Cell and Developmental Biology Commons^™

Using A Focus Measure To Automate The Location Of Biological Tissue Surfaces In Brightfield Microscopy, Daniel Toby Elozory

USF Tampa Graduate Theses and Dissertations

The study of microstructures in brightfield microscopy using unbiased stereology plays a large and growing role in bioscience research. Stereology enables objective quantitative analysis of biological structures within a tissue sample. A first step in the stereology process is to calculate the thickness of a tissue sample by locating the top and bottom surfaces of the sample. The aim of this project is to fully automate this location process by using the relative optical focus measure as an indicator of tissue surface boundary.

The current method for identification of focus bounding planes requires a trained user to manually select the …

Role Of Protein Kinase C-Iota In Neuroblastoma And The Effect Of Ica-1, A Novel Protein Kinase C-Iota Inhibitor On The Proliferation And Apoptosis Of Neuroblastoma Cells, Prajit P. Pillai

USF Tampa Graduate Theses and Dissertations

Protein Kinase C-iota (PKC-é), an atypical protein kinase C isoform manifests its potential as an oncogene by targeting various aspects of cancer cells such as growth, invasion and survival. PKC-é confers resistance to drug-induced apoptosis in cancer cells. The acquisition of drug resistance is a major obstacle to good prognosis in neuroblastoma. The focus of the dissertation was three-fold: First to study the role of PKC-é in the proliferation of neuroblastoma. Secondly, to identify the efficacy of [4-(5-amino-4-carbamoylimidazol-1-yl)-2,3-dihydroxycyclopentyl] methyl dihydrogen phosphate (ICA-1) as a novel PKC-é inhibitor in neuroblastoma cell proliferation and apoptosis. Finally, to analyze whether PKC-é could self-regulate …

An Upper Limit For Macromolecular Crowding Effects, Andrew C. Miklos, Congang Li, Courtney D. Sorell, L. Andrew Lyon, Gary J. Pielak

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Background: Solutions containing high macromolecule concentrations are predicted to affect a number of protein properties compared to those properties in dilute solution. In cells, these macromolecular crowders have a large range of sizes and can occupy 30% or more of the available volume. We chose to study the stability and ps-ns internal dynamics of a globular protein whose radius is similar to 2 nm when crowded by a synthetic microgel composed of poly(N-isopropylacrylamide-co-acrylic acid) with particle radii of similar to 300 nm.

Results: Our studies revealed no change in protein rotational or ps-ns backbone dynamics and only mild …

Biphasic Targeting And Cleavage Furrow Ingression Directed By The Tail Of A Myosin Ii, X. Fang, J. Y. Luo, R. Nishihama, C. Wloka, C. Dravis, M. Travaglia, M. Iwase, Elizabeth Ann Vallen, E. Bi

Biology Faculty Works

Cytokinesis in animal and fungal cells utilizes a contractile actomyosin ring (AMR). However, how myosin II is targeted to the division site and promotes AMR assembly, and how the AMR coordinates with membrane trafficking during cytokinesis, remains poorly understood. Here we show that Myo1 is a two-headed myosin II in Saccharomyces cerevisiae, and that Myo1 localizes to the division site via two distinct targeting signals in its tail that act sequentially during the cell cycle. Before cytokinesis, Myo1 localization depends on the septin-binding protein Bni5. During cytokinesis, Myo1 localization depends on the IQGAP Iqg1. We also show that the Myo1 …

Mosaic Analysis With Double Markers (Madm) As A Method To Map Cell Fates In Adult Mouse Taste Buds., Preston D. Moore

Electronic Theses and Dissertations

Taste buds are chemosensory endorgans embedded in the oral epithelium composed of cells that undergo continuous replacement. Mature taste cells live on average 10-14 days and are replaced by new cells when they die. However, the mechanism by which taste cells are produced and integrated into the taste bud as mature taste cells remains unknown. Previous studies approached this issue from either cell cycle gene expression properties or lineage tracing of precursor cells. In our study, we apply a new fate mapping technique that combines these two ideas. This technique, Mosaic Analysis with Double Markers, allows for simultaneous gene knockout …

