Biochemistry, Biophysics, and Structural Biology Commons^™

Ubiquitin Goes Green, Zhihua Hua, Richard D. Vierstra

Biology Faculty Publications & Presentations

Chloroplasts depend on the nucleus for much of their proteome. Consequently, strong transcriptional coordination exists between the genomes, which is attuned to the developmental and physiological needs of the organelle. Recent studies highlight that the post-translational modifier ubiquitin adds another layer to plastid homeostasis and even helps eliminate damaged chloroplasts.

Population Density Of The Florida Scrub Lizard (Sceloporus Woodi) In Managed Sand Pine Scrub And Longleaf Pine Sandhill Habitats, Matthew D. Kaunert, Lance D. Mcbrayer

Department of Biology Faculty Publications

Studies investigating managed landscapes are of increasing importance, as fragmentation is a known cause of biodiversity loss. From June to September 2012, we sampled populations of the rare, endemic Florida Scrub Lizard (Sceloporus woodi) across the Ocala National Forest (ONF) to compare lizard density across two managed habitat types. Florida Scrub habitat in the ONF is clearcut and roller-chopped, whereas Longleaf Pine habitat is managed via prescribed burning. We sampled 10 stands of Florida Scrub (2–3 y post disturbance) and 10 stands of Longleaf Pine (1 y post-disturbance) for lizards. We compared lizard density between the interior of …

Discovery & Born-Digital Archiving: Open Source Systems For Preservation And Access, L. Bryan Cooper, Margarita Perez-Martinez

Works of the FIU Libraries

The Everglades Explorer (EE) portal at http://ee.fiu.edu continues to evolve with the addition of the Internet Archive's Archive-It, and future planned alignment with the Digital Public Library of America (DPLA). The reasons for the development of the portal continue to exist, as originally perceived four years ago. Adaptations to ongoing change and system testing continues, and the use of Archive-It has broad institutional potential beyond EE. Cross-walking skills continue to grown, and will benefit future syndication and discovery system integration. The metadata normalization and harmonization will help save time for the end-user. Preservation and access to learning and research …

Transport Mechanisms For Human Fecal Indicator Bacteria In An Urban Stormwater Basin In Southeastern Wisconsin, Chelsea M. Corson

Theses and Dissertations

Discharge of stormwater runoff to receiving waters is a known source of human pathogens; however the primary mechanisms by which these pathogens enter the stormwater system have yet to be quantified. This study builds upon and utilizes prior research findings in an attempt to explain the influence of the age of the pipes within stormwater and sanitary conveyance systems, rainfall and hydrogeological characteristics, and select infrastructure variables that contribute to the observed contamination of an urban stormwater basin in Southeastern Wisconsin.

Over the course of approximately two years from 2012 to 2014, a total of 260 samples from 22 stormwater …

Expression Of Insulin Responsive Genes In Insulin Resistant Conditions, And The Effect Of Selenium On Gene Expression, David L. Ruff

Masters Theses

Chronically high blood glucose levels lead to many problems, such as insulin resistance, the hallmark of Type II diabetes. Increased flux through the hexosamine biosynthesis pathway is one mechanism by which high glucose as well as glucosamine has been shown to induce insulin resistance. This study tests the effects of glucosamine induced insulin resistance on insulin regulation of the metabolic genes glucose-6-phosphate dehydrogenase (G6PDH) and fatty acid synthase (FAS) as well as insulin responsive proteins tribbles homolog (TRIB3) and sterol regulatory element binding protein (SERBP-1c) 1c.

