Biochemistry, Biophysics, and Structural Biology Commons^™

The Replication Initiator Of The Cholera Pathogen’S Second Chromosome Shows Structural Similarity To Plasmid Initiators, Natalia Orlova, Matthew Gerding, Olha Ivashkiv, Paul Dominic B. Olinares, Brian T. Chait, Matthew K. Waldor, David Jeruzalmi

Publications and Research

The conserved DnaA-oriC system is used to initiate replication of primary chromosomes throughout the bacterial kingdom; however, bacteria with multipartite genomes evolved distinct systems to initiate replication of secondary chromosomes. In the cholera pathogen, Vibrio cholerae, and in related species, secondary chromosome replication requires the RctB initiator protein. Here, we show that RctB consists of four domains. The structure of its central two domains resembles that of several plasmid replication initiators. RctB contains at least three DNA binding winged-helix-turn-helix motifs, and mutations within any of these severely compromise biological activity. In the structure, RctB adopts a headto- head dimeric configuration …

Defining The Role Of Phosphorylation And Dephosphorylation In The Regulation Of Gap Junction Proteins, Hanjun Li

Theses & Dissertations

Gap junctions are intercellular channels that permit the free passage of ions, small metabolites, and signaling molecules between neighboring cells. In the diseased human heart, altered ventricular gap junction organization and connexin expression (i.e., remodeling) are key contributors to rhythm disturbances and contractile dysfunction. Connexin43 (Cx43) is the dominant gap junction protein isoform in the ventricle which is under tight regulation by serine/tyrosine phosphorylation. Phosphorylation and dephosphorylation regulate many aspects of Cx43 function including trafficking, assembly and disassembly, electrical and metabolic coupling at the plaque, as well as to modulate the interaction with other proteins.

Serine phosphorylation has long been …

Regulation Of Alteration/Deficiency In Activation 3 (Ada3) By Acetylation And Its Role In Cell Cycle Regulation And Oncogenesis, Shashank Srivastava

Theses & Dissertations

The ADA3 (Alteration/Deficiency in Activation 3) protein is a transcriptional adaptor protein that was initially discovered as a component of several HAT (Histone Acetyltransferase) complexes, the enzyme complex responsible for histone acetylation, which is a prerequisite for transcription. Earlier the studies from Dr. Band’s laboratory and that of others’ have deciphered a crucial role of ADA3 in cell cycle regulation (both through G₁/S and G₂/M phase transitions) and in maintaining the genomic stability.

While our laboratory investigated the mechanism behind the role of ADA3 in G₁/S transition, the same remained unknown for G_{2 …}

Mitogen And Morphogen Signaling Dysregulation: Pathophysiological Influence In Pancreatic Cancer And Alzheimer’S Disease, Eric Cruz

Theses & Dissertations

Although the etiology of a particular disease will vary, there are genetic and epigenetic bottlenecks that frequently converge resulting in dysregulation of mitogenic and morphogenetic signaling. This propensity is acutely experienced in malignancy and neurodegenerative disease.

Here, we have first investigated the role of dysregulated signaling in the context of pancreatic cancer (PC). Morphogenetic signaling has been regarded as a pleiotropic pathway with the potential to promote and inhibit metastatic features. Our investigation of bone morphogenetic protein 2 (BMP-2), an archetypical member of the BMP superfamily, has revealed the presence of extracellular, intracellular, and long non-coding RNA products. Our findings …

The Association Between Oxidative Stress, Cellular Differentiation And Galectins In Human Promyelocytic Leukemia Cells (Hl-60), James R. Vinnai

Electronic Thesis and Dissertation Repository

Galectins are a group of β-galactoside-binding proteins involved in different cellular processes including stress responses and differentiation. The role and expression of galectins under oxidative stress and during neutrophilic differentiation was examined in HL-60 cells. Galectin gene (LGALS), and galectin protein expression were determined using RT-qPCR and immunoblotting, respectively. Neutrophilic differentiation was measured via a spectrofluorometric assay. DNA methylation and JNK signaling were investigated as galectin regulatory mechanisms. Menadione-induced oxidative stress, DMSO-induced differentiation, DNA hypomethylation and JNK signaling all promoted similar galectin expression profiles. Antioxidant N-acetylcysteine attenuated the menadione-induced galectin expression but only partially attenuated DMSO-induced galectin expression. …

