Life Sciences Commons^™

A Laser Pointer Driven Microheater For Precise Local Heating And Conditional Gene Regulation In Vivo. Microheater Driven Gene Regulation In Zebrafish, Mike Placinta, Meng-Chieh Shen, Marc Achermann, Rolf O. Karlstrom

Rolf O Karlstrom

Background - Tissue heating has been employed to study a variety of biological processes, including the study of genes that control embryonic development. Conditional regulation of gene expression is a particularly powerful approach for understanding gene function. One popular method for mis-expressing a gene of interest employs heat-inducible heat shock protein (hsp) promoters. Global heat shock of hsp-promoter-containing transgenic animals induces gene expression throughout all tissues, but does not allow for spatial control. Local heating allows for spatial control of hsp-promoter-driven transgenes, but methods for local heating are cumbersome and variably effective. Results - We describe a simple, highly controllable, …

Graduate Education In Research Ethics For Scientists And Engineers: Final Report, Jorge Ferrer-Negron, William Frey, Efrain O'Neill-Carrillo, Didier Valdes, Carlos Rios-Valazquez

Ethics in Science and Engineering National Clearinghouse

No abstract provided.

Presence And Prevalence Of Viruses In Local And Migratory Honeybees (Apis Mellifera) In Massachusetts, Anna Welch, Francis Drummond, Sunil Tewari, Anne L. Averill, John P. Burand

Plant, Soil and Insect Sciences Graduate Student Publication Series

Migratory and local bees in Massachusetts were analyzed for seven viruses. three were detected: black queen cell virus (BQCV), deformed wing virus (DWV), and sacbrood virus (SBV). DWV was most common, followed closely by BQCV and then by SBV. BQCV and SBV were present at significantly higher rates in the migratory bees assayed, bringing into question the impact that these bees have on the health of local bee populations.

Toc Receptor Dimerization Participates In The Initiation Of Membrane Translocation During Protein Import Into Chloroplasts, Jeonghwa Lee, Fei Wang Wang, Danny Schnell

Danny Schnell

The post-translational import of nucleus-encoded preproteins into chloroplasts occurs through multimeric translocons in the outer (Toc) and inner (Tic) membranes. The high fidelity of the protein import process is maintained by specific recognition of the transit peptide of preproteins by the coordinate activities of two homologous GTPase Toc receptors, Toc34 and Toc159. Structural and biochemical studies suggest that dimerization of the Toc receptors functions as a component of the mechanism to control access of preproteins to the membrane translocation channel of the translocon. We show that specific mutations that disrupted receptor dimerization in vitro reduced the rate of protein import …

Phosphorylation And The N-Terminal Extension Of The Regulatory Light Chain Help Orient And Align The Myosin Heads In Drosophila Flight Muscle, Gerrie Farman, Mark Miller, Mary Reedy, Felipe Soto-Adames, Jim Vigoreaux, David Maughan, Thomas Irving

Mark S. Miller

X-ray diffraction of the indirect flight muscle (IFM) in living Drosophila at rest and electron microscopy of intact and glycerinated IFM was used to compare the effects of mutations in the regulatory light chain (RLC) on sarcomeric structure. Truncation of the RLC N-terminal extension (Dmlc2Δ2-46) or disruption of the phosphorylation sites by substituting alanines (Dmlc2S66A, S67A) decreased the equatorial intensity ratio (I20/I10), indicating decreased myosin mass associated with the thin filaments. Phosphorylation site disruption (Dmlc2S66A, S67A), but not N-terminal extension truncation (Dmlc2Δ2-46), decreased the 14.5 nm reflection intensity, indicating a spread of the axial distribution of the myosin heads. The …

Presence And Prevalence Of Viruses In Local And Migratory Honeybees (Apis Mellifera) In Massachusetts, John Burand, Anna Welch, Francis Drummond, Sunil Tewari, Anne Averill

John Burand

Migratory and local bees in Massachusetts were analyzed for seven viruses. Three were detected: black queen cell virus (BQCV), deformed wing virus (DWV), and sacbrood virus (SBV). DWV was most common, followed closely by BQCV and then by SBV. BQCV and SBV were present at significantly higher rates in the migratory bees assayed, bringing into question the impact that these bees have on the health of local bee populations.

