Cell and Developmental Biology Commons^™

Divergent Mechanisms Of Interaction Of Helicobacter Pylori And Campylobacter Jejuni With Mucus And Mucins, Julie Ann Naughton, Karina Mariño, Brendan Dolan, Colm Reid, Ronan Gough, Mary Gallagher, Michelle Kilcoyne, Jared Gerlachscience,, Lokesh Joshi, Pauline Rudd, Stephen Carrington, Billy Bourke, Marguerite Clyne

Articles

Helicobacter pylori and Campylobacter jejuni colonize the stomach and intestinal mucus, respectively. Using a combination of mucus-secreting cells, purified mucins, and a novel mucin microarray platform, we examined the interactions of these two organisms with mucus and mucins. H. pylori and C. jejuni bound to distinctly different mucins. C. jejuni displayed a striking tropism for chicken gastrointestinal mucins compared to mucins from other animals and preferentially bound mucins from specific avian intestinal sites (in order of descending preference: the large intestine, proximal small intestine, and cecum). H. pylori bound to a number of animal mucins, including porcine stomach mucin, but …

Proteomic And Biochemical Studies Of Estrogen-Mediated Signaling And Novel Estrogen Receptor-Interacting Proteins In Breast Cancer Cells, Zhenqi Zhou

Graduate Theses and Dissertations

Estrogen plays essential roles in the growth, development, and homeostasis of a number of tissues, and can also be linked to the growth of breast cancer. The biological activities of estrogen are mediated by estrogen receptors (ERs) ERá and ERâ, and also orphan G-protein-coupled receptor 30 (GPR30). In order to identify novel proteins that are involved in ER-mediated actions of estrogen, we used mass spectrometry-based quantitative proteomic methods to systematically profile global protein expression in responses to E2 (17â-estradiol) stimulation in human breast cancer cell, and identify and characterize cellular novel proteins that are associated with ERs in breast cancer …

Heterogeneous Nuclear Ribonucleoprotein K Supports Vesicular Stomatitis Virus Replication By Regulating Cell Survival And Cellular Gene Expression, Phat X. Dinh, Anshuman Das, Rodrigo Franco, Asit K. Pattnaik

School of Veterinary and Biomedical Sciences: Faculty Publications

The heterogeneous nuclear ribonucleoprotein K (hnRNP K) is a member of the family of hnRNPs and was recently shown in a genome-wide small interfering RNA (siRNA) screen to support vesicular stomatitis virus (VSV) growth. To decipher the role of hnRNP K in VSV infection, we conducted studies which suggest that the protein is required for VSV spreading. Virus binding to cells, entry, and nucleocapsid uncoating steps were not adversely affected in the absence of hnRNP K, whereas viral genome transcription and replication were reduced slightly. These results indicate that hnRNP K is likely involved in virus assembly and/or release from …

New Types Of Morpho-Physiological Changes In Cells Exposed To Nanosecond Pulsed Electric Field, Mikhail A. Rassokhin

Theses and Dissertations in Biomedical Sciences

Exposure of cells to a pulsed electric field (PEF) is the basis of multiple techniques and treatments. Nanosecond pulsed electric field (nsPEF) poses unique characteristics to induce subtle cellular effects while preserving cell integrity. Improving understanding of the mechanisms triggered by nsPEF in cells inspires new applications for the nanosecond pulse technology. Although many effects of nsPEF remain unknown, they can be inferred from morpho-physiologic changes, or cell reshaping, that accompany nsPEF exposure. During the exposure cells undergo reshaping that is manifested in swelling and diffuse blebbing. Recently we identified two new distinct forms of reshaping, pseudopod-like blebbing and microvesiculation, …

Germline Transgenic Methods For Tracking Cells And Testing Gene Function During Regeneration In The Axolotl, Shahryar Khattak, Maritta Schuez, Tobias Richter, Dunja Knapp, Saori L. Haigo, Tatiana Sandoval-Guzmán, Kristyna Hradlikova, Annett Duemmler, Ryan R. Kerney, Elly M. Tanaka