Selenium, a micronutrient has been shown to be an effective insulin mimetic in Type …

Characterization Of Sbip68: A Putative Tobacco Glucosyltransferase Protein And Its Role In Plant Defense Mechanisms, Abdulkareem O. Odesina

Electronic Theses and Dissertations

Plant secondary metabolites are essential for normal growth and development in plants ultimately affecting crop yield. They play roles ranging from appearance of the plants to defending against pathogen attack and herbivory. They have been used by humans for medicinal and recreational purposes amongst others. Glycosyltransferases catalyze the transfer of sugars from donor substrates to acceptors. Glucosyltransferases are a specific type of glycosyltransferases known to transfer glucose molecules from a glucose donor to a glucose acceptor (aglycone) producing the corresponding glucose secondary metabolite or glycone, in this case glucosides. It was hypothesized that SBIP68, a tobacco putative glucosyltransferase-like protein glucosylated …

Normal Glycolytic Enzyme Activity Is Critical For Hypoxia Inducible Factor-1a Activity And Provides Novel Targets For Inhibiting Tumor Growth, Geoffrey Grandjean Phd

Dissertations & Theses (Open Access)

Normal Glycolytic Enzyme Activity is Critical for Hypoxia Inducible Factor-1α Activity and Provides Novel Targets for Inhibiting Tumor Growth

By Geoffrey Grandjean

Advisory Professor: Garth Powis, D. Phil

Unique to proliferating cancer cells is the observation that their increased need for energy is provided by a high rate of glycolysis followed by lactic acid fermentation in a process known as the Warburg Effect, a process many times less efficient than oxidative phosphorylation employed by normal cells to satisfy a similar energy demand ^[1]. This high rate of glycolysis occurs regardless of the concentration of oxygen in the cell and …

Production And Harvest Of Microalgae In Wastewater Raceways With Resource Recycling, Alexander Colin Roberts

Master's Theses

Microalgae can be grown on municipal wastewater media to both treat the wastewater and produce feedstock for algae biofuel production. However the reliability of treatment must be demonstrated, as well as high areal algae productivity on recycled wastewater media and efficient sedimentation harvesting. This processes was studied at pilot scale in the present research.

A pilot facility was operated with nine CO₂-supplemented raceway ponds, each with a 33-m² surface area and a 0.3-m depth, continuously from March 6, 2013 through September 24, 2014. The ponds were operated as three sets of triplicates with two sets continuously fed …

Promoting Extracellular Matrix Crosslinking In Synthetic Hydrogels, Marcos M. Manganare

Masters Theses

The extracellular matrix (ECM) provides mechanical and biochemical support to tissues and cells. It is crucial for cell attachment, differentiation, and migration, as well as for ailment-associated processes such as angiogenesis, metastases and cancer development. An approach to study these phenomena is through emulation of the ECM by synthetic gels constructed of natural polymers, such as collagen and fibronectin, or simple but tunable materials such as poly(ethylene glycol) (PEG) crosslinked with short peptide sequences susceptible to digestion by metalloproteases and cell-binding domains. Our lab uses PEG gels to study cell behavior in three dimensions (3D). Although this system fosters cell …

Are Eastern Oysters Being Bored To Death? Influence Of Cliona Celata On Crassostrea Virginica Condition, Growth And Survival, John M. Carroll, Katheryn A. O'Shaughnessy, Grant A. Diedrich, Christopher M. Finelli

Department of Biology Faculty Publications

The boring sponge Cliona celata is a nuisance species that can have deleterious effects on eastern oyster Crassostrea virginica growth, condition, and survival. Surprisingly, however, these effects have not been well documented and when examined, results have been equi-vocal. In this study, we provide a direct comparison of growth, condition, and survival of sponge-colonized and uncolonized oysters in southeast North Carolina in 2 separate experiments. In the first experiment, sponge-colonized oysters exhibited significantly slower growth rates, reduced condition, and lower survival relative to uncolonized oysters, although results may have been confounded by oyster source. In the second experiment, using smaller …

Effect Of Genetic Background On The Dystrophic Phenotype In Mdx Mice., William D Coley, Laurent Bogdanik, Maria Candida Vila, Qing Yu, Terence A Partridge, Kanneboyina Nagaraju, +12 Additional Authors

Genomics and Precision Medicine Faculty Publications

Genetic background significantly affects phenotype in multiple mouse models of human diseases, including muscular dystrophy. This phenotypic variability is partly attributed to genetic modifiers that regulate the disease process. Studies have demonstrated that introduction of the γ-sarcoglycan null allele onto the DBA/2J background confers a more severe muscular dystrophy phenotype than the original strain, demonstrating the presence of genetic modifier loci in the DBA/2J background. To characterize the phenotype of dystrophin deficiency on the DBA/2J background, we created and phenotyped DBA/2J-congenic Dmdmdx mice (D2-mdx) and compared them to the original, C57BL/10ScSn-Dmdmdx (B10-mdx) model. These strains were compared to their respective …