Runx Proteins Regulate Foxp3 Expression, Janice T. Telfer

Janice C. Telfer

Runx proteins are essential for hematopoiesis and play an important role in T cell development by regulating key target genes, such as CD4 and CD8 as well as lymphokine genes, during the specialization of naive CD4 T cells into distinct T helper subsets. In regulatory T (T reg) cells, the signature transcription factor Foxp3 interacts with and modulates the function of several other DNA binding proteins, including Runx family members, at the protein level. We show that Runx proteins also regulate the initiation and the maintenance of Foxp3 gene expression in CD4 T cells. Full-length Runx promoted the de novo …

Review Of The Winter Harvest Handbook: Year-Round Vegetable Production Using Deep-Organic Techniques And Unheated Greenhouses, Madeleine K. Charney

University Libraries Publication Series

No abstract provided.

Proteogenomic Monitoring Of Geobacter Physiology During Stimulated Uranium Bioremdiation, Michael J. Wilkins, Nathan C. Verberkmes, Kenneth H. Williams, Stephen J. Callister, Paula J. Mouser, Hila Elifantz, A. Lucie N'Guessan, Brian C. Thomas, Carrie D. Nicora, Manesh B. Shah, Paul Abraham, Mary S. Lipton, Derek Lovley, Robert L. Hettich, Philip E. Long, Jillian F. Banfield

Derek Lovley

Implementation of uranium bioremediation requires methods for monitoring the membership and activities of the subsurface microbial communities that are responsible for reduction of soluble U(VI) to insoluble U(IV). Here, we report a proteomics-based approach for simultaneously documenting the strain membership and microbial physiology of the dominant Geobacter community members during in situ acetate amendment of the U-contaminated Rifle, CO, aquifer. Three planktonic Geobacter-dominated samples were obtained from two wells down-gradient of acetate addition. Over 2,500 proteins from each of these samples were identified by matching liquid chromatography-tandem mass spectrometry spectra to peptides predicted from seven isolate Geobacter genomes. Genome-specific peptides …

Ethical Issues In Graduate Research, William J. Frey

Ethics in Science and Engineering National Clearinghouse

(Caution! This module is being published in an incomplete, preliminary version. Later edited and fuller versions will follow.) "Graduate Education in Research Ethics for Scientists and Engineers" is a project funded by the National Science Foundation (SES 0629377) to design a pilot program in research ethics for graduate students in science and engineering. This project is built around three workshops: (1) a Graduate Awareness Workshop introduces students to fundamental ethical issues in research, (2) a Moral Deliberation Workshop acquaints students with the skills of moral deliberation, (3) a Case Analysis Workshop uses realistic scenarios to allow students to practice decision-making …

Tyrosine Phosphorylation Events In Mouse Sperm Capacitation, Enid Arcelay

Open Access Dissertations

Mammalian sperm are not able to fertilize immediately upon ejaculation; they become fertilization-competent after undergoing changes in the female reproductive tract collectively termed capacitation. Although it has been established that capacitation is associated with an increase in tyrosine phosphorylation, little is known about the role of this event in sperm function. In this work we used a combination of two dimensional gel electrophoresis and mass spectrometry to identify proteins that undergo tyrosine phosphorylation during capacitation. Some of the identified proteins are the mouse orthologues of human sperm proteins known to undergo tyrosine phosphorylation. Among them we identified VDAC, tubulin, PDH …

Ciliate Biodiversity And Phylogenetic Reconstruction Assessed By Multiple Molecular Markers, Micah Dunthorn

Open Access Dissertations

Ciliates provide a powerful system within microbial eukaryotes in which molecular genealogies can be compared to detailed morphological taxonomies. Two groups with such detailed taxonomies are the Colpodea and the Halteriidae. There are about 200 described Colpodea species that are found primarily in terrestrial habitats. In Chapters 1 and 2, taxon sampling is increased to include exemplars from all major subclades using nuclear small subunit rDNA (nSSU-rDNA) sequencing. Much of the morphological taxonomy is supported, but extensive non-monophyly is found throughout. The conflict between some nodes of the nSSU-rDNA genealogy and morphology-based taxonomy suggests the need for additional molecular marker. …

Development Of Structured Delivery Systems Using Nanolaminated Biopolymer Layers, Young-Hee Cho

Open Access Dissertations

The objectives of this study were to carry out research to better understand of the formation, stability and properties of multilayer emulsions containing nano-laminated biopolymer coatings, and to utilize this information to develop food-grade delivery systems. The effect of various preparation parameters on the formation and stability of multilayer emulsions was investigated: droplet concentration; mean droplet diameter; droplet charge; biopolymer concentration. β-lactoglobulin (β-Lg) stabilized emulsions (0.5 – 10 wt% oil) containing different pectin concentrations (0 to 0.5 wt%) were prepared at pH 7 (where lipid droplets and pectin molecules were both anionic) and pH 3.5 (where lipid droplets were cationic …