Biology Faculty Publications

The salamander is the only tetrapod that regenerates complex body structures throughout life. Deciphering the underlying molecular processes of regeneration is fundamental for regenerative medicine and developmental biology, but the model organism had limited tools for molecular analysis. We describe a comprehensive set of germline transgenic strains in the laboratory-bred salamander Ambystoma mexicanum(axolotl) that open up the cellular and molecular genetic dissection of regeneration. We demonstrate tissue-dependent control of gene expression in nerve, Schwann cells, oligodendrocytes, muscle, epidermis, and cartilage. Furthermore, we demonstrate the use of tamoxifen-induced Cre/loxP-mediated recombination to indelibly mark different cell types. Finally, we inducibly …

Truncation Of Type Iv Pilin Induces Mucoidy In Pseudomonas Aeruginosa Strain Pao579, T. Ryan Withers, F. Heath Damron, Yeshi Yin, Hongwei D. Yu

Biochemistry and Microbiology

Pseudomonas aeruginosa is a Gram negative, opportunistic pathogen that uses the overproduction of alginate, a surface polysaccharide, to form biofilms in vivo. Overproduction of alginate, also known as mucoidy, affords the bacterium protection from the host's defenses and facilitates the establishment of chronic lung infections in individuals with cystic fibrosis. Expression of the alginate biosynthetic operon is primarily controlled by the alternative sigma factor AlgU (AlgT/σ²²). In a nonmucoid strain, AlgU is sequestered by the transmembrane antisigma factor MucA to the cytoplasmic membrane. AlgU can be released from MucA via regulated intramembrane proteolysis by proteases AlgW and MucP …

Purification And Characterization Of Oxidation-Resistant Ribonuclease Inhibitor Variants, Alec W. Uebersohn

Lawrence University Honors Projects

Ribonuclease inhibitor (RI) is an intracellular mammalian protein which binds vertebrate-specific ribonucleases; this interaction is one of the tightest non-covalent interactions yet discovered. The biological activity of RI is poorly understood, but it is thought to regulate the biological functions of ribonucleases, which include initiating blood vessel growth, maintaining neuron viability, attacking pathogens, and mediating cell stress responses. RI is also involved in pathways unrelated to ribonucleases, including interactions with Drosha and PTEN, an anti-tumor protein.

One of the defining characteristics of RI is its oxidation sensitivity, a result of its unusually high cysteine content. The oxidation of RI is …

Validation Of Antibodies Used To Study Hypoxia Inducible Factors In Two Species Of Fundulus, Jenna D. Hill

LSU New Orleans Theses and Dissertations

Hypoxia inducible factors (HIFs) are transcription factors and the master regulators of oxygen-dependent gene expression in animals. The focus of this thesis is the distribution of HIF protein in tissues of the fish Fundulus heteroclitus and F. grandis, two widespread species that occur in naturally hypoxic waters. Polyclonal antibodies against HIF-1α, HIF-2α, and HIF-3α were tested on proteins made in vitro and on extracts made from several tissues of normoxic and hypoxic fish. Antibodies against HIF-1α and 3α bound specifically to full length protein made in vitro, and produced bands on western blots of nuclear extracts of near …

Dikar-Induced Synthetic Lethality In A Drosophila Model Of Cag Repeat Diseases Does Not Result From An Expression Feedback Loop, Daniel Camacho

Honors Scholar Theses

Human CAG repeat diseases manifest themselves through the common pathology of neurodeneration. This pathological link is attributed to the property shared by all nine of these diseases: an expanded polyglutamine (polyQ) tract. The most evident result of polyQ expansion is protein aggregation, and it is believed that this phenomenon is partly responsible for conferring cytotoxic properties on the mutated protein. Apart from sequestering the mutated protein, cellular aggregates are able to incorporate native proteins via polyQ-mediated aggregation, thus disrupting important cellular pathways. Using Drosophila melanogaster as a disease model, researchers have been able to compile collections of these so-called disease …

Investigations In Enhancing The Reproducibility Of Implantable Amperometric Glucose Biosensors For Mass Production, Dipesh Manharbhai Patel

Honors Scholar Theses

Implantable glucose sensors for Diabetes management should possess several attributes such as linearity, sensitivity, selectivity, long life time and should elicit no negative tissue response. Based on this, the UConn implantable glucose sensor team has developed a 5-layer sensor architecture that simultaneously affords the aforementioned requirements. However, the large scale production of these 5-layer sensors is inhibited by the lack of high sensor-to-sensor reproducibility. Herein, we investigate the origin of sensor-to-sensor irreproducibility and develop methodologies for large-scale fabrication of these devices.

