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, …

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 …

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 …

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 …

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 …

Leaf Herbivory Increases Floral Fragrance In Male But Not Female Cucurbita Peposubsp.Texana (Cucurbitaceae) Flowers, Nina Theis, Karen Kesler, Lynn Adler

Lynn Adler

Mutualisms are key interactions that affect population dynamics and structure communities, but the extent to which mutualists can attract potential partners may depend on community context. Many studies have shown that leaf herbivory reduces pollinator visitation and have focused on reduced fl oral visual display and rewards as potential mechanisms. However, olfactory display plays a critical role in mediating interactions between plants, herbivores, and pollinators. We simulated leaf damage in Cucurbita pepo subsp. texana and measured fragrance emission and other fl oral characters of both male and female fl owers. Contrary to our expectations, damage increased fragrance production, but only …

Exploring The Transcriptional Landscape Of Plant Circadian Rhythms Using Genome Tiling Arrays, Samuel P. Hazen, Felix Naef, Tom Quisel, Joshua M. Gendron, Huaming Chen, Joseph R. Ecker, Justin O. Borevitz, Steve A. Kay

Samuel P Hazen

Background - Organisms are able to anticipate changes in the daily environment with an internal oscillator know as the circadian clock. Transcription is an important mechanism in maintaining these oscillations. Here we explore, using whole genome tiling arrays, the extent of rhythmic expression patterns genome-wide, with an unbiased analysis of coding and noncoding regions of the Arabidopsis genome. Results - As in previous studies, we detected a circadian rhythm for approximately 25% of the protein coding genes in the genome. With an unbiased interrogation of the genome, extensive rhythmic introns were detected predominantly in phase with adjacent rhythmic exons, creating …

Intrakinetochore Stretch Is Associated With Changes In Kinetochore Phosphorylation And Spindle Assembly Checkpoint Activity, Thomas J. Maresca, E. D. Salmon

Thomas J. Maresca

Cells have evolved a signaling pathway called the spindle assembly checkpoint (SAC) to increase the fidelity of chromosome segregation by generating a “wait anaphase” signal until all chromosomes are properly aligned within the mitotic spindle. It has been proposed that tension generated by the stretch of the centromeric chromatin of bioriented chromosomes stabilizes kinetochore microtubule attachments and turns off SAC activity. Although biorientation clearly causes stretching of the centromeric chromatin, it is unclear whether the kinetochore is also stretched. To test whether intrakinetochore stretch occurs and is involved in SAC regulation, we developed a Drosophila melanogaster S2 cell line expressing …

Steroid Use And Human Performance: Lessons For Integrative Biologists, Jerry F. Husak, Duncan J. Irschick

Biology Department Faculty Publication Series

While recent studies have begun to address how hormones mediate whole-animal performance traits, the field conspicuously lags behind research conducted on humans. Recent studies of human steroid use have revealed that steroid use increases muscle cross-sectional area and mass, largely due to increases in protein synthesis, and muscle fiber hypertrophy attributable to an increased number of satellite cells and myonuclei per unit area. These biochemical and cellular effects on skeletal muscle morphology translate into increased power and work during weight-lifting and enhanced performance in burst, sprinting activities. However, there are no unequivocal data that human steroid use enhances endurance performance …

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

Biology Department Faculty Publication Series

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, and versatile …

A New Laser Pointer Driven Optical Microheater For Precise Local Heat Shock, Mike Placinta

Masters Theses 1911 - February 2014

The zebrafish has emerged as an important genetic model system for the study of vertebrate development. However, while genetics is a powerful tool for the study of early gene functions, the approach is more limited when it comes to understanding later functions of genes that have essential roles in early embryogenesis. There is thus a need to manipulate gene expression at different times, and ideally only in some regions of the developing embryo. Methods for conditional gene regulation have been established in Drosophila, C.elegans and the mouse, utilizing conditional gene activation systems such as the Gal4-UAS system (fly) and the …

Using Birds To Teach Biology, Bruce Byers

Science and Engineering Saturday Seminars

Cornell University curriculum materials http://www.birdsleuth.org/inquiry/

Illuminating Life: What's New And Noteworthy In, Pat O'Hara, Robert Blatchly

Science and Engineering Saturday Seminars

No abstract provided.

Rapid Screening For Temperature-Sensitive Alleles In Plants, L Vidali, Rc Augustine, Sn Fay, P Franco, Ka Pattavina, M Bezanilla

Biology Department Faculty Publication Series

No abstract provided.

Lifeact-Megfp Reveals A Dynamic Apical F-Actin Network In Tip Growing Plant Cells, L Vidali, Cm Rounds, Pk Hepler, M Bezanilla

Biology Department Faculty Publication Series

No abstract provided.

Rapid Formin-Mediated Actin-Filament Elongation Is Essential For Polarized Plant Cell Growth, L Vidali, Pac Van Gisbergen, C Guerin, P Franco, M Li, Gm Burkart, Rc Augustine, L Blanchoin, M Bezanilla

Biology Department Faculty Publication Series

No abstract provided.