Cytoplasmic Ph Measurement And Homeostasis In Bacteria And Archaea., Joan Slonczewski

Joan Slonczewski

Of all the molecular determinants for growth, the hydronium and hydroxide ions are found naturally in the widest concentration range, from acid mine drainage below pH 0 to soda lakes above pH 13. Most bacteria and archaea have mechanisms that maintain their internal, cytoplasmic pH within a narrower range than the pH outside the cell, termed "pH homeostasis." Some mechanisms of pH homeostasis are specific to particular species or groups of microorganisms while some common principles apply across the pH spectrum. The measurement of internal pH of microbes presents challenges, which are addressed by a range of techniques under varying …

Ph Of The Cytoplasm And Periplasm Of Escherichia Coli: Rapid Measurement By Green Fluorescent Protein Fluorimetry, Joan Slonczewski, Jessica C. Wilks

Joan Slonczewski

Cytoplasmic pH and periplasmic pH of Escherichia coli cells in suspension were observed with 4-s time resolution using fluorimetry of TorA-green fluorescent protein mutant 3* (TorA-GFPmut3*) and TetR-yellow fluorescent protein. Fluorescence intensity was correlated with pH using cell suspensions containing 20 mM benzoate, which equalizes the cytoplasmic pH with the external pH. When the external pH was lowered from pH 7.5 to 5.5, the cytoplasmic pH fell within 10 to 20 s to pH 5.6 to 6.5. Rapid recovery occurred until about 30 s after HCl addition and was followed by slower recovery over the next 5 min. As a …

Cytoplasmic Ph Response To Acid Stress In Individual Cells Of Escherichia Coli And Bacillus Subtilis Observed By Fluorescence Ratio Imaging Microscopy, Joan Slonczewski, Keith A. Martinez Ii, Ryan D. Kitko, J. Patrick Mershon, Haley E. Adcox, Kotiba A. Malek, Melanie B. Berkmen

Joan Slonczewski

The ability of Escherichia coli and Bacillus subtilis to regulate their cytoplasmic pH is well studied in cell suspensions but is poorly understood in individual adherent cells and biofilms. We observed the cytoplasmic pH of individual cells using ratiometric pHluorin. A standard curve equating the fluorescence ratio with pH was obtained by perfusion at a range of external pH 5.0 to 9.0, with uncouplers that collapse the transmembrane pH difference. Adherent cells were acid stressed by switching the perfusion medium from pH 7.5 to pH 5.5. The E. coli cytoplasmic pH fell to a value that varied among individual cells …

Ph Regulates Genes For Flagellar Motility, Catabolism, And Oxidative Stress In Escherichia Coli K-12†, Joan Slonczewski

Joan Slonczewski

Gene expression profiles of Escherichia coli K-12 W3110 were compared as a function of steady-state external pH. Cultures were grown to an optical density at 600 nm of 0.3 in potassium-modified Luria-Bertani medium buffered at pH 5.0, 7.0, and 8.7. For each of the three pH conditions, cDNA from RNA of five independent cultures was hybridized to Affymetrix E. coli arrays. Normalized expression indices were calculated for each gene and intergenic region (IG). Differential expression among the three pH classes was observed for 763 genes and 353 IGs. Hierarchical clustering yielded six well-defined clusters of pH profiles, designated Acid High …

Generalization Of Dna Microarray Dispersion Properties: Microarray Equivalent Of T-Distribution, Joan Slonczewski

Joan Slonczewski

Background: DNA microarrays are a powerful technology that can provide a wealth of gene expression data for disease studies, drug development, and a wide scope of other investigations. Because of the large volume and inherent variability of DNA microarray data, many new statistical methods have been developed for evaluating the significance of the observed differences in gene expression. However, until now little attention has been given to the characterization of dispersion of DNA microarray data. Results: Here we examine the expression data obtained from 682 Affymetrix GeneChips® with 22 different types and we demonstrate that the Gaussian (normal) frequency distribution …

