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2015-2016

7 hits

  • Thumbnail for An Exploration of the Molecular Mechanisms of Behavior in Apteronotus leptorhynchus, or the Brown Ghost Knifefish
    An Exploration of the Molecular Mechanisms of Behavior in Apteronotus leptorhynchus, or the Brown Ghost Knifefish by Silver, Brianna Danielle

    Behavioral, molecular, and hormonal mechanisms work together to impact sensory processes, communication, and mate choice. This study investigates the evolution of communication and sexual dimorphism through an analysis of intrasexual variation in behavioral and molecular mechanisms in Apteronotus leptorhynchus, or the brown ghost knifefish. Although knifefish behavior is well described, gene expression studies in the brain are fairly novel, and few studies have looked at the correlation between behavior and gene expression in the brain within individuals. The brown ghost knifefish has unique, sexually dimorphic communicative behaviors which involve electric signals and can be systematically quantified by measuring electric organ discharge frequency (EODf) and chirp rates (rapid frequency modulations). In this study, we investigated the hypothesis that intrasexual variation in behavioral and molecular traits will be higher in males than in females due to sexual selection pressures, and that this variation is the result of changes in hormone receptor expression. To this end, we show that behavior is dimorphic in brown ghost knifefish, both in terms of EODf and chirp rates. Males have higher baseline frequencies and chirp rates than females. In addition, there is significantly more intrasexual variation in chirp rates within males. To determine if this sexually dimorphic behavior is due to changes in hormone receptor expression, we looked molecularly at expression of the androgen receptor, and of the estrogen receptors ESR1 and ESR2A. Preliminarily, our data suggest that perhaps receptor gene expression levels are not dimorphic in the brain, and therefore a different molecular mechanism seems to be driving this dimorphic behavior. Upon further investigation, we found that hormones, and specifically 11-ketotestosterone, seem to be likely candidates. This work supports the hypothesis that behavioral variation is greater in male knifefish, but the presence or absence of molecular dimorphism in the brain specifically could not be confirmed.

  • Thumbnail for Experimental and observational evidence that dissolved organic matter loading induces heterotrophy in a high elevation, oligotrophic lake
    Experimental and observational evidence that dissolved organic matter loading induces heterotrophy in a high elevation, oligotrophic lake by De La Rosa, Gabriel Tumonis

    Dissolved organic matter (DOM) is an important regulator of metabolism in aquatic ecosystems. The influx of DOM in oligotrophic, high alpine lakes during rain events has been shown to alter lake metabolism and increase net heterotrophy. DOM loading is expected to increase for high-elevation Sierra Nevada lakes as precipitation shifts from snow to rain. To explore the effects of DOM loading on ecosystem metabolism in high-elevation lakes, we characterize ecosystem metabolic response to DOM loading in both in situ incubation experiments and through whole lake measurements of dissolved oxygen made during a summer with unusually high levels precipitation. Experimental in situ incubations show net ecosystem production (NEP) decrease linearly as DOM increases, causing heterotrophy to grow stronger. Gross primary production (GPP) responded in a light-dependent manner, where GPP was stimulated across the DOM loading gradient during sunny, but not cloudy, weeks. ER was also stimulated and exceeded GPP across the entire DOM gradient, causing heterotrophy. ER was coupled to GPP in a light dependent manner, suggesting consumers continue to consume autochthonous carbon even when allochthonous carbon is present. Seasonal metabolism measurements support that DOM shifts a lake towards heterotrophy at the whole lake level, and a large rain event was sufficient to shift lake metabolism from autotrophy to heterotrophy for at least three weeks. Increased DOM loading due to climate induced changes in vegetation cover and the timing and magnitude of precipitation events could alter the amount of carbon cycled in these lakes.