Jmh Dissertation 2016.Pdf, Jennifer Hayashi

Jennifer Hayashi

Pharmacokinetics, Tissue Distribution, Synergistic Activity, And Antitumor Activity Of Two Isomeric Flavones, Crystal L. Whitted

Electronic Theses and Dissertations

Flavonoids are polyphenolic secondary metabolites found in plants that have bioactive properties including antiviral, antioxidant, and anticancer. Two isomeric flavone were extracted from Gnaphalium elegans and Achyrocline bogotensis, plants used by the people from the Andean region of South America as remedies for cancer. 5,7-dihydroxy-3,6,8-trimethoxy-2-phenyl-4H-chromen-4-one (5, 7–dihydroxy- 3, 6, 8 trimethoxy flavone/ flavone A) and 3,5-dihydroxy-6,7,8-trimethoxy-2-phenyl-4H-chromen-4-one (3, 5–dihydroxy-6, 7, 8–trimethoxy flavone/ flavone B) have shown antineoplastic activity against colon cancer cell lines dependent upon their differentiation status. Pharmacokinetic studies reported herein were used to determine dosing for antitumor assays, as well as determine target tissue concentration. These included the …

Investigating The Effects Of Ph On Alphaviral E3-E2 Glycoprotein Association, Organization, And Cellular Tropism, Jason Michael Sequra

Open Access Dissertations

In alphaviruses the role of E3 is required in protecting the fusion peptide region of E1 during intracellular transport. Throughout viral processing, the association of E2 and E3 is required for the successful trafficking and incorporation of E1 into the mature virion. This E3-E2 association has been observed to extend to mature virions in the solved structure for the envelope of Semliki Forest virus (SFV) and supported by the solved structure for the entire Venezuelan equine encephalitis virion (VEEV) with exclusive contacts being made between E3-E2. Immunization with monoclonal antibodies against VEEV E3 provided protection for mice challenged by lethal …

Characterization Of Vesicular Monoamine Transporter 2 And Its Role In Parkinson's Disease Pathogenesis Using Drosophila, Antonio Joel Tito Jr., Sheng Zhang

Dissertations & Theses (Open Access)

Parkinson’s disease (PD) is a progressive neurodegenerative disorder caused by the selective loss of the dopaminergic neurons in the Substantia nigra pars compacta region of the brain. PD is also the most common neurodegenerative disorder and the second most common movement disorder. PD patients exhibit the cardinal symptoms, including tremor of the extremities, rigidity, slowness of movement, and postural instability, after 70-80% of DA neurons degenerate. It is, therefore, imperative to elucidate the underlying mechanisms involved in the selective degeneration of DA neurons. Although increasing numbers of PD genes have been identified, why these largely widely expressed genes induce …

Punctuated Evolution Within A Eurythermic Genus (Mesenchytraeus) Of Segmented Worms: Genetic Modification Of The Glacier Ice Worm F1f0 Atp Synthase, Shirley A. Lang

Graduate School of Biomedical Sciences Theses and Dissertations

Segmented worms (Annelida) are among the most successful animal inhabitants of extreme environments worldwide. An unusual group of Mesenchytraeus worms endemic to the Pacific Northwest of North America occupy geographically proximal ecozones ranging from low elevation temperate rainforests to high altitude glaciers. Along this altitudinal transect, Mesenchytraeus representatives from disparate habitat types were collected and subjected to deep mitochondrial and nuclear phylogenetic analyses. Evidence presented here employing modern bioinformatic analyses (i.e., maximum likelihood, Bayesian inference, multi-species coalescent) supports a Mesenchytraeus “explosion” in the upper Miocene (5-10 million years ago) that gave rise to ice, snow and terrestrial worms, derived from …

Engineered Human Acidic Fibroblast Growth Factor (Fgf1) With An Enhanced Thermal And Proteolytic Stability, Duaa Abdullah Almansaf