Escherichia Coli K-12 Survives Anaerobic Exposure At Ph 2 Without Rpos, Gad, Or Hydrogenases, But Shows Sensitivity To Autoclaved Broth Products, Joan Slonczewski, Daniel P. Riggins, Maria J. Narvaez, Keith A. Martinez, Mark M. Harden

Joan Slonczewski

Escherichia coli and other enteric bacteria survive exposure to extreme acid (pH 2 or lower) in gastric fluid. Aerated cultures survive via regulons expressing glutamate decarboxylase (Gad, activated by RpoS), cyclopropane fatty acid synthase (Cfa) and others. But extreme-acid survival is rarely tested under low oxygen, a condition found in the stomach and the intestinal tract. We observed survival of E. coli K-12 W3110 at pH 1.2–pH 2.0, conducting all manipulations (overnight culture at pH 5.5, extreme-acid exposure, dilution and plating) in a glove box excluding oxygen (10% H2, 5% CO2, balance N2). With dissolved O2 concentrations maintained below 6 …

Hydrogenase-3 Contributes To Anaerobic Acid Resistance Of Escherichia Coli, Joan Slonczewski, Ken Noguchi, Daniel P. Riggins, Khalid C. Eldahan, Ryan D. Kitko

Joan Slonczewski

Background: Hydrogen production by fermenting bacteria such as Escherichia coli offers a potential source of hydrogen biofuel. Because H2 production involves consumption of 2H+ , hydrogenase expression is likely to involve pH response and regulation. Hydrogenase consumption of protons in E. coli has been implicated in acid resistance, the ability to survive exposure to acid levels (pH 2–2.5) that are three pH units lower than the pH limit of growth (pH 5–6). Enhanced survival in acid enables a larger infective inoculum to pass through the stomach and colonize the intestine. Most acid resistance mechanisms have been defined using aerobic cultures, …

Osmolytes Contribute To Ph Homeostasis Of Escherichia Coli, Joan Slonczewski, Ryan D. Kitko, Jessica C. Wilks, Gian M. Garduque

Joan Slonczewski

Background: Cytoplasmic pH homeostasis in Escherichia coli includes numerous mechanisms involving pH-dependent catabolism and ion fluxes. An important contributor is transmembrane K+ flux, but the actual basis of K+ compensation for pH stress remains unclear. Osmoprotection could mediate the pH protection afforded by K+ and other osmolytes. Methods and Principal Findings: The cytoplasmic pH of E. coli K-12 strains was measured by GFPmut3 fluorimetry. The wild-type strain Frag1 was exposed to rapid external acidification by HCl addition. Recovery of cytoplasmic pH was enhanced equally by supplementation with NaCl, KCl, proline, or sucrose. A triple mutant strain TK2420 defective for the …

Oxygen Limitation Modulates Ph Regulation Of Catabolism And Hydrogenases, Multidrug Transporters, And Envelope Composition In Escherichia Coli K-12, Joan Slonczewski, Everett T. Hayes, Jessica C. Wilks, Piero Sanfilippo, Elizabeth Yohannes, Daniel P. Tate, Brian D. Jones, Michael D. Radmacher, Sandra S. Bondurant

Joan Slonczewski

Background: In Escherichia coli, pH regulates genes for amino-acid and sugar catabolism, electron transport, oxidative stress, periplasmic and envelope proteins. Many pH-dependent genes are co-regulated by anaerobiosis, but the overall intersection of pH stress and oxygen limitation has not been investigated. Results: The pH dependence of gene expression was analyzed in oxygen-limited cultures of E. coli K-12 strain W3110. E. coli K-12 strain W3110 was cultured in closed tubes containing LBK broth buffered at pH 5.7, pH 7.0, and pH 8.5. Affymetrix array hybridization revealed pH-dependent expression of 1,384 genes and 610 intergenic regions. A core group of 251 genes …

