Showing 41 - 50 of 257 Items
The role of adjuvant in mediating antigen structure and stability
Date: 2012-01-01
Creator: Latoya Jones Braun, Aimee M. Eldridge, Jessica Cummiskey, Kelly K. Arthur, Deborah S., Wuttke
Access: Open access
- The purpose of this study was to probe the fate of a model antigen, a cysteine-free mutant of bacteriophage T4 lysozyme, to the level of fine structural detail, as a consequence of its interaction with an aluminum (Al)-containing adjuvant. Fluorescence spectroscopy and differential scanning calorimetry were used to compare the thermal stability of the protein in solution versus adsorbed onto an Al-containing adjuvant. Differences in accessible hydrophobic surface areas were investigated using an extrinsic fluorescence probe, 8-Anilino-1-naphthalenesulfonic acid (ANS). As has been observed with other model antigens, the apparent thermal stability of the protein decreased following adsorption onto the adjuvant. ANS spectra suggested that adsorption onto the adjuvant caused an increase in exposure of hydrophobic regions of the protein. Electrostatic interactions drove the adsorption, and disruption of these interactions with high ionic strength buffers facilitated the collection of two-dimensional 15N heteronuclear single quantum coherence nuclear magnetic resonance data of protein released from the adjuvant. Although the altered stability of the adsorbed protein suggested changes to the protein's structure, the fine structure of the desorbed protein was nearly identical to the protein's structure in the adjuvant-free formulation. Thus, the adjuvant-induced changes to the protein that were responsible for the reduced thermal stability were not observed upon desorption. © 2011 Wiley Periodicals, Inc.
Arginine methylation of yeast mRNA-binding protein Npl3 directly affects its function, nuclear export, and intranuclear protein interactions
Date: 2005-09-02
Creator: Anne E. McBride, Jeffrey T. Cook, Elizabeth A. Stemmler, Kate L. Rutledge, Kelly A., McGrath, Jeffrey A. Rubens
Access: Open access
- Arginine methylation can affect both nucleocytoplasmic transport and protein-protein interactions of RNA-binding proteins. These effects are seen in cells that lack the yeast hnRNP methyltransferase (HMT1), raising the question of whether effects on specific proteins are direct or indirect. The presence of multiple arginines in individual methylated proteins also raises the question of whether overall methylation or methylation of a subset of arginines affects protein function. We have used the yeast mRNA-binding protein Npl3 to address these questions in vivo. Matrix-assisted laser desorption/ionization Fourier transform mass spectrometry was used to identify 17 methylated arginines in Npl3 purified from yeast: whereas 10 Arg-Gly-Gly (RGG) tripeptides were exclusively dimethylated, variable levels off methylation were found for 5 RGG and 2 RG motif arginines. We constructed a set of Npl3 proteins in which subsets of the RGG arginines were mutated to lysine. Expression of these mutant proteins as the sole form of Npl3 specifically affected growth of a strain that requires Hmtl. Although decreased growth generally correlated with increased numbers of Arg-to-Lys mutations, lysine substitutions in the N terminus of the RGG domain showed more severe effects. Npl3 with all 15 RGG arginines mutated to lysine exited the nucleus independent of Hmtl, indicating a direct effect of methylation on Npl3 transport. These mutations also resulted in a decreased, methylation-independent interaction of Npl3 with transcription elongation factor Tho2 and inhibited Npl3 self-association. These results support a model in which arginine methylation facilitates Npl3 export directly by weakening contacts with nuclear proteins. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc.
The Role of the Golgi ELMO Proteins in Cell Adhesion in Arabidopsis thaliana
Date: 2021-01-01
Creator: Wesley James Hudson
Access: Open access
- Proper growth and development of plant cells is dependent upon successful cell adhesion between cells, and this is mostly mediated by pectin in the plant cell wall. Previously, the Kohorn Laboratory identified a non-enzymatic Golgi protein named ELMO1 as it is required for cell adhesion, likely acting as a scaffold for cell wall polymer synthesis. Plants with mutant ELMO1 demonstrate a weak defective cellular adhesion phenotype as well as reduced mannose content in the cell wall. ELMO1 has homologous proteins in at least 29 different vascular plants. These homologues have 2 possible deletions in their amino acid sequence, but protein modeling determined that these variations will not affect protein structure. There are 5 homologous ELMO1 proteins in Arabidopsis thaliana that have been aptly named ELMO2, ELMO3, ELMO4, ELMO5. elmo2-/-mutants revealed no mutant adhesion phenotypes, while elmo1-/-elmo2-/-double mutants revealed strong defects in adhesion. Confocal microscopy of propidium iodide-stained seedlings confirmed the lack of a phenotype for elmo2-/-mutants and showed disorganized gapping cells for the elmo1-/-elmo2-/-mutant. Additionally, while elmo2-/-did not have any change to root or hypocotyl length, elmo1-/-elmo2-/- mutants were significantly shorter in both regards. Taken together, these data support that ELMO2 and ELMO1 are partially redundant.
