Investigating the Attraction of Caenorhabditis elegans to Bacillus thuringiensis Through Volatile Organic Compounds
In this study we aimed to investigate the mechanism by which Bacillus thuringiensis, a known Caenorhabditis elegans pathogen, attracts the worm. We hypothesized that B. thuringiensis produces volatile organic compounds that attract the worm. We hoped to identify these compounds by extracting them from B. thuringiensis using methanol and attempted to identify them using gas chromatographic and mass spectroscopic analysis. Results showed that in a chemotaxis assay a methanol extract from B. thuringiensis does indeed attract the worms. Increasing the concentration of extract by ten-fold showed an increase in attraction, given by an increase in mean chemotaxis index of 0.191 to 0.416, a statistically significant increase. Upon analyzing the extract using gas chromatography there were reproducible traces of volatile compounds in multiple methanol extracts.
1 Dept of Biology, Belmont University, Nashville, TN.
213 • KelviNeisha L. Williams1, Timothy Odom1, Teodora Best2, Nicole Zembower2, Ketia Shumaker1, John Carlson2
Comparative Analysis of Ozone Effects and Genomic Expression of Two Hardwood Trees
The goal of this study was a comparative analysis of ozone effects on two species of hardwood trees: Liriodendron tulipifera, L. (Tulip poplar) and Nyssa sylvatica, Marsh. (Black gum). The two species were exposed to four levels of ozone that best represented a conceivable spectrum of exposure (10, 80, 125, 225 ppb). In terms of foliar injury, Tulip poplar was more sensitive to the ozone effects than Black gum. From a comparative analysis of the transcriptoms of the two species, results are expected to show that the Black gum tree is more equipped to handle ozone stress than the Tulip poplar.
1 The University of West Alabama; 2 Pennsylvania State University
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214 • Sasha Gogoli, Renee J. Chosed
Modeling the MLL (Mixed-Lineage Leukemia) Complex in Saccharomyces Cerevisiae
Both yeast and humans have highly conservative SET domain containing methyltransferases that are part of large multi-protein complexes. These protein complexes are responsible for the methylation of Histone H3 on the fourth lysine residue (H3K4). This methylation allows for the regulation of HOX genes that initiate the growth of hematopoietic precursor cells from stem cells. Human MLL1 and Saccharomyces cerevisiae Set1 are the methyltransferase enzymes within these protein complexes, which are also composed of several other proteins. The non-catalytic accessory proteins of each complex are thought to play a role in regulating the methyltransferase activity of each complex, yet their roles are not well defined. Saccharomyces cerevisiae contain a single protein complex referred to as COMPASS that is homologous to that of the MLL1 protein complex in mammalian cells. To elucidate the roles of the accessory proteins in the mammalian MLL1 complex, we replaced proteins of COMPASS in yeast with those of the mammalian MLL1 complex. We then assayed for the activity of the protein complex by detecting histone H3K4 methylation by Western blotting. The results of our experiments showed that Set1 and Bre2 are both necessary for the proper function of COMPASS. Replacement of yeast Set1 with its human homologue MLL1, led to a partial rescue of the observed H3K4 methylation deficiency. Studies are ongoing to recover methylation by introduction of Ash2L into the yeast strain deficient in Bre2.
Dept of Biology, Furman University, Greenville, SC
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215 • Jacob Hanna, Renee J. Chosed
Ruthenium Complex KP1019 and Its Effects on Saccharomyces cerevisiae
The anticancer ruthenium complex trans-[tetrachlorobis(1H-indazole)ruthenate(III)], also known as KP1019, has been previously shown to induce DNA damage and cell death consistent with the formation of DNA interstrand cross-linkage. These conclusions were based off of the analysis of the hypersensitivity of yeast strains defective in DNA to treatment with KP1019. However, DNA interstrand cross-linkage does not explain KP1019’s ability to retain its toxicity against cancer cells resistant to such DNA damage. This suggests that addition KP1019 possesses additional cytotoxicity factors. Using Saccharomyces cerevisiae as a model organism we examined the alterations in histone modifications following treatment with KP1019. We utilized W303a, Δpdr1, and Δpdr3 S. cerevisiae strains. We found noticeable variation in multiple histone modifications including H3K4 tri-methylation and H4 acetylation. These data suggest that KP1019 may alter histone post-translational modifications in addition to forming DNA adducts.
Dept of Biology, Furman University, Greenville, SC
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216 • Rebekah L. Robinson1, Claudia Y. Alvarado1, Mario Johnson1, Scott C. Mateer1, Ron E. Garner2, Traci L. Ness1
Construction of a Single Nucleotide Polymorphism in the Human TLR4 Gene
Pattern recognition receptors (PRRs) are responsible for recognizing pathogens and initiating appropriate immune responses in many cell types. Toll-like receptor four (TLR4), a well-studied PRR, is the receptor for lipopolysaccharide (LPS) found in the outer membrane of Gram-negative bacteria. However, it has been demonstrated that the fungal polysaccharide mannan is also capable of binding and signaling through TLR4, but the details of this interaction are poorly understood. TLR4-D299G is a naturally-occurring polymorphism which has been shown to alter the response to LPS. Individuals with this polymorphism are highly susceptible to Gram-negative bacterial infections. Our lab is interested in determining if this polymorphism will also eliminate mannan binding and signaling through TLR4 in human cells. The goal of this project was to construct the TLR4-D299G polymorphism in the plasmid pUNO-hTLR04a using circular mutagenesis. A single nucleotide change (adenine → guanine) was introduced which changed the amino acid at site 299 from an aspartic acid (D) to a glycine (G) residue. The success of our mutagenesis strategy has been verified by antibiotic resistance of bacterial transformants, Pst I digestion of the isolated plasmids, and direct sequencing of the plasmid. Future studies will include transfection of the TLR4-D299G plasmid into a reporter cell line to test mannan’s ability to bind and activate this polymorphic form of the TLR4 receptor.
1 Dept of Biology, Armstrong Atlantic State University, Savannah, Georgia; 2 Mercer University School of Medicine, Savannah, Georgia
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217 • D’angeleau Newsome, Nick Ragsdale
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