Graduate Theses and Dissertations

Fibroblast growth factor receptor (FGFR) is made up of three significant domains. The most important domain is the intracellular domain where the dimerization and autophosphorylation occur. Fibroblast growth factor (FGF) interacts with specific FGFR to regulate many cellular processes during the embryonic stage. Furthermore, FGF is significant for adults because FGF plays an important role in regulating cellular differentiation as well as wound healing. The cellular regulating processes are initiated through binding FGF to heparin followed by binding FGF/heparin to FGFR to form FGF/heparin/FGFR complex. Thus, FGFR is dimerized and autophosphorylated. The phosphorylation of FGFR triggers downstream signaling pathways, which …

Consensus Paper: Cerebellar Development., K Leto, M Arancillo, Ebe Becker, A Chiang, Et Al.

Faculty Publications

The development of the mammalian cerebellum is orchestrated by both cell-autonomous programs and inductive environmental influences. Here, we describe the main processes of cerebellar ontogenesis, highlighting the neurogenic strategies used by developing progenitors, the genetic programs involved in cell fate specification, the progressive changes of structural organization, and some of the better-known abnormalities associated with developmental disorders of the cerebellum.

Influence Of Ph And Acidic Side Chain Charges On The Behavior Of Designed Model Peptides In Lipid Bilayer Membranes, Venkatesan Rajagopalan

Graduate Theses and Dissertations

The molecular properties of transmembrane proteins and their interactions with lipids regulate biological function. Of particular interest are interfacial aromatic residues and charged residues in the core helix whose functions range from stabilizing the native structure to regulating ion channels. This dissertation addresses the pH dependence and influence of potentially negatively charged tyrosine, glutamic acid or aspartic acid side chains. We have employed GWALP23 (acetyl-GGALW5LALALALALALALW19LAGA-amide) as favorable host peptide framework. We have substituted W5 with Tyr (Y5GWALP23) and Leu residues with Glu (L12E, L14E or L16E) or Asp (L14D or L16D), and have incorporated specific 2H-labeled alanine residues within the …

Molecular And Functional Analysis Of The Pixb Gene In Xenorhabdus Nematophila, John Lucas

Theses and Dissertations

Xenorhabdus nematophila and the entomopathogenic nematode Steinernema carpocapsae form a mutualistic relationship facilitating the infection, death and consumption of an insect host. The infective juvenile (IJ) form of S. carpocapsae invades the insect host through natural openings and proceeds to the hemocoel where exposure to hemolymph stimulates the release of X. nematophila from the anterior vesicle. Excreted X. nematophila releases immunosuppressive compounds and insect toxins into the insect hemolymph that facilitates death of the host. As X. nematophila reaches high cell density it secretes exoenzymes that degrade insect tissues and produces antibiotics that reduce microbial competition. S. carpocapsae utilizes the …

Rna Sequencing Analysis Of The Developing Chicken Retina, Christophe Langouet-Astrie*, Annamarie Meinsen*, Emily R. Grunwald*, Stephen Turner, Raymond A. Enke

Ray Enke Ph.D.

Pt-Mal-Lhrh Mediates Breast Cancer Cell Cytotoxicity Through Increased Apoptosis, Kendall E. Collins

Posters-at-the-Capitol

In the United States one in eight women will be afflicted with breast cancer. It is estimated that in 2016 there will be approximately 246,600 new invasive breast cancer cases and 61,000 new non-invasive cases. Triple negative breast cancers account for 15% of all breast cancers and are significantly more aggressive than other subtypes. Treatment options for triple negative breast cancer are limited due to the cancers not expressing the estrogen, progestogen, or herceptin receptors making them unresponsive to hormonal therapy. Our recent work centers around developing a novel chemotherapeutic agent that will direct therapy selectively to triple negative (4T1) …

Understanding The Differences Between Neuronal Calcium Sensor Proteins: A Comparison Of Neurocalcin Delta And Hippocalcin, Jeffrey M. Viviano

Graduate School of Biomedical Sciences Theses and Dissertations

Many neuronal functions, including learning and memory are driven by changes in intracellular Ca2+concentrations. The Neuronal Calcium Sensor (NCS) family of proteins is responsible for mediating the response to calcium. They are typically comprised of 4 EF hands; of which EF 2, 3, and 4 bind calcium.