Analysis Of The Chondroitinase Operon Of Flavobacterium Columnare, Erin L. Sorlien

Senior Honors Projects

Analysis of the chondroitinase operon of Flavobacterium columnare

Erin Sorlien

Major

Cell and Molecular Biology, Chemistry

Advisor

Dr. David R. Nelson

Date

May 2, 2013

Keywords

Flavobacterium columnare, columnaris disease, chondroitin AC lyase, complementation, csl operon

Abstract
Flavobacterium columnare, an opportunistic bacterial pathogen of fish, is the causative agent of columnaris disease (CD). The bacterium is a Gram-negative rod that exhibits gliding motility and avidly forms biofilms. CD affects both wild and cultured freshwater fish, and continues to cause large economic losses to the fish farming industry. According to an investigation conducted by the National Animal Health Monitoring System, CD …

Mechanisms Underlying The Heterogeneous Sensitivities Of Cancer Cells To Proteasome Inhibitors, Matthew C. White

Dissertations and Theses (Open Access)

The mechanisms underlying cellular response to proteasome inhibitors have not been clearly elucidated in solid tumor models. Evidence suggests that the ability of a cell to manage the amount of proteotoxic stress following proteasome inhibition dictates survival. In this study using the FDA-approved proteasome inhibitor bortezomib (Velcade®) in solid tumor cells, we demonstrated that perhaps the most critical response to proteasome inhibition is repression of global protein synthesis by phosphorylation of the eukaryotic initiation factor 2-α subunit (eIF2α). In a panel of 10 distinct human pancreatic cancer cells, we showed marked heterogeneity in the ability of cancer cells to induce …

Characterizing The Role Of Cortactin In Actin Pedestal Assembly By Enterohemorrhagic Escherichia Coli (Ehec), Sarah E. Grout

Honors Scholar Theses

Enterohemorrhagic Escherichia coli (EHEC) is a major foodborne cause of bloody diarrhea and renal failure. During colonization of the intestine, EHEC injects the transmembrane receptor protein Tir and the cytoplasmic effector protein EspFU into host cells to reorganize the actin cytoskeleton into adhesion “pedestals.” EspFU has been shown to bind and activate the actin nucleation factor N-WASP to drive actin polymerization into pedestals. However, EspFU can still assemble pedestals in cells lacking N-WASP, suggesting that this effector protein is able to also trigger N-WASP-independent pathways of actin polymerization during infection. Cortactin is an atypical nucleation factor that localizes to pedestals, …

Characterization Of Aebp2 Mutant Mice, Arundhati Bakshi

Honors Capstones

No abstract provided.

Nuclear Localization Of Cpi-17, A Protein Phosphatase-1 Inhibitor Protein, Affects Histone H3 Phosphorylation And Corresponds To Proliferation Of Cancer And Smooth Muscle Cells., Masumi Eto, Jason A Kirkbride, Rishika Chugh, Nana Kofi Karikari, Jee In Kim

Department of Molecular Physiology and Biophysics Faculty Papers

CPI-17 (C-kinase-activated protein phosphatase-1 (PP1) inhibitor, 17kDa) is a cytoplasmic protein predominantly expressed in mature smooth muscle (SM) that regulates the myosin-associated PP1 holoenzyme (MLCP). Here, we show CPI-17 expression in proliferating cells, such as pancreatic cancer and hyperplastic SM cells. Immunofluorescence showed that CPI-17 was concentrated in nuclei of human pancreatic cancer (Panc1) cells. Nuclear accumulation of CPI-17 was also detected in the proliferating vascular SM cell culture and cells at neointima of rat vascular injury model. The N-terminal 21-residue tail domain of CPI-17 was necessary for the nuclear localization. Phospho-mimetic Asp-substitution of CPI-17 at Ser12 attenuated the nuclear …

Effects Of Synthetic Histone Deacetylase Inhibitors On 3t3l1 Preadipocyte Adipogenic Differentiation Potential, Vikramjeet Singh

Honors Capstones

No abstract provided.