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

Biology Department Faculty Publication Series

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, …

Exploring The Transcriptional Landscape Of Plant Circadian Rhythms Using Genome Tiling Arrays, Samuel P. Hazen, Felix Naef, Tom Quisel, Joshua M. Gendron, Huaming Chen, Joseph R. Ecker, Justin O. Borevitz, Steve A. Kay

Biology Department Faculty Publication Series

Background

Organisms are able to anticipate changes in the daily environment with an internal oscillator know as the circadian clock. Transcription is an important mechanism in maintaining these oscillations. Here we explore, using whole genome tiling arrays, the extent of rhythmic expression patterns genome-wide, with an unbiased analysis of coding and noncoding regions of the Arabidopsis genome.

Results

As in previous studies, we detected a circadian rhythm for approximately 25% of the protein coding genes in the genome. With an unbiased interrogation of the genome, extensive rhythmic introns were detected predominantly in phase with adjacent rhythmic exons, creating a transcript …

Hormonal Regulation Of Whole-Animal Performance: Implications For Selection, Jerry F. Husak, Duncan J. Irschick, Stephen D. Mccormick, Ignacio T. Moore

Biology Department Faculty Publication Series

No abstract provided.

A Dynamic Gli Code Interprets Hh Signals To Regulate Induction, Patterning, And Endocrine Cell Specification In The Zebrafish Pituitary, Christine A. Devine, Jennifer L. Sorogna, Burcu Guner, Marcey Osgood, Meng-Chieh Shen, Rolf O. Karlstrom

Biology Department Faculty Publication Series

Hedgehog (Hh) signaling is necessary for the induction and functional patterning of the pituitary placode, however the mechanisms by which Hh signals are interpreted by placodal cells are unknown. Here we show distinct temporal requirements for Hh signaling in endocrine cell differentiation and describe a dynamic Gli transcriptional response code that interprets these Hh signals within the developing adenohypophysis. Gli1 is required for the differentiation of selected endocrine cell types and acts as the major activator of Hh-mediated pituitary induction, while Gli2a and Gli2b contribute more minor activator functions. Intriguingly, this Gli response code changes as development proceeds. Gli1 continues …

A Caax Motif Can Compensate For The Ph Domain Of Num1 For Cortical Dynein Attachment, Xianying Tang, Jesse J. Punch, Wei-Lih Lee

Biology Department Faculty Publication Series

During mitosis in budding yeast, cortically anchored dynein exerts pulling forces on cytoplasmic microtubules, moving the mitotic spindle into the mother-bud neck. Anchoring of dynein requires the cortical patch protein Num1, which is hypothesized to interact with PI(4,5)P₂ via its C-terminal pleckstrin homology (PH) domain. Here we show that the PH domain and PI(4,5)P₂ are required for the cortical localization of Num1, but are not sufficient to mediate the cortical assembly of Num1 patches. A GFP fusion to the PH domain localizes to the cortex in foci containing ~2 molecules, whereas patches of full-length Num1-GFP contain ~14 molecules. …

Functional Overlap Of Microtubule Assembly Factors In Chromatin-Promoted Spindle Assembly, Thomas J. Maresca, A. C. Groen, J. C. Gatlin, E. D. Salmon

Thomas J. Maresca

Distinct pathways from centrosomes and chromatin are thought to contribute in parallel to microtubule nucleation and stabilization during animal cell mitotic spindle assembly, but their full mechanisms are not known. We investigated the function of three proposed nucleation/stabilization factors, TPX2, γ-tubulin and XMAP215, in chromatin-promoted assembly of anastral spindles in Xenopus laevis egg extract. In addition to conventional depletion-add back experiments, we tested whether factors could substitute for each other, indicative of functional redundancy. All three factors were required for microtubule polymerization and bipolar spindle assembly around chromatin beads. Depletion of TPX2 was partially rescued by the addition of excess …

Disruption Of Osysl15 Leads To Iron Inefficiency In Rice Plants, Elsbeth Walker, S. Lee, J. C. Chiecko, Y. Lee, M. L. Guerinot, G. An

Elsbeth Walker

Uptake and translocation of metal nutrients are essential processes for plant growth. Graminaceous species release phytosiderophores that bind to Fe3+; these complexes are then transported across the plasma membrane. We have characterized OsYSL15, one of the rice (Oryza sativa) YS1-like (YSL) genes that are strongly induced by iron (Fe) deficiency. The OsYSL15 promoter fusion to β-glucuronidase showed that it was expressed in all root tissues when Fe was limited. In low-Fe leaves, the promoter became active in all tissues except epidermal cells. This activity was also detected in flowers and seeds. The OsYSL15:green fluorescent protein fusion was localized to the …