Hypothesis: NCS proteins carry out unique, non-overlapping functions, and that specific characteristics of the family can be mapped to precise regions of the proteins.

Results: The effect on the following properties were investigated primarily on two highly similar NCS proteins, Neurocalcin Delta (NCALD) and Hippocalcin (HPCA): (1) Response to calcium was determined through …

Pleckstrin Homology (Ph) Domain Leucine-Rich Repeat Protein Phosphatase Controls Cell Polarity By Negatively Regulating The Activity Of Atypical Protein Kinase C, Xiaopeng Xiong, Xin Li, Yang-An Wen, Tianyan Gao

Markey Cancer Center Faculty Publications

The proper establishment of epithelial polarity allows cells to sense and respond to signals that arise from the microenvironment in a spatiotemporally controlled manner. Atypical PKCs (aPKCs) are implicated as key regulators of epithelial polarity. However, the molecular mechanism underlying the negative regulation of aPKCs remains largely unknown. In this study, we demonstrated that PH domain leucine-rich repeat protein phosphatase (PHLPP), a novel family of Ser/Thr protein phosphatases, plays an important role in regulating epithelial polarity by controlling the phosphorylation of both aPKC isoforms. Altered expression of PHLPP1 or PHLPP2 disrupted polarization of Caco2 cells grown in 3D cell cultures …

An Arginine Finger Regulates The Sequential Action Of Asymmetrical Hexameric Atpase In The Double-Stranded Dna Translocation Motor, Zhengyi Zhao, Gian Marco De-Donatis, Chad T. Schwartz, Huaming Fang, Jingyuan Li, Peixuan Guo

Pharmaceutical Sciences Faculty Publications

Biological motors are ubiquitous in living systems. Currently, how the motor components coordinate the unidirectional motion is elusive in most cases. Here, we report that the sequential action of the ATPase ring in the DNA packaging motor of bacteriophage ϕ29 is regulated by an arginine finger that extends from one ATPase subunit to the adjacent unit to promote noncovalent dimer formation. Mutation of the arginine finger resulted in the interruption of ATPase oligomerization, ATP binding/hydrolysis, and DNA translocation. Dimer formation reappeared when arginine mutants were mixed with other ATPase subunits that can offer the arginine to promote their interaction. Ultracentrifugation …

Molecular Analysis Of Ftsz-Ring Assembly In E. Coli Cytokinesis, Kuo-Hsiang Huang

Dissertations, Theses, and Capstone Projects

An essential first step in bacterial division is the assembly of a cytokinetic ring (Z-ring) formed by the tubulin-like FtsZ at midcell. The highly conserved core domain of FtsZ has been reported to mediate assembly of FtsZ polymers in vivo and in vitro. Species-specific differences in the FtsZ C-terminal domain such as the FtsZ CTV region and interactions with several modulatory proteins such as ZapC and ZapD, restricted to certain bacterial classes, also serve as key determinants of FtsZ protofilament bundling. Here, we characterize (i) the roles of the FtsZ CTV region in mediating both longitudinal and lateral interactions …

Characterization Of The Catalytic Ck2 Subunits With Substitutions At Residues Involved In Inhibitor Binding, Paul Desormeaux

Electronic Thesis and Dissertation Repository

CK2 is a constitutively active, ubiquitously expressed and pleiotropic serine/threonine protein kinase that is implicated in many cellular functions including tumorigenesis. CK2 has two catalytic subunits, CK2a and CK2a’, that carry out its function in the cell. Previous studies have indicated that inhibitor-refractory mutants have been effective in recovering residual CK2 activity, in the presence of inhibitors, when compared to wild type CK2. Based on these observations, inhibitor-refractory mutants were created for both CK2a and CK2a’ and tested with various concentrations with two CK2-specific inhibitors, CX-4945 and inhibitor VIII. The CK2a triple mutant (V66A/I174A/H160D) was tested in inducible U2OS Flp-In …