The Herpesvirus Vp1/2 Protein Is An Effector Of Dynein-Mediated Capsid Transport And Neuroinvasion, Sofia V. Zaichick, Kevin P. Bohannon, Ami Hughes, Patricia J. Sollars, Gary E. Pickard, Gregory A. Smith

School of Veterinary and Biomedical Sciences: Faculty Publications

Microtubule transport of herpesvirus capsids from the cell periphery to the nucleus is imperative for viral replication and, in the case of many alphaherpesviruses, transmission into the nervous system. Using the neuroinvasive herpesvirus, pseudorabies virus (PRV), we show that the viral protein 1/2 (VP1/2) tegument protein associates with the dynein/dynactin microtubule motor complex and promotes retrograde microtubule transport of PRV capsids. Functional activation of VP1/2 requires binding to the capsid protein pUL25 or removal of the capsid-binding domain. A proline-rich sequence within VP1/2 is required for the efficient interaction with the dynein/ dynactin microtubule motor complex as well as for …

Pathology In Practice, Timothy L. Cushing, David J. Steffen, Gerald E. Duhamel

School of Veterinary and Biomedical Sciences: Faculty Publications

A 1.5-year-old second-parity Large Black X Tamworth cross sow from a well-managed 20-sow, unvaccinated, pasture-raised herd of pigs in upstate New York aborted a litter of 7 variably mummified near-term fetuses and stillborn piglets. This sow had no signs of ill health other than abortion; it was housed in a group with 4 other sows and a 2-year-old Gloucestershire Old Spot boar. One of the stillborn piglets from this litter was submitted to the New York State Animal Health Diagnostic Center for necropsy; no placental tissue was submitted with the piglet. A second sow from this group had aborted a …

Autophagy Plays An Essential Role In Mediating Regression Of Hypertrophy During Unloading Of The Heart, Nirmala Hariharan, Yoshiyuki Ikeda, Chull Hong, Ralph R. Alcendor, Soichiro Usui, Shumin Gao, Yasuhiro Maejima, Junichi Sadoshima

Publications and Research

Autophagy is a bulk degradation mechanism for cytosolic proteins and organelles. The heart undergoes hypertrophy in response to mechanical load but hypertrophy can regress upon unloading. We hypothesize that autophagy plays an important role in mediating regression of cardiac hypertrophy during unloading. Mice were subjected to transverse aortic constriction (TAC) for 1 week, after which the constriction was removed (DeTAC). Regression of cardiac hypertrophy was observed after DeTAC, as indicated by reduction of LVW/BW and cardiomyocyte cross-sectional area. Indicators of autophagy, including LC3-II expression, p62 degradation and GFP-LC3 dots/cell, were significantly increased after DeTAC, suggesting that autophagy is induced. Stimulation …

Omega-3 Fatty Acids As Therapeutic Options For The Treatment Of B-Cell Chronic Lymphocytic Leukemia, Johannes Francois Fahrmann

Theses, Dissertations and Capstones

B-cell chronic lymphocytic leukemia (CLL) is the most common form of adult leukemia in the western world. CLL is often diagnosed in the asymptomatic (early-stage) stages. However, approximately 50% of these patients will progress to advanced, symptomatic disease and require therapy. Current treatment options are limited due to progressive drug resistance and severe drug-induced toxicities which are often too toxic for the elderly or those with co-morbidities. Therefore, a non-toxic therapeutic intervention that could slow the progression of asymptomatic CLL to symptomatic CLL or enhance the effects of actively used chemo-therapeutic drugs in patients who require therapy would be clinically …

Detection Of Viable Microorganisms Using Propidium Monoazide, Erik J. Mcfarland, Adrian Ponce Dr.