  • Thumbnail for Experimental and observational evidence that dissolved organic matter loading induces heterotrophy in a high-elevation, oligotrophic lake
    Experimental and observational evidence that dissolved organic matter loading induces heterotrophy in a high-elevation, oligotrophic lake by De La Rosa, Gabriel Tumonis

    Dissolved organic matter (DOM) is an important regulator of metabolism in aquatic ecosystems. The influx of DOM in oligotrophic, high alpine lakes during rain events has been shown to alter lake metabolism and increase net heterotrophy. DOM loading is expected to increase for high-elevation Sierra Nevada lakes as precipitation shifts from snow to rain. To explore the effects of DOM loading on ecosystem metabolism in high-elevation lakes, we characterize ecosystem metabolic response to DOM loading in both in situ incubation experiments and through whole lake measurements of dissolved oxygen made during a summer with unusually high levels precipitation. Experimental in situ incubations show net ecosystem production (NEP) decrease linearly as DOM increases, causing heterotrophy to grow stronger. Gross primary production (GPP) responded in a light-dependent manner, where GPP was stimulated across the DOM loading gradient during sunny, but not cloudy, weeks. ER was also stimulated and exceeded GPP across the entire DOM gradient, causing heterotrophy. ER was coupled to GPP in a light dependent manner, suggesting consumers continue to consume autochthonous carbon even when allochthonous carbon is present. Seasonal metabolism measurements support that DOM shifts a lake towards heterotrophy at the whole lake level, and a large rain event was sufficient to shift lake metabolism from autotrophy to heterotrophy for at least three weeks. Increased DOM loading due to climate induced changes in vegetation cover and the timing and magnitude of precipitation events could alter the amount of carbon cycled in these lakes.

  • Thumbnail for Paralogs of nos-1 in Caenorhabditis elegans Exhibit Genetic Redundancy as Translational Regulators in Dendrite Morphogenesis
    Paralogs of nos-1 in Caenorhabditis elegans Exhibit Genetic Redundancy as Translational Regulators in Dendrite Morphogenesis by Barney, Julia Nicole

    Neurological disorders often present with defects in dendritic arborization, illustrating the significance of dendrites and synaptic connections for cognition. As such, it is important to the study of these disorders to understand the mechanism by which neurons establish proper dendritic morphology. Because dendrites are distantly located from the nucleus and exhibit independent changes in growth and connectivity, regulation of dendrite morphogenesis likely occurs through a localized post-transcriptional mechanism. RNA-binding proteins are important mediators of post-transcriptional regulation and several such proteins have been implicated in dendrite development. The RNA-binding protein, Nanos, has been previously characterized as important for dendrite morphogenesis in Drosophila. However, knockout of nos-1, the strongest nanos homolog in C. elegans, does not reveal significant dendrite defects. Due to the presence of two additional nos paralogs in the C. elegans genome, nos-2 and nos-3, we suspected that the lack of dendrite defects upon knockout of nos-1 indicated the possibility of genetic redundancy. Our results support the notion that the nos paralogs are genetically redundant and further identifies the redundant function of these proteins as important for fourth order dendrite morphogenesis in C. elegans. Furthermore, this study also identifies nos-3 alone as important for the morphogenesis of second and third order dendrites. The expression pattern of the three NOS proteins in the C. elegans PVD neuron is also consistent with the predicted function of NOS as a translational regulator, suggesting a role for the nos paralogs in dendrite development and maintenance.

  • Thumbnail for The Functional Significance of Type IV Pilus Proteins for Competence and Effects of Environmental Conditions on Competence in Acinetobacter baylyi
    The Functional Significance of Type IV Pilus Proteins for Competence and Effects of Environmental Conditions on Competence in Acinetobacter baylyi by Leong, Colleen Gong