Ph-Dependent Expression Of Periplasmic Proteins And Amino Acid Catabolism In Escherichia Coli, Joan Slonczewski, Lauren M. Stancik, Dawn M. Stancik, Brian Schmidt, D. Michael Barnhart, Yuliya N. Yoncheva

Joan Slonczewski

Escherichia coli grows over a wide range of pHs (pH 4.4 to 9.2), and its own metabolism shifts the external pH toward either extreme, depending on available nutrients and electron acceptors. Responses to pH values across the growth range were examined through two-dimensional electrophoresis (2-D gels) of the proteome and through lac gene fusions. Strain W3110 was grown to early log phase in complex broth buffered at pH 4.9, 6.0, 8.0, or 9.1. 2-D gel analysis revealed the pH dependence of 19 proteins not previously known to be pH dependent. At low pH, several acetate-induced proteins were elevated (LuxS, Tpx, …

Ph-Dependent Catabolic Protein Expression During Anaerobic Growth Of Escherichia Coli K-12, Joan Slonczewski, Elizabeth Yohannes, D. Michael Barnhart

Joan Slonczewski

During aerobic growth of Escherichia coli, expression of catabolic enzymes and envelope and periplasmic proteins is regulated by pH. Additional modes of pH regulation were revealed under anaerobiosis. E. coli K-12 strain W3110 was cultured anaerobically in broth medium buffered at pH 5.5 or 8.5 for protein identification on proteomic two-dimensional gels. A total of 32 proteins from anaerobic cultures show pH-dependent expression, and only four of these proteins (DsbA, TnaA, GatY, and HdeA) showed pH regulation in aerated cultures. The levels of 19 proteins were elevated at the high pH; these proteins included metabolic enzymes (DhaKLM, GapA, TnaA, HisC, …

Polyamine Stress At High Ph In Escherichia Coli K-12, Joan Slonczewski, Elizabeth Yohannes, Amy E. Thurber, Jessica C. Wilks, Daniel P. Tate

Joan Slonczewski

Background: Polyamines such as spermine and spermidine are required for growth of Escherichia coli; they interact with nucleic acids, and they bind to ribosomes. Polyamines block porins and decrease membrane permeability, activities that may protect cells in acid. At high concentrations, however, polyamines impair growth. They impair growth more severely at high pH, probably due to their increased uptake as membrane-permeant weak bases. The role of pH is critical in understanding polyamine stress. Results: The effect of polyamines was tested on survival of Escherichia coli K-12 W3110 in extreme acid or base (pH conditions outside the growth range). At pH …

Isotopic Discrimination In The Double-Crested Cormorant (Phalacrocorax Auritus), Elizabeth C. Craig, Brian S. Dorr, Katie C. Hanson-Dorr, Jed P. Sparks, Paul D. Curtis

Brian S Dorr

The diet-tissue discrimination factor is the amount by which a consumer’s tissue varies isotopically from its diet, and is therefore a key element in models that use stable isotopes to estimate diet composition. In this study we measured discrimination factors in blood (whole blood, red blood cells and plasma), liver, muscle and feathers of Double- crested Cormorants (Phalacrocorax auritus) for stable isotope ratios of carbon, nitrogen and sulfur. Cormorants exhibited discrimination factors that differed significantly among tissue types (for carbon and nitrogen), and differed substantially (in the context of the isotopic variation among relevant prey species) from those observed in …

Two Distinct Modes Of Metal Ion Binding In The Nuclease Active Site Of A Viral Dna-Packaging Terminase: Insight Into The Two-Metal-Ion Catalytic Mechanism, Haiyan Zhao, Zihan Lin, Anna Y. Lynn, Brittany Varnado, John A. Beutler, Ryan P. Murelli, Stuart F.J. Le Grice, Liang Tang

Publications and Research

Many dsDNA viruses encode DNA-packaging terminases, each containing a nuclease domain that resolves concatemeric DNA into genome-length units. Terminase nucleases resemble the RNase H-superfamily nucleotidyltransferases in folds, and share a two-metal-ion catalytic mechanism. Here we show that residue K428 of a bacteriophage terminase gp2 nuclease domain mediates binding of the metal cofactor Mg2+. A K428A mutation allows visualization, at high resolution, of a metal ion binding mode with a coupled-octahedral configuration at the active site, exhibiting an unusually short metal-metal distance of 2.42 A° . Such proximity of the two metal ions may play an essential role in catalysis by …