Investigations Into The Potential For 3,4-Methylenedioxymethamphetamine To Induce Neurotoxic Terminal Damage To Serotonergic Neurons, Dominik Biezonski

Open Access Dissertations

High doses of 3,4-methylenedioxymethamphetamine (MDMA; "Ecstasy") are known to reduce levels of various serotonergic markers outside of the raphe nuclei. To test the hypothesis that these deficits reflect a degeneration of distal axons/terminals, we investigated the effects of an MDMA binge (10mg/kg x 4) on the relative protein and genetic expression of several serotonergic markers in rats, as well as the effects of this compound on the quantity of serotonergic terminals in these animals. In experiment I, we examined whether MDMA alters serotonin transporter (SERT) levels as determined by lysate binding and immunoblotting analyses. Both methods of analysis revealed MDMA-induced …

Distribution And Diversity Of Planktonic Ciliates: Patterns And Processes, Mary Doherty

Open Access Dissertations

The nature and extent of microbial biodiversity remain controversial with persistent debates over patterns of distributions (i.e. cosmopolitanism vs. endemism) and the processes that structure these patterns (neutrality vs. selection). We used culture-independent approaches to address these issues focusing on two groups of ciliates, the Oligotrichia (Spirotrichea) and Choreotrichea (Spirotrichea). To assess the diversity of these ciliates, we designed primers specific to SSU rDNA of ciliates within these clades, and investigated (1) geographic and temporal distributions along three coastal sites in the Northwest Atlantic; (2) the relationship between ciliate communities in the benthos and the plankton along the New England …

Seasonal Home Range Sizes, Transboundary Movements And Conservation Of Elephants In Northern Tanzania, Alfred P. Kikoti

Open Access Dissertations

Although the unprotected lands of northern Tanzania support large numbers of elephants, and provide critical linkages for wildlife movements across the region, there is little information on the dispersal patterns of elephants in these unprotected lands. Our home range measures (100% MCP) of 21 elephants with satellite collars in four study regions were highly variable (191 to 3,698 km2). Home range sizes (95% fixed kernel) of bulls were typically larger than those of females, and wet season ranges were typically larger than dry season ranges. There were large differences in average home range sizes reflected varying strategies for obtaining food …

Piping Plover (Charadrius Melodius) Conservation On The Barrier Islands Of New York: Habitat Quality And Implications In A Changing Climate, Jennifer Ruth Seavey

Open Access Dissertations

Habitat loss is the leading cause of species extinction. Protecting and managing habitat quality is vital to an organism's persistence, and essential to endangered species recovery. We conducted an investigation of habitat quality and potential impacts from climate change to piping plovers (Charadrius melodius) breeding on the barrier island ecosystem of New York, during 2003-2005. Our first step in this analysis was to examined the relationship between two common measures of habitat quality: density and productivity (Chapter 1). We used both central and limiting tendency data analysis to find that density significantly limited productivity across many spatial scales, especially broader …

Investigation Of Chemotaxis Genes And Their Functions In Geobacter Species, Hoa T. Tran

Open Access Dissertations

Geobacter species are δ-Proteobacteria and are often predominant in the Fe(III) reduction zone of sedimentary environments. Their abilities to remediate contaminated environments and to produce electricity have inspired extensive studies. Cell motility, biofilm formation, and type IV pili, which have been shown to be regulated by chemotaxis genes in other bacteria, all appear important for the growth of Geobacter species in changing environments and for electricity production. The genomes of Geobacter species show the presence of a significant number of chemotaxis gene homologs, suggesting important roles for them in the physiology of Geobacter species, although gene functions are not yet …

Fabrication Of Protein-Polysaccharide Particulates Through Thermal Treatment Of Associative Complexes, Owen Griffith Jones

Open Access Dissertations

Mixed solutions of β-lactoglobulin and anionic polysaccharides, specifically pectin, were formed into associative complexes through pH reduction from neutral conditions. Thermal treatment of these associative complexes was investigated as a function of biopolymer composition, heating conditions, pH, and ionic strength. Thermal treatment of β-lactoglobulin-pectin complexes at pH 4.5 – 5.0 was found to create protein-based particulates of consistent and narrow size distribution (diameter ~ 150 – 400 nm). These particulates were relatively stable to further pH adjustment and to high levels of salt (200 NaCl). Particle characteristics were maintained after re-suspending them in aqueous solutions after they have been either …