Identification Of Thyroid Hormone Receptor Binding Sites And Target Genes Using Chip-On-Chip In Developing Mouse Cerebellum, H. Dong, C. L. Yauk, A. Rowan-Carroll, S. H. You, R. Thomas Zoeller, I Lambert, M. G. Wade

R. Thomas Zoeller

Thyroid hormone (TH) is critical to normal brain development, but the mechanisms operating in this process are poorly understood. We used chromatin immunoprecipitation to enrich regions of DNA bound to thyroid receptor beta (TRβ) of mouse cerebellum sampled on post natal day 15. Enriched target was hybridized to promoter microarrays (ChIP-on-chip) spanning −8 kb to +2 kb of the transcription start site (TSS) of 5000 genes. We identified 91 genes with TR binding sites. Roughly half of the sites were located in introns, while 30% were located within 1 kb upstream (5′) of the TSS. Of these genes, 83 with …

Rapid Screening For Temperature-Sensitive Alleles In Plants, L Vidali, Rc Augustine, Sn Fay, P Franco, Ka Pattavina, M Bezanilla

Magdalena Bezanilla

We developed a simple and fast method to identify temperature-sensitive alleles of essential plant genes. We used primary and tertiary structure information to identify residues in the core of the protein of interest. These residues were mutated and tested for temperature sensitivity, taking advantage of the exceptionally rapid 1-week complementation assay in the moss Physcomitrella patens. As test molecules, we selected the actin-binding proteins profilin and actin-depolymerizing factor, because they are essential and their loss-of-function phenotype can be fully rescued. Screening a small number of candidate mutants, we successfully identified temperature-sensitive alleles of both profilin and actin-depolymerizing factor. Plants harboring …

Lifeact-Megfp Reveals A Dynamic Apical F-Actin Network In Tip Growing Plant Cells, L Vidali, Cm Rounds, Pk Hepler, M Bezanilla

Magdalena Bezanilla

Background

Actin is essential for tip growth in plants. However, imaging actin in live plant cells has heretofore presented challenges. In previous studies, fluorescent probes derived from actin-binding proteins often alter growth, cause actin bundling and fail to resolve actin microfilaments.

Methodology/Principal Findings

In this report we use Lifeact-mEGFP, an actin probe that does not affect the dynamics of actin, to visualize actin in the moss Physcomitrella patens and pollen tubes from Lilium formosanum and Nicotiana tobaccum. Lifeact-mEGFP robustly labels actin microfilaments, particularly in the apex, in both moss protonemata and pollen tubes. Lifeact-mEGFP also labels filamentous actin structures in …

Investigation Of Gene Expression In C2c12 Myotubes Following Simvastatin Application And Mechanical Strain, Ji-Gu Yu, Kimberly Sewright, Monica J. Hubal, Jing-Xia Liu, Lawrence M. Schwartz, Eric P. Hoffman, Priscilla M. Clarkson

Lawrence M. Schwartz

Aim: The 3-hydroxy-3methylgutaryl coenzyme A (HMG-CoA) reductase inhibitors (statins) are the most effective prescribed drugs for lowering serum cholesterol; however, although statins are extremely safe medications and have brought significant benefits to patients with hypercholesterolemia, they have been shown to produce myalgia, cramps, exercise intolerance and fatigue. The aim of the study was to investigate the molecular mechanisms that may mediate statin myopathy. Methods: We used DNA microarray analysis to examine the changes in gene expression profiles induced by 1 hour and 6 hours of statin treatment on differentiated C2C12 myotubes. Four genes were selected for analysis at the protein …

Why Public Health Agencies Cannot Depend On Good Laboratory Practices As A Criterion For Selecting Data: The Case Of Bisphenol A, I Chahoud, F. S. Vom Saal, B. T. Akingbemi, S. M. Belcher, D. A. Crain, D. Crews, L. C. Guidice, P. A. Hunt, F. Farabollini, L. J. Guillette Jr., T. Hassold, S. M. Ho, K. Arizono, T. Colborn, T. Iguchi, S. Jobling, J. Kanno, H. Laufer, M. Marcus, A. Nadal, J. A. Mclachlan, J Oehlmann, N. Olea, P. Palanza, S. Parmigiani, B. S. Rubin, G. Schoenfelder, C. Sonnenschein, A. M. Soto, C. E. Talsness, J. A. Taylor, L. N. Vandenberg, J. G. Vandenbergh, S. Vogel, C. S. Watson, W. V. Welshons, R. Thomas Zoeller

R. Thomas Zoeller

Background In their safety evaluations of bisphenol A (BPA), the U.S. Food and Drug Administration (FDA) and a counterpart in Europe, the European Food Safety Authority (EFSA), have given special prominence to two industry-funded studies that adhered to standards defined by Good Laboratory Practices (GLP). These same agencies have given much less weight in risk assessments to a large number of independently replicated non-GLP studies conducted with government funding by the leading experts in various fields of science from around the world. Objectives We reviewed differences between industry-funded GLP studies of BPA conducted by commercial laboratories for regulatory purposes and …