Reactive Oxygen Species-Mediated Neurodegeneration Is Independent Of The Ryanodine Receptor In Caenorhabditis Elegans, Lyndsay E.A. Young, Daniel C. Williams

Journal of the South Carolina Academy of Science

Despite the significant impacts on human health caused by neurodegeneration, our understanding of the degeneration process is incomplete. The nematode Caenorhabditis elegans is emerging as a genetic model organism well suited for identification of conserved cellular mechanisms and molecular pathways of neurodegeneration. Studies in the worm have identified factors that contribute to neurodegeneration, including excitotoxicity and stress due to reactive oxygen species (ROS). Disruption of the gene unc-68, which encodes the ryanodine receptor, abolishes excitotoxic cell death, indicating a role for calcium (Ca²⁺) signaling in neurodegeneration. We tested the requirement for unc-68 in ROS-mediated neurodegeneration using the …

Incipient Speciation In Freshwater Fish Species From Two Isolated Watersheds, Paula Gore Miller

Dissertations, Theses, and Capstone Projects

The process of speciation occurs as a result of restricted gene flow between segments of an interbreeding population occupying different geographic areas. This separation may result in isolated populations which undergo genetic and phenotypic changes. The Wisconsin glacial period, which ended approximately 17,500 years ago, dramatically altered the geography of North America. The glacier covered almost the entire North America as it advanced. Areas that were not covered with ice provided suitable habitats (refugia) for relict species that were previously widespread in the northern section of the continent. As the ice sheet retreated, animals and plants were able to return …

Alternative Strategies In Response To Saline Stress In Two Varieties Of Portulaca Oleracea (Purslane), Dana A. Dudle, Kristina R. Mulry, Bryan A. Hanson

Biology Faculty publications

Purslane (Portulaca oleracea) is a globally-distributed plant with a long history of use in folk medicine and cooking. We have developed purslane as a model system for exploring plant responses to stress. We exposed two varieties of purslane to saline stress with the objective of identifying differences between the varieties in the plasticity of morphological and physiological traits. The varieties responded to saline stress with significantly different changes in the measured traits, which included inter alia biomass, flower counts, proline concentrations and betalain pigment concentrations. The alternative responses of the two varieties consisted of complex, simultaneous changes in …

A Forward Genetic Screen Identifies Factors Associated With Fever Pathogenesis In Plasmodium Falciparum, Phaedra J. Thomas

USF Tampa Graduate Theses and Dissertations

Infectious diseases that spread from person-to-person and continent-to-continent are a cause for concern for any health entity. One such disease is malaria, a mosquito-borne infection instigated by the protozoan parasite, Plasmodium falciparum. Hundreds of millions of people are affected annually and it is responsible for nearly 1 million deaths. It is the most fatal species causing malaria and proliferates in human red blood cells with a life cycle occurring every 48 hours. At this time, the parasite’s late stage form or schizont bursts from the erythrocyte releasing immune-inducing particles and infective forms (merozoites) into the bloodstream. The merozoites go …

Precise Repair Of Mping Excision Sites Is Facilitated By Target Site Duplication Derived Microhomology, David M. Gilbert, M. Catherine Bridges, Ashley E. Strother, Courtney E. Burckhalter, James M. Burnette Iii, C. Nathan Hancock

Faculty Publications

A key difference between the Tourist and Stowaway families of miniature inverted repeat transposable elements (MITEs) is the manner in which their excision alters the genome. Upon excision, Stowaway-like MITEs and the associated Mariner elements usually leave behind a small duplication and short sequences from the end of the element. These small insertions or deletions known as “footprints” can potentially disrupt coding or regulatory sequences. In contrast, Tourist-like MITEs and the associated PIF/Pong/Harbinger elements generally excise precisely, returning the genome to its original state. The purpose of this study was to determine the mechanisms underlying these …