The EOL Enhancer Activates Eya Expression to Mediate Visual System Development in Drosophila melanogaster This record is embargoed.
- Embargo End Date: 2027-05-16
Date: 2024-01-01
Creator: Benjamin Sewell-Grossman
Access: Embargoed
A histological investigation of Arceuthobium pusillum infections in Picea rubens and Picea glauca
Date: 2024-01-01
Creator: Sade K. McClean
Access: Open access
- Arceuthobium pusillum is a hemiparasite that infects select Picea species. The hosts of A. pusillum do not experience the same symptoms of infection. A. pusillum infections are more fatal to P. marinara, and P. glauca. P. rubens, on the other hand, can survive longer with sustained infection. This presents itself as a contemporary issue because P. glauca, one of the parasite’s most vulnerable hosts, was untethered from ecological competition when old growth forests were subjected to large scale anthropogenic disturbances. These disturbances allowed P. glauca to proliferate, with A. pusillum following. A deeper understanding of the host-species specific responses to A. pusillum infection can broaden general knowledge of parasitic growth and development while also potentially inspiring conservation techniques. This study took advantage of the intrinsic differences between host and parasite to visualize infections in P. rubens and P. glauca, highlighting differences in infection outcome. By illuminating lignin and callose within cross sections of infected P. rubens and P. glauca branches, it was revealed that P. rubens forms dense bands of cells around the cortical strands of infection. These bands form more frequently in P. rubens than in P. glauca and are of a significantly larger area in P. rubens than in P. glauca (t(8), p=0.003, p=0.005). The discovery of the exterior bands is novel and exciting, as the bands are possibly made of callose and potentially facilitate P. rubens survival against A. pusillum infection. The foundational discoveries and results of this study should inspire, and warrant, further analysis.
Mitigation of Negative Effects of Ocean Change on Oysters by Eelgrass and its Implications for Aquaculture in Midcoast Maine
Date: 2022-01-01
Creator: Fiona G Ralph
Access: Open access
- Species interactions are important to organisms and to the ecosystems they inhabit. These interactions, sometimes facilitations, can result in increased resiliency for both species. When facilitation occurs, organisms co-assist with physiological and environmental stressors. As anthropogenic impacts become more stressful for modern organisms, these interactions could offer a solution for many species. Ocean acidification has been shown to be detrimental to many calcifying organisms including oysters. More acidic conditions can slow the process of shell calcification, which can slow growth rates. This effect could directly impact the robust oyster farming business in Midcoast Maine. Because of its possible importance to oyster crops, we assessed the potential of Zostera marina, or eelgrass, to ameliorate the stresses of ocean acidification on farmed Eastern Oysters (Crassotrea virginica). Photosynthesizing organisms such as seagrasses have been shown to locally raise pH, which could create growth refugia for calcifying organisms. While eelgrass has the potential to enhance oyster growth rates, its meadows could also be influencing food availability. To better understand these dynamics, we grew C. virginica in two locations in Harpswell, ME. Crassostrea virginica were split into three habitats at each location: seagrass, fringe, and mudflat, and placed on surface or benthic arrays. We found that seagrass presence and depth interacted to increase shell growth rate. Similarly, Z. marina improved condition index of C. virginica. As ocean acidification worsens, oyster farmers might have to turn to mitigation strategies to ensure profit yield from their labors. Zostera marina could be the solution to their future problems.