The Roles Of Elevated Bcl-2 In Ovarian Cancer, Nicole Shree Anderson

USF Tampa Graduate Theses and Dissertations

Ovarian cancer (OC) is the second most common gynecologic cancer; however it is responsible for the most gynecologic cancer-related deaths. Apoptosis evasion is an important mechanism in OC tumorigenesis, and the prototypic anti-apoptotic protein, B-cell lymphoma 2 (Bcl-2), is often overexpressed in OC tumors. Gaining a better understanding of the mechanism(s) behind Bcl-2 overexpression and potential extra-anti-apoptotic functions of Bcl-2 could elucidate the importance of elevated Bcl-2 in OC. In the current study, I show through immunohistochemical analysis of normal, benign, and OC tissue sections, that both epithelial and stromal Bcl-2 expression decreases with OC progression. However, the number of …

Development Of A Genetic Modification System In Clostridium Scatologenes Atcc 25775 For Generation Of Mutants, Prasanna Tamarapu Parthasarathy

Masters Theses & Specialist Projects

3-Methyl indole (3-MI) is a malodorant in food and animal waste and Clostridium scatologenes ATCC 25775 is the model organism for the study of 3-MI production. 3-MI is an anaerobic degradation product of L-tryptophan and can cause pulmonary disorders and death in cattle and goats. To elucidate the 3-MI biosynthesis pathway and the underlying genes, it is necessary to develop a system to allow genetic modification in Clostridium scatologenes ATCC 25775. Bacteriophages and transposons are useful tools to achieve this goal. Isolation of Clostridium scatologenes ATCC 25775 bacteriophage was attempted by prophage induction and enrichments using environmental sources. To induce …

Analyzing The Role Of Αvβ8 Integrin In Glioma-Induced Angiogenesis, Jeremy H. Tchaicha

Dissertations and Theses (Open Access)

In this study we aimed to determine the functional roles for αvβ8 integrin in astrocytoma-induced angiogenesis. These studies originate from our analyses of αvβ8 integrin in developmental brain angiogenesis. αv and β8 knockout (KO) mice develop brain-specific vascular phenotypes that resemble vascular pathologies observed in the malignant astrocytoma, glioblastoma multiforme (GBM). Indeed, a murine xenograft model of astrocytoma suggested a role for the integrin in glioma-induced angiogenesis. Primary mouse astroglia were cultured from wild type (WT) and β8 KO neonates and were immortalized (HPV:E6/E7) and transformed (HRas:G12V). WT and β8 KO transformed astroglia were intracranially injected into athymic mice. WT …

Dynamic Remodeling Of The Stressed Heart: Role Of Protein Degradation Pathways, Deborah Vela

Dissertations and Theses (Open Access)

The heart is a remarkable organ. In order to maintain its function, it remodels in response to a variety of environmental stresses, including pressure overload, volume overload, mechanical or pharmacological unloading and hormonal or metabolic disturbances. All these responses are linked to the inherent capacity of the heart to rebuild itself. Particularly, cardiac pressure overload activates signaling pathways of both protein synthesis and degradation. While much is known about regulators of protein synthesis, little is known about regulators of protein degradation in hypertrophy. The ubiquitin-proteasome system (UPS) selectively degrades unused and abnormal intracellular proteins. I speculated that the UPS may …

Mcam/Muc18 Regulates Melanoma Progression By Modulating The Expression Of Id-1, Maya Zigler

Dissertations and Theses (Open Access)

The acquisition of the metastatic melanoma phenotype is associated with increased expression of the melanoma cell adhesion molecule MCAM/MUC18 (CD146). However, the mechanism by which MUC18 contributes to melanoma metastasis remains unclear. Herein, we stably silenced MUC18 expression utilizing lentivirus-incorporated small hairpin RNA, in two metastatic melanoma cell lines, A375SM and C8161, and conducted cDNA microarray analysis. We identified and validated that the transcriptional regulator, Inhibitor of DNA Binding-1 (Id-1), previously shown to function as an oncogene in several malignancies, was downregulated by 5.6-fold following MUC18 silencing. Additionally, we found that MUC18 regulated Id-1 expression at the transcriptional level via …

Production, Purification And Crystallization Of Membrane Integrated Multimeric Bax, Adelbert Mark Villoso

UNLV Theses, Dissertations, Professional Papers, and Capstones

Apoptosis, or programmed cell death, is a vital process intimately involved in the embryonic development and homeostatic maintenance of all multicellular organisms. The committing step to apoptosis is regulated by a key protein, Bax, and its ability to integrate and form a pore structure at the outer mitochondrial membrane.