Experimental Infection Of Conventional Nursing Pigs And Their Dams With Porcine Deltacoronavirus, Sarah Vitosh-Sillman, John Dustin Loy, Bruce W. Brodersen, Clayton L. Kelling, Alan R. Doster, Christina L. Topliff, Eric A. Nelson, Jianfa Bai, Erin Schirtzinger, Elizabeth Poulsen, Barbara Meadors, Joseph Anderson, Benjamin Hause, Gary Anderson, Richard Hesse

School of Veterinary and Biomedical Sciences: Faculty Publications

Porcine deltacoronavirus (PDCoV) is a newly identified virus that has been detected in swine herds of North America associated with enteric disease. The aim of this study was to demonstrate the pathogenicity, course of infection, virus kinetics, and aerosol transmission of PDCoV using 87 conventional piglets and their 9 dams, including aerosol and contact controls to emulate field conditions. Piglets 2–4 days of age and their dams were administered an oronasal PDCoV inoculum with a quantitative real-time reverse transcription (qRT)-PCR quantification cycle (Cq) value of 22 that was generated from a field sample having 100% nucleotide identity to USA/Illinois121/2014 determined …

Regulation Of Nachrs And Stemness By Nicotine And E-Cigarettes In Nsclc, Courtney Schaal

USF Tampa Graduate Theses and Dissertations

Lung cancer is the leading cause of cancer-related death in both men and women, nationally and internationally and kills more people each year than breast, prostate, and colon cancers combined. Non-small cell lung carcinoma (NSCLC) is the most common histological subtype of lung cancer, and accounts for 85% of all cases. Cigarette smoking is the single greatest risk factor for lung cancer, and is correlated with 80-90% of all lung cancer deaths. Nicotine, the addictive component of tobacco smoke, is not a carcinogen and cannot initiate tumors itself; however, it is known to act as a tumor promoter, by enhancing …

Role Of Ddr1 In Pancreatic Cancer, Huocong Huang

Theses & Dissertations

Pancreatic ductal adenocarcinomas are highly malignant cancers, characterized by extensive invasion into surrounding tissues, metastasis to distant organs at a very early stage, and a limited response to therapy. One of the main features of pancreatic ductal adenocarcinomas is desmoplasia, which leads to extensive deposition of collagen I. We have demonstrated that collagen I can induce epithelial-mesenchymal transition (EMT) in pancreatic cancer cells. A hallmark of EMT is an increase in the expression of a mesenchymal cadherin, N-cadherin. Our previous studies have shown that up-regulation of N-cadherin can promote tumor cell invasion and that collagen I-induced EMT is through two …

Modeling And Analysis Of Germ Layer Formations Using Finite Dynamical Systems, Alexander Garza, Megan Eberle, Eric A. Eager

Spora: A Journal of Biomathematics

The development of an embryo from a fertilised egg to a multicellular organism proceeds through numerous steps, with the formation of the three germ layers (endoderm, mesoderm, ectoderm) being one of the first. In this paper we study the mesendoderm (the tissue that collectively gives rise to both mesoderm and endoderm) gene regulatory network for two species, \textit{Xenopus laevis} and the axolotl (\textit{Ambystoma mexicanum}) using Boolean networks. We find that previously-established bistability found in these networks can be reproduced using this Boolean framework, provided that some assumptions used in previously-published differential equations models are relaxed. We conclude by discussing our …

Cal And Magi Pdz Protein Regulation Of Crfr1 And 5-Ht2ar Trafficking And Signaling, Maha Mahmoud Hammad

Electronic Thesis and Dissertation Repository

PDZ (PSD95/Disc Large/Zona Occludens) domain-containing proteins are scaffolding proteins that play important roles in regulating the activity of G protein-coupled receptors. Corticotropin Releasing Factor Receptor 1 (CRFR1) and Serotonin 2A Receptor (5-HT_2AR) are two GPCRs that are commonly associated with mental disorders. Both receptors also contain a class I PDZ-binding motif at the carboxyl terminal tail. In the first chapter, we investigate the effects of CAL (CFTR-associated ligand) on regulating the trafficking and signaling of CRFR1. We demonstrate a role for CAL in inhibiting CRFR1 endocytosis, cell surface expression, and CRF-mediated ERK1/2 signaling via the CRFR1 PDZ-binding motif. …