A genomewide survey argues that every zygotic gene product is dispensable for the initiation of somatic homolog pairing in Drosophila
Date: 2008-11-01
Creator: Jack R. Bateman, C. Ting Wu
Access: Open access
- Studies from diverse organisms show that distinct interchromosomal interactions are associated with many developmental events. Despite recent advances in uncovering such phenomena, our understanding of how interchromosomal interactions are initiated and regulated is incomplete. During the maternal-to-zygotic transition (MZT) of Drosophila embryogenesis, stable interchromosomal contacts form between maternal and paternal homologous chromosomes, a phenomenon known as somatic homolog pairing. To better understand the events that initiate pairing, we performed a genomewide assessment of the zygotic contribution to this process. Specifically, we took advantage of the segregational properties of compound chromosomes to generate embryos lacking entire chromosome arms and, thus, all zygotic gene products derived from those arms. Using DNA fluorescence in situ hybridization (FISH) to assess the initiation of pairing at five separate loci, this approach allowed us to survey the entire zygotic genome using just a handful of crosses. Remarkably, we found no defect in pairing in embryos lacking any chromosome arm, indicating that no zygotic gene product is essential for pairing to initiate. From these data, we conclude that the initiation of pairing can occur independently of zygotic control and may therefore be part of the developmental program encoded by the maternal genome. Copyright © 2008 by the Genetics Society of America.
Site-specific transformation of Drosophila via φC31 integrase-mediated cassette exchange
Date: 2006-06-30
Creator: Jack R. Bateman, Anne M. Lee, C. Ting Wu
Access: Open access
- Position effects can complicate transgene analyses. This is especially true when comparing transgenes that have inserted randomly into different genomic positions and are therefore subject to varying position effects. Here, we introduce a method for the precise targeting of transgenic constructs to predetermined genomic sites in Drosophila using the φC31 integrase system in conjunction with recombinase-mediated cassette exchange (RMCE). We demonstrate the feasibility of this system using two donor cassettes, one carrying the yellow gene and the other carrying GFP. At all four genomic sites tested, we observed exchange of donor cassettes with an integrated target cassette carrying the mini-white gene. Furthermore, because RMCE-mediated integration of the donor cassette is necessarily accompanied by loss of the target cassette, we were able to identify integrants simply by the loss of mini-white eye color. Importantly, this feature of the technology will permit integration of unmarked constructs into Drosophila, even those lacking functional genes. Thus, φC31 integrase-mediated RMCE should greatly facilitate transgene analysis as well as permit new experimental designs. Copyright © 2006 by the Genetics Society of America.
Assembly of the precursor and processed light-harvesting chlorophyll a/b protein of Lemna into the light-harvesting complex II of barley etiochloroplasts.
Date: 1986-01-01
Creator: P. R. Chitnis, E. Harel, B. D. Kohorn, E. M. Tobin, J. P., Thornber
Access: Open access
- When the in vitro synthesized precursor of a light-harvesting chlorophyll a/b binding protein (LHCP) from Lemna gibba is imported into barley etiochloroplasts, it is processed to a single form. Both the processed form and the precursor are found in the thylakoid membranes, assembled into the light-harvesting complex of photosystem II. Neither form can be detected in the stromal fraction. The relative amounts of precursor and processed forms observed in the thylakoids are dependent on the developmental stage of the plastids used for uptake. The precursor as well as the processed form can also be detected in thylakoids of greening maize plastids used in similar uptake experiments. This detection of a precursor in the thylakoids, which has not been previously reported, could be a result of using rapidly developing plastids and/or using an heterologous system. Our results demonstrate that the extent of processing of LHCP precursor is not a prerequisite for its inclusion in the complex. They are also consistent with the possibility that the processing step can occur after insertion of the protein into the thylakoid membrane.
Levels of DNA polymorphism vary with mating system in the nematode genus Caenorhabditis
Date: 2002-01-01
Creator: Andrew Graustein, John M. Caspar, James R. Walters, Michael F. Palopoli
Access: Open access
- Self-fertilizing species often harbor less genetic variation than cross-fertilizing species, and at least four different models have been proposed to explain this trend. To investigate further the relationship between mating system and genetic variation, levels of DNA sequence polymorphism were compared among three closely related species in the genus Caenorhabditis: two self-fertilizing species, Caenorhabditis elegans and C. briggsae, and one cross-fertilizing species, C. remanei. As expected, estimates of silent site nucleotide diversity were lower in the two self-fertilizing species. For the mitochondrial genome, diversity in the selfing species averaged 42% of diversity in C. remanei. Interestingly, the reduction in genetic variation was much greater for the nuclear than for the mitochondrial genome. For two nuclear genes, diversity in the selfing species averaged 6 and 13% of diversity in C. remanei. We argue that either population bottlenecks or the repeated action of natural selection, coupled with high levels of selfing, are likely to explain the observed reductions in species-wide genetic diversity.