Unfortunately, the molecular details of apoptosis remain largely unclear due to the lack of structural data of integral membrane (IM) Bax. Experimental limitations of membrane protein production have slowed the pursuit of an IM-Bax structure simply because standard protocols for producing recombinant IM-Bax are inefficient in producing adequate quantities of IM-Bax …

Role Of Alpha 7 Nicotinic Acetylcholine Receptor In Calcium Signaling Induced By Prion Protein Interaction With Stress-Inducible Protein 1, Flavio H. Beraldo, Camila P. Arantes, Tiago G. Santos, Nicolle G. T. Queiroz, Kirk Young, Jane R. Rylett, Regina P. Markus, Marco A. M. Prado, Vilma R. Martins

Anatomy and Cell Biology Publications

The prion protein (PrP(C)) is a conserved glycosylphosphatidyl-inositol-anchored cell surface protein expressed by neurons and other cells. Stress-inducible protein 1 (STI1) binds PrP(C) extracellularly, and this activated signaling complex promotes neuronal differentiation and neuroprotection via the extracellular signal-regulated kinase 1 and 2 (ERK1/2) and cAMP-dependent protein kinase 1 (PKA) pathways. However, the mechanism by which the PrPC-STI1 interaction transduces extracellular signals to the intracellular environment is unknown. We found that in hippocampal neurons, STI1-PrP(C) engagement induces an increase in intracellular Ca(2+) levels. This effect was not detected in PrP(C)-null neurons or wild-type neurons treated with an STI1 mutant unable to …

Proteoliposome Proton Flux Assays Establish Net Conductance, Ph-Sensitivity, And Functional Integrity Of A Novel Truncate Of The M2 Ion "Channel" Of Influenza A, Emily Peterson

Theses and Dissertations

A novel truncate of Influenza A M2 protein (residues 22-62), incorporated into a uniquely tailored proteoliposome proton uptake assay, demonstrated proton flux more characteristic of an ion transporter than a traditional ion "channel." The liposome paradigm was essential for testing the conductance activity of this M2 truncate at a range of extraphysiological pHs appropriate for channel vs. transport function determination. In addition to transporter-typical proton flux, M2(22-62) showed the key characteristics of functional integrity: selective proton uptake into liposomes and block of uptake by amantadine. Two sets of proteoliposome proton flux assays were carried out, Set 1 at pH values …

Characterization Of The Lkb1-Mo25-Strad Ampkk Complex In Adult Mouse Skeletal Muscle, Cody Don Smith

Theses and Dissertations

In liver tissue, the AMP-activated protein kinase kinase (AMPKK) complex was identified as the association of LKB1, MO25α/β, and STRADα/β proteins; however, this complex has yet to be characterized in skeletal muscle. In this report, we demonstrate the expression of the LKB1-MO25-STRAD AMPKK complex in adult skeletal muscle, confirm the absence of mRNA splice variants, and report the relative mRNA expression levels of these complex-forming proteins. To facilitate this characterization we used control (ctrl) and muscle-specific LKB1 knockout (LKB1-/-) mice. LKB1 detection in untreated ctrl and LKB1-/- muscle lysates revealed two protein bands at approximately 50 and 60 kDa; although, …

Virulence Of An Emerging Respiratory Pathogen, Genus Pandoraea, In Vivo And Its Interactions With Lung Epithelial Cells, Gillian Herbert, Anne Costello, Lydia Fabunmi, Kirsten Schaffer, Kevin Kavanagh, Emma M. Caraher, Máire Callaghan, Siobhan Mcclean

Articles

Pandoraea species have emerged as opportunistic pathogens among cystic fibrosis (CF) and non-CF patients. Pandoraea pulmonicola is the predominant Pandoraea species among Irish CF patients. The objective of this study was to investigate the pathogenicity and potential mechanisms of virulence of Irish P. pulmonicola isolates and strains from other Pandoraea species. Three patients from whom the P. pulmonicola isolates were isolated have since died. The in vivo virulence of these and other Pandoraea strains was examined by determining the ability to kill Galleria mellonella larvae. The P. pulmonicola strains generally were the most virulent of the species tested, with three …