Notch1 Modulation Of Lymphoid Target Genes, Ok Hyun Cho

Open Access Dissertations

Over the past decades, information has accumulated concerning the mechanism how an exterior signal induced by ligand on neighboring cells is transmitted to the nucleus through the Notch receptor and the cellular effects of Notch signaling on the regulation of differentiation, proliferation and apoptosis in many cell types. However, the function and the mechanism of Notch signaling in peripheral T cells still remains to be addressed. Therefore, we asked whether Notch1 is involved in CD8+ cytolytic effector T cell (CTLs) maturation and effector functions and how Notch1 exerts its cellular function in the nucleus and in the cytoplasm. The maturation …

The Activity Of Eg5 And Dynein During Mammalian Mitosis, Nicholas P. Ferenz

Open Access Dissertations

The development and maintenance of multicellular organisms depends fundamentally on cell division, a series of events largely mediated by the mitotic spindle. Errors in spindle formation and/or function are often associated with severe consequences, most notably cancer. In order to elucidate the cause of such errors and the potential for therapeutic intervention, it is imperative to attain a clear understanding of how cell division normally operates. In this regard, this dissertation focuses on the activity of two microtubule-based motor proteins, Eg5 and dynein, prior to and immediately following nuclear envelope breakdown during mitosis. I show that prophase microtubules are remarkably …

Determinants For Stop-Transfer And Post-Import Pathways For Protein Targeting To The Chloroplast Inner Envelope Membrane, Antonio Americo Barbosa Viana

Open Access Dissertations

Chloroplast biogenesis relies on the import of thousands of nuclear encoded proteins into the organelle and proper sorting to their sub-organellar compartment. The majority of nucleus-encoded chloroplast proteins are synthesized in the cytoplasm and imported into the organelle via the Toc-Tic translocation systems of the chloroplast envelope. In many cases, these proteins are further targeted to subcompartments of the organelle (e.g. the thylakoid membrane and lumen or inner envelope membrane) by additional targeting systems that function downstream of the import apparatus. The inner envelope membrane (IEM) plays key roles in controlling metabolite transport between the organelle and cytoplasm, and is …

Professional Ethics-Responsible Conduct Of Research Training: Making Sense Of Complex Problems, Michael D. Mumford, Shane Connelly, Ryan P. Brown, Lynn D. Devenport, Stephen T. Murphy, Alison L. Antes

Ethics in Science and Engineering National Clearinghouse

General Course Abstract

This two-day seminar format course exposes students to the complexities involved in real-world ethical decision-making. It provides students with strategies, or tools, for understanding and thinking through ethical problems to arrive at a decision. Students practice working with these strategies by applying them to realistic, complex cases.

Day 1 Abstract

During the first day of the course, guidelines and principles for ethical research practices are discussed. It is emphasized that students must apply guidelines in a context to arrive at a decision, and the course provides guidance on this decision-making process. Next, students learn about the personal …

Federal Register: National Science Foundation, Responsible Conduct Of Research, National Science Foundation

Ethics in Science and Engineering National Clearinghouse

The National Science Foundation (NSF) is announcing its implementation of Section 7009 of the America Creating Opportunities to Meaningfully Promote Excellence in Technology, Education, and Science (COMPETES) Act (42 U.S.C. 1862o–1). This section of the Act requires that ‘‘each institution that applies for financial assistance from the Foundation for science and engineering research or education describe in its grant proposal a plan to provide appropriate training and oversight in the responsible and ethical conduct of research to undergraduate students, graduate students, and postdoctoral researchers participating in the proposed research project.’’

[Abstract by author]

Persistence Of Colicinogenic Escherichia Coli In The Mouse Gastrointestinal Tract, Margaret Riley, Osnat Gillor, Itamar Giladi

Margaret Riley

Background: The ability of a bacterial strain to competitively exclude or displace other strains can be attributed to the production of narrow spectrum antimicrobials, the bacteriocins. In an attempt to evaluate the importance of bacteriocin production for Escherichia coli strain residence in the gastrointestinal tract, a murine model experimental evolution study was undertaken. Results: Six colicin-producing, yet otherwise isogenic, E. coli strains were administered and established in the large intestine of streptomycin-treated mice. The strains' persistence, population density, and doubling time were monitored over a period of 112 days. Early in the experiment only minor differences in population density between …

Sinorhizobium Meliloti Cpdr1 Is Critical For Co-Ordinating Cell Cycle Progression And The Symbiotic Chronic Infection, H. Kobayashi, N. J. Kobayashi, Peter Chien, L. A. Simmons, G. C. Walker