STAR Program Research Presentations

Propidium monoazide (PMA) is a molecular tool used to assess viability of microorganisms. Currently, PMA is thought to discern viability through membrane permeability; PMA enters only membrane compromised cells, irreversibly crosslinks to theirDNAand precipitates theDNAout of solution, preventing it from being amplified during polymerase chain reaction (PCR). Using PMA on a sample of live and dead microorganisms results in only theDNAof living organisms being amplified and identified. Therefore, a comparison ofPCRresults with and without PMA allows one to determine the live fraction and total population, respectively.

Current literature provides conflicting evidence as to the effectiveness of the technique. Our research …

Generation And Characterization Of Peptide Fusion Proteins, Brianna L. Probasco

Graduate School of Biomedical Sciences Theses and Dissertations

Pathogenic Th17 cells drive progression of many autoimmune diseases. Th17 cells develop from naïve T cells in the immune system after antigen-driven stimulation in a specific cytokine environment. Normally, T cells act to fight off infection, but when not properly controlled, they can cause disease. The cytokine interleukin-23 (IL-23) plays an essential role in the expansion of pathogenic Th17 cells. IL-23 is a heterodimeric protein, composed of a p19 alpha chain and a p40 beta chain. The p40 is also part of IL-12 and binds to the IL-12 receptor beta 1 (IL-12Rβ1) subunit. Thus, it follows that the IL-23 receptor …

Extracellular Proteases Are Key Mediators Of Staphylococcus Aureus Virulence Via The Global Modulation Of Virulence-Determinant Stability, Stacey L. Kolar, J. Antonio Ibarra, Frances E. Rivera, Joe M. Mootz, Jessica E. Davenport, Stanley M. Stevens Jr., Alexander R. Horswill, Lindsey N. Shaw

Molecular Biosciences Faculty Publications

Staphylococcus aureus is a highly virulent and successful pathogen that causes a diverse array of diseases. Recently, an increase of severe infections in healthy subjects has been observed, caused by community-associated methicillin-resistant S. aureus (CA-MRSA). The reason for enhanced CA-MRSA virulence is unclear; however, work suggests that it results from hypersecretion of agr-regulated toxins, including secreted proteases. In this study, we explore the contribution of exo-proteases to CA-MRSA pathogenesis using a mutant lacking all 10 enzymes. We show that they are required for growth in peptide-rich environments, serum, in the presence of antimicrobial peptides (AMPs), and in human blood. …

A Transient Α-Helical Molecular Recognition Element In The Disordered N-Terminus Of The Sgs1 Helicase Is Critical For Chromosome Stability And Binding Of Top3/Rmi1, Jessica A. Kennedy, Gary W. Daughdrill, Kristina H. Schmidt

Molecular Biosciences Faculty Publications

The RecQ-like DNA helicase family is essential for the maintenance of genome stability in all organisms. Sgs1, a member of this family in Saccharomyces cerevisiae, regulates early and late steps of double-strand break repair by homologous recombination. Using nuclear magnetic resonance spectroscopy, we show that the N-terminal 125 residues of Sgs1 are disordered and contain a transient α-helix that extends from residue 25 to 38. Based on the residue-specific knowledge of transient secondary structure, we designed proline mutations to disrupt this α-helix and observed hypersensitivity to DNA damaging agents and increased frequency of genome rearrangements. In vitro binding assays …

Identification Of Proteins At Active, Stalled, And Collapsed Replication Forks Using Isolation Of Proteins On Nascent Dna (Ipond) Coupled With Mass Spectrometry, Bianca M. Sirbu, W. Hayes Mcdonald, Huzefa Dungrawala, Akosua Badu-Nkansah, Gina M. Kavanaugh, Yaoyi Chen, David L. Tabb, David Cortez