Nanobubbles Provide Theranostic Relief To Cancer Hypoxia, Christopher M. Long, Pushpak N. Bhandari, Joseph Irudayaraj

The Summer Undergraduate Research Fellowship (SURF) Symposium

Hypoxia is a common motif among tumors, contributing to metastasis, angiogenesis, cellular epigenetic abnormality, and resistance to cancer therapy. Hypoxia also plays a pivotal role in oncological studies, where it can be used as a principal target for new anti-cancer therapeutic methods. Oxygen nanobubbles were designed in an effort to target the hypoxic tumor regions, thus interrupting the hypoxia-inducible factor-1α (HIF-1α) regulatory pathway and inhibiting tumor progression. At less than 100nm, oxygen nanobubbles act as a vehicle for site-specific oxygen delivery, while also serving as an ultrasound contrast agent for advanced imaging purposes. Through in vitro and in vivo studies, …

P-Rex1 Promotes Resistance To Vegf/Vegfr-Targeted Therapy In Prostate Cancer, Hira Lal Goel, Bryan M. Pursell, Leonard D. Shultz, Dale L. Greiner, Rolf A. Brekken, Craig W. Vander Kooi, Arthur M. Mercurio

Arthur M. Mercurio

Autocrine VEGF signaling is critical for sustaining prostate and other cancer stem cells (CSCs), and it is a potential therapeutic target, but we observed that CSCs isolated from prostate tumors are resistant to anti-VEGF (bevacizumab) and anti-VEGFR (sunitinib) therapy. Intriguingly, resistance is mediated by VEGF/neuropilin signaling, which is not inhibited by bevacizumab and sunitinib, and it involves the induction of P-Rex1, a Rac GEF, and consequent Rac1-mediated ERK activation. This induction of P-Rex1 is dependent on Myc. CSCs isolated from the PTEN(pc-/-) transgenic model of prostate cancer exhibit Rac1-dependent resistance to bevacizumab. Rac1 inhibition or P-Rex1 downregulation increases the sensitivity …

Inhibition Of Nuclear Factor-Kappa B Enhances The Tumor Growth Of Ovarian Cancer Cell Line Derived From A Low-Grade Papillary Serous Carcinoma In P53-Independent Pathway, Xue Xiao, Gong Yang, Peng Bai, Shunping Gui, Tri M. Bui Nguyen, +8 Additional Authors

Biochemistry and Molecular Medicine Faculty Publications

Background: NF-kB can function as an oncogene or tumor suppressor depending on cancer types. The role of NF-kB in low-grade serous ovarian cancer, however, has never been tested. We sought to elucidate the function of NF-kB in the low-grade serous ovarian cancer.

Methods: The ovarian cancer cell line, HOC-7, derived from a low-grade papillary serous carcinoma. Introduction of a dominant negative mutant, IkBαM, which resulted in decrease of NF-kB function in ovarian cancer cell lines. The transcription ability, tumorigenesis, cell proliferation and apoptosis were observed in derivative cell lines in comparison with parental cells.

Results: Western blot analysis indicated increased …

Intestinal Cytoplasmic Lipid Droplets, Associated Proteins, And The Regulation Of Dietary Fat Absorption, Theresa M. D'Aquila

Open Access Dissertations

Dietary fat provides essential nutrients, contributes to energy balance, and regulates blood lipid concentrations. These functions are important to health, but can also become dysregulated and contribute to diseases such as obesity, diabetes, and cardiovascular disease. The small intestine absorbs dietary fat through an efficient multi step process of digestion, uptake, metabolism, and secretion or storage. When dietary fat is taken up by the absorptive cells of the small intestine, enterocytes, it can be secreted into circulation where it contributes to blood lipid levels or temporarily stored in cytoplasmic lipid droplets (CLDs). The objective of this dissertation is to investigate …