Cyanidin-3-Glucoside Inhibits Ethanol-Induced Invasion Of Breast Cancer Cells Overexpressing Erbb2, Mei Xu, Kimberly A. Bower, Siying Wang, Jacqueline A. Frank, Gang Chen, Min Ding, Shiow Wang, Xianglin Shi, Zunji Ke, Jia Luo

Internal Medicine Faculty Publications

BACKGROUND: Ethanol is a tumor promoter. Both epidemiological and experimental studies suggest that ethanol may enhance the metastasis of breast cancer cells. We have previously demonstrated that ethanol increased the migration/invasion of breast cancer cells expressing high levels of ErbB2. Amplification of ErbB2 is found in 20-30% of breast cancer patients and is associated with poor prognosis. We sought to identify agents that can prevent or ameliorate ethanol-induced invasion of breast cancer cells. Cyanidin-3-glucoside (C3G), an anthocyanin present in many vegetables and fruits, is a potent natural antioxidant. Ethanol exposure causes the accumulation of intracellular reactive oxygen species (ROS). This …

Quantitative Trait Loci Mapping Reveals Candidate Pathways Regulating Cell Cycle Duration In Plasmodium Falciparum, Heather B. Reilly Ayala, Mark A. Wacker, Geoffrey Siwo, Michael T. Ferdig

Faculty Publications - Department of Biological & Molecular Science

Background: Elevated parasite biomass in the human red blood cells can lead to increased malaria morbidity. The genes and mechanisms regulating growth and development of Plasmodium falciparum through its erythrocytic cycle are not well understood. We previously showed that strains HB3 and Dd2 diverge in their proliferation rates, and here use quantitative trait loci mapping in 34 progeny from a cross between these parent clones along with integrative bioinformatics to identify genetic loci and candidate genes that control divergences in cell cycle duration. Results: Genetic mapping of cell cycle duration revealed a four-locus genetic model, including a major genetic effect …

Molecular Mechanism Of Agc Kinases In Human Malignant, Shaokun Shu

USF Tampa Graduate Theses and Dissertations

The maintenance of normal cell function and tissue homeostasis is dependent on the precise regulation of multiple signaling pathways that control cellular decisions to either proliferate, differentiate, arrest cell growth, or initiate programmed cell death (apoptosis). Cancer arises when clones of mutated cells escape this balance and proliferate inappropriately without compensatory apoptosis. Deregulated cell growth occurs as a result of perturbed signal transduction that modulates or alters cellular behavior or function to keep the critical balance between the rate of cell-cycle progression (cell division) and cell growth (cell mass) on one hand, and programmed cell death (apoptosis, autophagy) on the …

Ruthenium Complexes As Dna Photonucleases In Treatment Of Malignant Skin Carcinoma, Devi Kalyan Karumanchi

All Capstone Projects

A variety of Ruthenium complexes possessing terpyridine complexes as ligands in their coordination sphere have been investigated extensively as DNA photo nucleases in vitro. This work has led to the realization that compounds of Ruthenium bind to DNA purines and that the interactions are of an unprecedented bridging variety. Owing to the molar absorptivity of these complexes in the visible region, direct cleavage of duplex DNA by these complexes requires irradiation in the range of 400-500 nm.

Although the observation of DNA photo cleavage in vitro makes these systems good candidates for further investigation, it does not provide any information …

Characterization Of Metastasis-Associated Cell Surface Glycoproteins In Prostate Cancer, Lifang Yang

Theses and Dissertations in Biomedical Sciences

Prostate cancer (PCa) is a major health problem in males in the United States. Its lethality is mostly attributed to the primary tumor metastasizing to distant sites that are highly resistant to conventional therapies. Serum Prostate Specific Antigen (PSA) is the only protein biomarker used in clinic for prediction of prostate cancer recurrence following local therapies. Nonetheless, PSA lacks the ability to predict the behavior of an individual tumor in an individual patient. Therefore, development of reliable biomarkers for detection of metastatic potential in primary tumors, as well as discovery of new therapeutic targets, is in a great need for …