Peter Chien

ATP-driven proteolysis plays a major role in regulating the bacterial cell cycle, development and stress responses. In the nitro -fixing symbiosis with host plants, Sinorhizobium meliloti undergoes a profound cellular differentiation, including endoreduplication of the ome. The regulatory mechanisms governing the alterations of the S. meliloti cell cycle in planta are largely unknown. Here, we report the characterization of two cpdR homologues, cpdR1 and cpdR2, of S. meliloti that encode single-domain response regulators. In Caulobacter crescentus, CpdR controls the polar localization of the ClpXP protease, thereby mediating the regulated proteolysis of key protein(s), such as CtrA, involved in cell cycle …

Genomic Analysis Of The Basal Lineage Fungus Rhizopus Oryzae Reveals A Whole-Genome Duplication, Li-Jun Ma, Ashraf S. Ibrahim, Christopher Skory, Manfred G. Grabherr, Gertraud Burger, Margi Butler, Marek Elias, Alexander Idnurm, B. Franz Lang, Teruo Sone, Ayumi Abe, Sarah E. Calvo, Luis M. Corrochano, Reinhard Engels, Jianmin Fu, Wilhelm Hansberg, Jung-Mi Kim, Chinnappa D. Kodira, Michael J. Koehrsen, Bo Liu, Diego Miranda-Saavedra, Sinead O'Leary, Lucila Ortiz-Castellanos, Russell Poulter, Julio Rodriguez-Romero, José Ruiz-Herrera, Yao-Qing Shen, Qiandong Zeng, James Galagan, Bruce W. Birren, Christina A. Cuomo, Brian L. Wickes

Li-Jun Ma

Rhizopus oryzae is the primary cause of mucormycosis, an emerging, life-threatening infection characterized by rapid angioinvasive growth with an overall mortality rate that exceeds 50%. As a representative of the paraphyletic basal group of the fungal kingdom called “zygomycetes,” R. oryzae is also used as a model to study fungal evolution. Here we report the genome sequence of R. oryzae strain 99–880, isolated from a fatal case of mucormycosis. The highly repetitive 45.3 Mb genome assembly contains abundant transposable elements (TEs), comprising approximately 20% of the genome. We predicted 13,895 protein-coding genes not overlapping TEs, many of which are paralogous …

Fluorescence Correlation Spectroscopy Of Phosphatidylinositolspecific Phospholipase C Monitors The Interplay Of Substrate And Activator Lipid Binding, Mingming Pu, Mary F. Roberts, Anne Gershenson

Anne Gershenson

Phosphatidylinositol-specific phospholipase C (PI-PLC) enzymes simultaneously interact with the substrate, PI, and with non-substrate lipids such as phosphatidylcholine (PC). For Bacillus thuringiensis PI-PLC these interactions are synergistic with maximal catalytic activity observed at low to moderate mole fractions of PC (XPC) and maximal binding occurring at low mole fractions of anionic lipids. It has been proposed that residues in α helix B help modulate membrane binding and that dimerization on the membrane surface both increases affinity for PC and activates PI-PLC yielding the observed PI/PC synergy. Vesicle binding and activity measurements using a variety of PI-PLC mutants support many aspects …

Modular Laboratory Exercises To Analyze The Development Of Zebrafish Motor Behavior, Kelly Anne Mckeown, Gerald B. Downes, Lara D. Hutson

Gerald B. Downes

The embryonic zebrafish is an excellent research model to examine the neural networks that coordinate locomotive behavior. It demonstrates robust locomotive behavior early in development, its nervous system is relatively simple and accessible compared to mammalian systems, and there are mutants available with specific molecular and motor deficits. We have developed a series of four exercises that provide students with a basic understanding of locomotive behavior development, nervous system organization, development of neurotransmitter responsiveness, and genetics. The first two exercises can be performed in one 3-h laboratory period, and the third and fourth exercises, which build on the first two, …

A Drosophila Resource Of Transgenic Rnai Lines For Neurogenetics, Quan Jian Ni, Lu Ping Liu, Richard Binari, Robert Hardy, Hye Seok Shim, Amanda Cavallaro, Matthew Booker, Barret D. Pfeiffer, Michele Markstein, Hui Wang, Christians Villalta, Todd R. Laverty, Lizabeth A. Perkins, Norbert Perrimon

Michele Markstein

Conditional expression of hairpin constructs in Drosophila is a powerful method to disrupt the activity of single genes with a spatial and temporal resolution that is impossible, or exceedingly difficult, using classical genetic methods. We previously described a method (Ni et al. 2008) whereby RNAi constructs are targeted into the genome by the phiC31-mediated integration approach using Vermilion-AttB-Loxp-Intron-UAS-MCS (VALIUM), a vector that contains vermilion as a selectable marker, an attB sequence to allow for phiC31-targeted integration at genomic attP landing sites, two pentamers of UAS, the hsp70 core promoter, a multiple cloning site, and two introns. As the level of …