    Acinetobacter baylyi are naturally competent soil bacteria. Natural transformation is the acquisition of new genetic material via the uptake of foreign, exogenous DNA. Competence is the physiological state some bacterial species may realize in order for natural transformation to occur. Natural transformation, and therefore competence, is clinically relevant, as natural transformation serves as a chief method by which antibiotic-resistant genes are dispersed amongst the bacterial population. A. baylyi serves as an ideal model organism to model natural transformation. A. baylyi are easy to cultivate in vitro, may be genetically modified with ease, and there is a complete library of single gene deletion mutants available for research use. Our first goal was to test the role of Type IV pilus (T4P) proteins in competence using a novel surface-associated quantitative protocol. From the French collection, we obtained knockout mutants lacking proteins predicted to be important for comprising a T4P or for uptake of DNA across the inner membrane. Transformation of cells on a nutritive agar surface allowed for quantitative determination of transformation efficiency over nine orders of magnitude. Using this method we determined which genes were necessary for competence. Under the conditions we tested, genes absolutely required for transformation in A. baylyi include genes encoding the basal apparatus of a T4P (comM, pilF, pilC, pilU, and pilT), the gene encoding the inner membrane DNA translocation protein (comA), the gene encoding the major pilin (comP), genes encoding minor pilins (pilV, fimT), and the gene encoding the pilus tip protein (comC). Mutations in genes encoding for a periplasmic protein that helps target DNA to the comA channel (comEA), conserved hypothetical protein (CHP), genes encoding for a signal transduction response and regulatory receiver (pilG, pilR, and pilS), and a gene encoding for a minor pilins, comF and comE, resulted in a 2-4 log loss in competence. Using transformation on a surface instead of in liquid, we have discovered that a T4P, including its major pilin, is required for transformation A. baylyi. Further, in order to determine conditions under which A. baylyi are most competent in LB media, a simple broth comprised of three ingredients in an easy ratio, we tested additives to LB broth. Though A. baylyi may be grown in a variety of different medias, our laboratory chooses to grow ADP1 cells in LB because of its ubiquity in bacterial labs, low cost, and high rates of natural transformation. Our experiments address whether we may increase the efficacy of LB by altering growing temperature or infusing it with a variety of experimental additives. Overall, we found that A. baylyi cells are competent in nearly all LB conditions tested, but notably, rates of transformation slightly increase in LB+succinate but drastically decrease in LBK+Fe-deficient.

  • Thumbnail for The Interaction Between Invariant Natural Killer T Cells and B Cells in Adipose Tissue
    The Interaction Between Invariant Natural Killer T Cells and B Cells in Adipose Tissue by Kumagai, Abigail Mieko

    Invariant natural killer T cells (iNKT) cells can play an anti-inflammatory role and secrete the cytokine interleukin 10 (IL-10). IL-10 acts to inhibit signaling from inflammatory cytokines. Loss of iNKT cells has been implicated in cancer, autoimmune diseases, diabetes, and obesity. During early responses to immune challenge, iNKT cells provide help to B cells via two main pathways. Direct help from iNKT cells to B cells follows an innate pathway, while indirect help from iNKT cells to B cells follows an adaptive pathway. The specific interaction between B cells and iNKT cells within adipose tissue is not understood. Here, the interaction between iNKT cells and B cells within white adipose tissue and brown adipose tissue was examined using αGalCer, a potent stimulator of iNKT cells. In both brown and white adipose tissue, αGalCer stimulation resulted in an expansion of tissue-resident, IL-10 producing iNKT cells. However, tissue-resident B cells did not significantly increase in number or IL-10 production. Instead, a population of B220+CD19-CD1d+ lymphocytes were found to have an iNKT-dependent expansion in response to αGalCer. Our data suggests that iNKT cells provide indirect, adaptive help to B cells within both brown and white adipose tissue. These findings have implications in possible treatments for diabetes and obesity, as activated iNKT cells could be used to reduce tissue inflammation.

  • Thumbnail for Toward imaging live, competent Acinetobacter baylyi with Liquid Cell Atomic Force Microscopy
    Toward imaging live, competent Acinetobacter baylyi with Liquid Cell Atomic Force Microscopy by Tenente, Ricardo

    Imaging live bacteria with an atomic force microscope (AFM) is a challenging process, taking into account the lateral forces exerted by the probe on cells that must be immobilized but still immersed in liquid. The Lang-Lostroh labs use AFM to examine Acinetobacter baylyi cells that are competent (able to take up DNA from the environment). In order to image cells while they are competent, it is necessary to find an effective combination of liquid media and sample preparation method while maintaining the cells alive, competent, and attached to the AFM pucks with porcine gelatin. Many combinations were tested using different washing and gelatin immobilization media (distilled deionized water, and phosphate-buffer saline), as well as using different media (distilled deionized water, phosphate-buffer saline, and diverse dilutions of Luria-Bertani broth with and without sodium chloride) during imaging itself. The combinations that presented the most satisfying images of immobilized cells with AFM were submitted to membrane integrity and competence assays. Washing and immobilizing the cells with distilled deionized water and imaging them with 50% Luria-Bertani broth without sodium chloride was the most successful combination. Even though the cells did not show detectable competence ability, they were alive yet inactive since after the imaging procedure they were still able to form colonies on plain Luria-Bertani agar plates. It provides insight on a viability spectrum that must be taken into account when distinguishing cell viability. Further research should focus on testing smoother media transitions with access to food source to avoid osmotic pressure stress and starvation.