Molecular Biosciences Faculty Publications

Both DNA and chromatin need to be duplicated during each cell division cycle. Replication happens in the context of defects in the DNA template and other forms of replication stress that present challenges to both genetic and epigenetic inheritance. The replication machinery is highly regulated by replication stress responses to accomplish this goal. To identify important replication and stress response proteins, we combined isolation of proteins on nascent DNA (iPOND) with quantitative mass spectrometry. We identified 290 proteins enriched on newly replicated DNA at active, stalled, and collapsed replication forks. Approximately 16% of these proteins are known replication or DNA …

Recruitment Of The Intracellular Ca2+ By Ultrashort Electric Stimuli: The Impact Of Pulse Duration, Iurii Semenov, Shu Xiao, Olga N. Pakhomova, Andrei G. Pakhomov

Bioelectrics Publications

Nanosecond-duration electric stimuli are distinguished by the ability to permeabilize intracellular membranes and recruit Ca²⁺ from intracellular stores. We quantified this effect in non-excitable cells (CHO) using ratiometric Ca²⁺ imaging with Fura-2. In a Ca²⁺-free medium, 10-, 60-, and 300-ns stimuli evoked Ca2⁺ transients by mobilization of Ca²⁺ from the endoplasmic reticulum. With 2 mM external Ca²⁺, the transients included both extra- and intracellular components. The recruitment of intracellular Ca²⁺ increased as the stimulus duration decreased. At the threshold of 200–300 nM, the transients were amplified by calcium-induced calcium release. We …

Regulation Of The Glycine Transporter 1 (Glyt1) By Pkca-Dependent Ubiquitination, Susana Barrera

Open Access Theses & Dissertations

Glycine is one of the major inhibitory neurotransmitters in the central nervous system. It is implicated in the regulation of motor-sensory function such as; pain perception, reflex responses, and is essential for early development of the CNS. In addition to its inhibitory actions, it acts as an obligatory co-agonist for the activation of glutamatergic NMDARs, which are of pivotal importance in the process of learning and memory, its dysfunction leads to several neurodegenerative diseases. Two high affinity Na+/Cl- dependent transporters GlyT1 and GlyT2 tightly regulate the availability of glycine present at the synapse for the activation of NMDARs and GlyRs. …

Interaction Between Centromeric Histone H3 Variant And Shugoshin, Visarut Buranasudja

Open Access Theses

Precise and faithful segregation of chromosome segregation during mitosis depends on the ability of the cell to regulate chromosome bi-orientation on the mitotic spindle. Shugoshin (Sgo1), the protector of meiotic centromeric cohesin, is required for proper establishment of chromosome bi-orientation. Sgo1 plays a crucial role as part of a mitotic tension sensor between sister chromatids. Recently, Sgo1 has been reported to interact with histone H3 at the pericentromere region, as an important factor for tension sensing and chromosome segregation. However, the role of Sgo1 in tension sensing at centromere is still elusive. The centromere is the region of attachment of …

Characterizing The Effects Of Glutaraldehyde On The Ryanodine Receptor Calcium Release Channel, Joshua Dov Strauss

Legacy Theses & Dissertations (2009 - 2024)

Ryanodine receptors (RyR) are large intracellular calcium release channels, which

Rhodotorula Minuta Fungemia In A Ewe Lamb, C. G. Chitko-Mckown, K. A. Leymaster, M. P. Heaton, D. D. Griffin, J. K. Veatch, S. A. Jones, M. L. Clawson

School of Veterinary and Biomedical Sciences: Faculty Publications

An 8-month-old crossbred ewe, normal upon physical examination, was humanely euthanized for tissue collection. After approximately 3 weeks in tissue culture, fungi began budding out of cells obtained from the choroid plexus. After an additional 3 weeks, budding was observed in kidney cell cultures and eventually in monocyte cultures as well. Serum from the lamb was submitted to the Veterinary Diagnostic Laboratory at Colorado State University for fungal diagnosis and was found negative for Aspergillus, Blastomyces, Coccidioidomycosis and Histoplasmosis. DNA was isolated from fungi collected from tissue culture supernatants and used in a set of pan-fungal PCR assays with …