments for slowing or stopping the disease. However, recent research has shown that plaque formation is correlated to concentrations of amyloid beta. This discovery suggests that limiting amyloid beta production could potentially halt the disease. One promising avenue for slowing amyloid beta production is serotonergic signaling.

This dissertation presents evidence for a direct sequence …

Characterization Of Efferosome Maturation And The Processing Of Apoptotic Bodies, Yohan Kim

Electronic Thesis and Dissertation Repository

Every day billions of cells in our bodies undergo apoptosis and are cleared through efferocytosis – a phagocytosis-like process in which phagocytes engulf and degrade apoptotic cells. Proper processing of efferosomes prevents inflammation and immunogenic presentation of antigens. In this thesis I determined that the early stages of efferosome maturation parallel that of the pro-immunogenic phagocytosis of pathogens. Mass spectrometry analysis of later maturation stages identified unique regulatory proteins on efferosomes and phagosomes. Keys among these were Rab17 and Rab45 on efferosomes versus Rab6b and PI-4-Kinase on phagosomes. The later would allow for antigen presentation from phagosomes, while the former …